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by Keyword: Tissue

Fischer NG, Aparicio C, (2022). Junctional epithelium and hemidesmosomes: Tape and rivets for solving the “percutaneous device dilemma” in dental and other permanent implants Bioactive Materials 18, 178-198

The percutaneous device dilemma describes etiological factors, centered around the disrupted epithelial tissue surrounding non-remodelable devices, that contribute to rampant percutaneous device infection. Natural percutaneous organs, in particular their extracellular matrix mediating the “device”/epithelium interface, serve as exquisite examples to inspire longer lasting long-term percutaneous device design. For example, the tooth's imperviousness to infection is mediated by the epithelium directly surrounding it, the junctional epithelium (JE). The hallmark feature of JE is formation of hemidesmosomes, cell/matrix adhesive structures that attach surrounding oral gingiva to the tooth's enamel through a basement membrane. Here, the authors survey the multifaceted functions of the JE, emphasizing the role of the matrix, with a particular focus on hemidesmosomes and their five main components. The authors highlight the known (and unknown) effects dental implant – as a model percutaneous device – placement has on JE regeneration and synthesize this information for application to other percutaneous devices. The authors conclude with a summary of bioengineering strategies aimed at solving the percutaneous device dilemma and invigorating greater collaboration between clinicians, bioengineers, and matrix biologists. © 2022 The Authors

JTD Keywords: amino-acid-sequence, bioinspired surfaces, cell-secreted protein, growth-factor receptor, hemidesmosome, integrin beta-4 subunit, junctional epithelium, keratinocyte-derived chemokine, laminin-binding integrins, marginal bone loss, percutaneous implant, pressure wound therapy, soft-tissue integration, Bioinspired surfaces, Bullous-pemphigoid antigen, Hemidesmosome, Junctional epithelium, Percutaneous device, Percutaneous implant


Martínez-Miguel M, Castellote-Borrell M, Köber M, Kyvik AR, Tomsen-Melero J, Vargas-Nadal G, Muñoz J, Pulido D, Cristóbal-Lecina E, Passemard S, Royo M, Mas-Torrent M, Veciana J, Giannotti MI, Guasch J, Ventosa N, Ratera I, (2022). Hierarchical Quatsome-RGD Nanoarchitectonic Surfaces for Enhanced Integrin-Mediated Cell Adhesion Acs Applied Materials & Interfaces 14, 48179-48193

The synthesis and study of the tripeptide Arg-Gly-Asp (RGD), the binding site of different extracellular matrix proteins, e.g., fibronectin and vitronectin, has allowed the production of a wide range of cell adhesive surfaces. Although the surface density and spacing of the RGD peptide at the nanoscale have already shown a significant influence on cell adhesion, the impact of its hierarchical nanostructure is still rather unexplored. Accordingly, a versatile colloidal system named quatsomes, based on fluid nanovesicles formed by the self-assembling of cholesterol and surfactant molecules, has been devised as a novel template to achieve hierarchical nanostructures of the RGD peptide. To this end, RGD was anchored on the vesicle's fluid membrane of quatsomes, and the RGD-functionalized nanovesicles were covalently anchored to planar gold surfaces, forming a state of quasi-suspension, through a long poly(ethylene glycol) (PEG) chain with a thiol termination. An underlying self-assembled monolayer (SAM) of a shorter PEG was introduced for vesicle stabilization and to avoid unspecific cell adhesion. In comparison with substrates featuring a homogeneous distribution of RGD peptides, the resulting hierarchical nanoarchitectonic dramatically enhanced cell adhesion, despite lower overall RGD molecules on the surface. The new versatile platform was thoroughly characterized using a multitechnique approach, proving its enhanced performance. These findings open new methods for the hierarchical immobilization of biomolecules on surfaces using quatsomes as a robust and novel tissue engineering strategy.

JTD Keywords: activation, arg-gly-asp (rgd), cell adhesion, extracellular-matrix, growth, integrins, ligands, nanopatterns, quatsomes, scaffolds, self-assembled monolayers, surface engineering, tissue engineering, Arg-gly-asp (rgd), Cell adhesion, Integrins, Nano-structured surfaces, Nanovesicles, Quatsomes, Self-assembled monolayers, Surface engineering, Tissue engineering


López-Canosa, Adrián, Pérez-Amodio, Soledad, Engel, Elisabeth, Castaño, Oscar, (2022). Microfluidic 3D Platform to Evaluate Endothelial Progenitor Cell Recruitment by Bioactive Materials Acta Biomaterialia 151, 264-277

Fernández-Garibay, Xiomara, Gomez-Florit, Manuel, Domingues, Rui M A, Gomes, Manuela, Fernandez-Costa, Juan M., Ramon, Javier, (2022). Xeno-free bioengineered human skeletal muscle tissue using human platelet lysate-based hydrogels Biofabrication 14, 045015

Abstract Bioengineered human skeletal muscle tissues have emerged in the last years as new in vitro systems for disease modeling. These bioartificial muscles are classically fabricated by encapsulating human myogenic precursor cells in a hydrogel scaffold that resembles the extracellular matrix. However, most of these hydrogels are derived from xenogenic sources, and the culture media is supplemented with animal serum, which could interfere in drug testing assays. On the contrary, xeno-free biomaterials and culture conditions in tissue engineering offer increased relevance for developing human disease models. In this work, we used human platelet lysate-based nanocomposite hydrogels (HUgel) as scaffolds for human skeletal muscle tissue engineering. These hydrogels consist of human platelet lysate reinforced with cellulose nanocrystals (a-CNC) that allow tunable mechanical, structural, and biochemical properties for the 3D culture of stem cells. Here, we developed hydrogel casting platforms to encapsulate human muscle satellite stem cells in HUgel. The a-CNC content was modulated to enhance matrix remodeling, uniaxial tension, and self-organization of the cells, resulting in the formation of highly aligned, long myotubes expressing sarcomeric proteins. Moreover, the bioengineered human muscles were subjected to electrical stimulation, and the exerted contractile forces were measured in a non-invasive manner. Overall, our results demonstrated that the bioengineered human skeletal muscles could be built in xeno-free cell culture platforms to assess tissue functionality, which is promising for drug development applications.

JTD Keywords: 3d culture, generation, identification, image, manipulate, matrigel, mechanics, model, platelet lysate, scaffolds, tissue engineering, xeno-free, Platform, Skeletal muscle


Cable, J, Arlotta, P, Parker, KK, Hughes, AJ, Goodwin, K, Mummery, CL, Kamm, RD, Engle, SJ, Tagle, DA, Boj, SF, Stanton, AE, Morishita, Y, Kemp, ML, Norfleet, DA, May, EE, Lu, A, Bashir, R, Feinberg, AW, Hull, SM, Gonzalez, AL, Blatchley, MR, Pulido, NM, Morizane, R, McDevitt, TC, Mishra, D, Mulero-Russe, A, (2022). Engineering multicellular living systems-A Keystone Symposia report Annals Of The New York Academy Of Sciences , 1-13

The ability to engineer complex multicellular systems has enormous potential to inform our understanding of biological processes and disease and alter the drug development process. Engineering living systems to emulate natural processes or to incorporate new functions relies on a detailed understanding of the biochemical, mechanical, and other cues between cells and between cells and their environment that result in the coordinated action of multicellular systems. On April 3-6, 2022, experts in the field met at the Keystone symposium "Engineering Multicellular Living Systems" to discuss recent advances in understanding how cells cooperate within a multicellular system, as well as recent efforts to engineer systems like organ-on-a-chip models, biological robots, and organoids. Given the similarities and common themes, this meeting was held in conjunction with the symposium "Organoids as Tools for Fundamental Discovery and Translation".

JTD Keywords: Brain organoids, Cell diversity, Computational, Dynamics, Engineered living, Engineered organs, Heart, Maturation, Model, Multicellular, Mycobacterium-tuberculosis, Quantitative-analysis, Systems, Tissue deformation


Larrañaga, Enara, Fernández‐Majada, Vanesa, Ojosnegros, Samuel, Comelles, Jordi, Martinez, Elena, (2022). Ephrin Micropatterns Exogenously Modulate Cell Organization in Organoid‐Derived Intestinal Epithelial Monolayers Advanced Materials Interfaces , 2201301

Fernández‐Costa, Juan M., Ortega, María A., Rodríguez‐Comas, Júlia, Lopez‐Muñoz, Gerardo, Yeste, Jose, Mangas‐Florencio, Lluís, Fernández‐González, Miriam, Martin‐Lasierra, Eduard, Tejedera‐Villafranca, Ainoa, Ramon‐Azcon, Javier, (2022). Training-on-a-Chip: A MultiOrgan Device to Study the Effect of Muscle Exercise on Insulin Secretion in Vitro Advanced Materials Technologies , 2200873

Ordoño J, Pérez-Amodio S, Ball K, Aguirre A, Engel E, (2022). The generation of a lactate-rich environment stimulates cell cycle progression and modulates gene expression on neonatal and hiPSC-derived cardiomyocytes Biomaterials Advances 139, 213035

In situ tissue engineering strategies are a promising approach to activate the endogenous regenerative potential of the cardiac tissue helping the heart to heal itself after an injury. However, the current use of complex reprogramming vectors for the activation of reparative pathways challenges the easy translation of these therapies into the clinic. Here, we evaluated the response of mouse neonatal and human induced pluripotent stem cell-derived cardiomyocytes to the presence of exogenous lactate, thus mimicking the metabolic environment of the fetal heart. An increase in cardiomyocyte cell cycle activity was observed in the presence of lactate, as determined through Ki67 and Aurora-B kinase. Gene expression and RNA-sequencing data revealed that cardiomyocytes incubated with lactate showed upregulation of BMP10, LIN28 or TCIM in tandem with downregulation of GRIK1 or DGKK among others. Lactate also demonstrated a capability to modulate the production of inflammatory cytokines on cardiac fibroblasts, reducing the production of Fas, Fraktalkine or IL-12p40, while stimulating IL-13 and SDF1a. In addition, the generation of a lactate-rich environment improved ex vivo neonatal heart culture, by affecting the contractile activity and sarcomeric structures and inhibiting epicardial cell spreading. Our results also suggested a common link between the effect of lactate and the activation of hypoxia signaling pathways. These findings support a novel use of lactate in cardiac tissue engineering, modulating the metabolic environment of the heart and thus paving the way to the development of lactate-releasing platforms for in situ cardiac regeneration.Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.

JTD Keywords: cardiac regeneration, cardiac tissue engineering, cell cycle, failure, growth, heart regeneration, induced pluripotent stem cells, ischemia, lactate, metabolic environment, metabolism, mouse, proliferation, repair, Bone morphogenetic protein-10, Cardiac tissue engineering, Cardiomyocytes, Cell cycle, Induced pluripotent stem cells, Lactate, Metabolic environment


Duch, P, Diaz-Valdivia, N, Ikemori, R, Gabasa, M, Radisky, ES, Arshakyan, M, Gea-Sorli, S, Mateu-Bosch, A, Bragado, P, Carrasco, JL, Mori, H, Ramirez, J, Teixido, C, Reguart, N, Fillat, C, Radisky, DC, Alcaraz, J, (2022). Aberrant TIMP-1 overexpression in tumor-associated fibroblasts drives tumor progression through CD63 in lung adenocarcinoma Matrix Biology 111, 207-225

Tissue inhibitor of metalloproteinase-1 (TIMP-1) is an important regulator of extracellular matrix turnover that has been traditionally regarded as a potential tumor suppressor owing to its inhibitory effects of matrix metal-loproteinases. Intriguingly, this interpretation has been challenged by the consistent observation that increased expression of TIMP-1 is associated with poor prognosis in virtually all cancer types including lung cancer, supporting a tumor-promoting function. However, how TIMP-1 is dysregulated within the tumor micro-environment and how it drives tumor progression in lung cancer is poorly understood. We analyzed the expression of TIMP-1 and its cell surface receptor CD63 in two major lung cancer subtypes: lung adenocarci-noma (ADC) and squamous cell carcinoma (SCC), and defined the tumor-promoting effects of their interac-tion. We found that TIMP-1 is aberrantly overexpressed in tumor-associated fibroblasts (TAFs) in ADC compared to SCC. Mechanistically, TIMP-1 overexpression was mediated by the selective hyperactivity of the pro-fibrotic TGF-61/SMAD3 pathway in ADC-TAFs. Likewise, CD63 was upregulated in ADC compared to SCC cells. Genetic analyses revealed that TIMP-1 secreted by TGF-61-activated ADC-TAFs is both nec-essary and sufficient to enhance growth and invasion of ADC cancer cells in culture, and that tumor cell expression of CD63 was required for these effects. Consistently, in vivo analyses revealed that ADC cells co-injected with fibroblasts with reduced SMAD3 or TIMP-1 expression into immunocompromised mice attenu-ated tumor aggressiveness compared to tumors bearing parental fibroblasts. We also found that high TIMP1 and CD63 mRNA levels combined define a stronger prognostic biomarker than TIMP1 alone. Our results identify an excessive stromal TIMP-1 within the tumor microenvironment selectively in lung ADC, and implicate it in a novel tumor-promoting TAF-carcinoma crosstalk, thereby pointing to TIMP-1/CD63 interaction as a novel therapeutic target in lung cancer. (c) 2022 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)

JTD Keywords: Angiogenesis, Cancer cells, Cancer-associated fibroblast, Cd63, Expression, Fibrosis, Hepatocellular-carcinoma, Metalloproteinases, Nintedanib, Prognostic-significance, Protein, Smad3, Squamous-cell carcinoma, Tgf-? 1, Tgf-β1, Timp-1, Tissue inhibitor, Tumor microenvironment


Blanco-Fernandez, B, Rey-Vinolas, S, Bagci, G, Rubi-Sans, G, Otero, J, Navajas, D, Perez-Amodio, S, Engel, E, (2022). Bioprinting Decellularized Breast Tissue for the Development of Three-Dimensional Breast Cancer Models Acs Applied Materials & Interfaces 14, 29467-29482

The tumor extracellular matrix (ECM) plays a vital role in tumor progression and drug resistance. Previous studies have shown that breast tissue-derived matrices could be an important biomaterial to recreate the complexity of the tumor ECM. We have developed a method for decellularizing and delipidating a porcine breast tissue (TDM) compatible with hydrogel formation. The addition of gelatin methacrylamide and alginate allows this TDM to be bioprinted by itself with good printability, shape fidelity, and cytocompatibility. Furthermore, this bioink has been tuned to more closely recreate the breast tumor by incorporating collagen type I (Col1). Breast cancer cells (BCCs) proliferate in both TDM bioinks forming cell clusters and spheroids. The addition of Col1 improves the printability of the bioink as well as increases BCC proliferation and reduces doxorubicin sensitivity due to a downregulation of HSP90. TDM bioinks also allow a precise three-dimensional printing of scaffolds containing BCCs and stromal cells and could be used to fabricate artificial tumors. Taken together, we have proven that these novel bioinks are good candidates for biofabricating breast cancer models.

JTD Keywords: 3d in vitro cancer model, Bioink, Bioprinting, Breast tissue, Crosstalk, Decellularization, Extracellular-matrix, Growth, Hydrogels, In-vitro, Inhibition, Mechanical-properties, Metastasis, Proliferation


Clua-Ferre, L, De Chiara, F, Rodriguez-Comas, J, Comelles, J, Martinez, E, Godeau, AL, Garcia-Alaman, A, Gasa, R, Ramon-Azcon, J, (2022). Collagen-Tannic Acid Spheroids for beta-Cell Encapsulation Fabricated Using a 3D Bioprinter Advanced Materials Technologies 7, 2101696

Chulia-Peris, L, Carreres-Rey, C, Gabasa, M, Alcaraz, J, Carretero, J, Pereda, J, (2022). Matrix Metalloproteinases and Their Inhibitors in Pulmonary Fibrosis: EMMPRIN/CD147 Comes into Play International Journal Of Molecular Sciences 23, 6894

Pulmonary fibrosis (PF) is characterized by aberrant extracellular matrix (ECM) deposition, activation of fibroblasts to myofibroblasts and parenchymal disorganization, which have an impact on the biomechanical traits of the lung. In this context, the balance between matrix metalloproteinases (MMPs) and their tissue inhibitors of metalloproteinases (TIMPs) is lost. Interestingly, several MMPs are overexpressed during PF and exhibit a clear profibrotic role (MMP-2, -3, -8, -11, -12 and -28), but a few are antifibrotic (MMP-19), have both profibrotic and antifibrotic capacity (MMP7), or execute an unclear (MMP-1, -9, -10, -13, -14) or unknown function. TIMPs are also overexpressed in PF; hence, the modulation and function of MMPs and TIMP are more complex than expected. EMMPRIN/CD147 (also known as basigin) is a transmembrane glycoprotein from the immunoglobulin superfamily (IgSF) that was first described to induce MMP activity in fibroblasts. It also interacts with other molecules to execute non-related MMP aactions well-described in cancer progression, migration, and invasion. Emerging evidence strongly suggests that CD147 plays a key role in PF not only by MMP induction but also by stimulating fibroblast myofibroblast transition. In this review, we study the structure and function of MMPs, TIMPs and CD147 in PF and their complex crosstalk between them.

JTD Keywords: Basigin, Cd147, Cell-surface, Emmprin, Extracellular-matrix, Gelatinase-b, Gene-expression profiles, Growth-factor-beta, Immunoglobulin superfamily, Induced lung injury, Inducer emmprin, Mmps, Pulmonary fibrosis, Timps, Tissue inhibitor, Transforming growth-factor-beta-1


Kim, YH, Dawson, JI, Oreffo, ROC, Tabata, Y, Kumar, D, Aparicio, C, Mutreja, I, (2022). Gelatin Methacryloyl Hydrogels for Musculoskeletal Tissue Regeneration Bioengineering (Basel) 9, 332-332

Musculoskeletal disorders are a significant burden on the global economy and public health. Hydrogels have significant potential for enhancing the repair of damaged and injured musculoskeletal tissues as cell or drug delivery systems. Hydrogels have unique physicochemical properties which make them promising platforms for controlling cell functions. Gelatin methacryloyl (GelMA) hydrogel in particular has been extensively investigated as a promising biomaterial due to its tuneable and beneficial properties and has been widely used in different biomedical applications. In this review, a detailed overview of GelMA synthesis, hydrogel design and applications in regenerative medicine is provided. After summarising recent progress in hydrogels more broadly, we highlight recent advances of GelMA hydrogels in the emerging fields of musculoskeletal drug delivery, involving therapeutic drugs (e.g., growth factors, antimicrobial molecules, immunomodulatory drugs and cells), delivery approaches (e.g., single-, dual-release system), and material design (e.g., addition of organic or inorganic materials, 3D printing). The review concludes with future perspectives and associated challenges for developing local drug delivery for musculoskeletal applications.

JTD Keywords: Drug delivery, Gelatin, Gelma, Hydrogel, Musculoskeletal tissue


Rosales-Rojas, R, Zuniga-Bustos, M, Salas-Sepulveda, F, Galaz-Araya, C, Zamora, RA, Poblete, H, (2022). Self-Organization Dynamics of Collagen-like Peptides Crosslinking Is Driven by Rose-Bengal-Mediated Electrostatic Bridges Pharmaceutics 14, 1148

The present work focuses on the computational study of the structural micro-organization of hydrogels based on collagen-like peptides (CLPs) in complex with Rose Bengal (RB). In previous studies, these hydrogels computationally and experimentally demonstrated that when RB was activated by green light, it could generate forms of stable crosslinked structures capable of regenerating biological tissues such as the skin and cornea. Here, we focus on the structural and atomic interactions of two collagen-like peptides (collagen-like peptide I (CLPI), and collagen-like peptide II, (CLPII)) in the presence and absence of RB, highlighting the acquired three-dimensional organization and going deep into the stabilization effect caused by the dye. Our results suggest that the dye could generate a ternary ground-state complex between collagen-like peptide fibers, specifically with positively charged amino acids (Lys in CLPI and Arg in CLPII), thus stabilizing ordered three-dimensional structures. The discoveries generated in this study provide the structural and atomic bases for the subsequent rational development of new synthetic peptides with improved characteristics for applications in the regeneration of biological tissues during photochemical tissue bonding therapies.

JTD Keywords: Anastomosis, Collagen-like peptide, Crosslinking, Green light, Mm simulations, Molecular dynamics, Molecular-dynamics, Photochemical tissue bonding therapies, Qm, Rose bengal


Raymond, Y, Lehmann, C, Thorel, E, Benitez, R, Riveiro, A, Pou, J, Manzanares, MC, Franch, J, Canal, C, Ginebra, MP, (2022). 3D printing with star-shaped strands: A new approach to enhance in vivo bone regeneration Biomaterials Advances 137, 212807

Concave surfaces have shown to promote bone regeneration in vivo. However, bone scaffolds obtained by direct ink writing, one of the most promising approaches for the fabrication of personalized bone grafts, consist mostly of convex surfaces, since they are obtained by microextrusion of cylindrical strands. By modifying the geometry of the nozzle, it is possible to print 3D structures composed of non-cylindrical strands and favor the presence of concave surfaces. In this work, we compare the in vivo performance of 3D-printed calcium phosphate scaffolds with either conventional cylindrical strands or star-shaped strands, in a rabbit femoral condyle model. Mono cortical defects, drilled in contralateral positions, are randomly grafted with the two scaffold configurations, with identical composition. The samples are explanted eight weeks post-surgery and assessed by ??-CT and resin embedded histological observations. The results reveal that the scaffolds containing star-shaped strands have better osteoconductive properties, guiding the newly formed bone faster towards the core of the scaffolds, and enhance bone regeneration, although the increase is not statistically significant (p > 0.05). This new approach represents a turning point towards the optimization of pore shape in 3D-printed bone grafts, further boosting the possibilities that direct ink writing technology offers for patient-specific applications.

JTD Keywords: 3d printing, Architecture, Biomimetic calcium phosphate, Bone regeneration, Calcium-phosphate scaffolds, Geometry, Growth, Implants, In vivo, Induction, Microporosity, Osteoinduction, Pore architecture, Scaffold, Surfaces, Tissue


Arque, X, Torres, MDT, Patino, T, Boaro, A, Sanchez, S, de la Fuente-Nunez, C, (2022). Autonomous Treatment of Bacterial Infections in Vivo Using Antimicrobial Micro- and Nanomotors Acs Nano 16, 7547-7558

The increasing resistance of bacteria to existing antibiotics constitutes a major public health threat globally. Most current antibiotic treatments are hindered by poor delivery to the infection site, leading to undesired off-target effects and drug resistance development and spread. Here, we describe micro- and nanomotors that effectively and autonomously deliver antibiotic payloads to the target area. The active motion and antimicrobial activity of the silica-based robots are driven by catalysis of the enzyme urease and antimicrobial peptides, respectively. These antimicrobial motors show micromolar bactericidal activity in vitro against different Gram-positive and Gram-negative pathogenic bacterial strains and act by rapidly depolarizing their membrane. Finally, they demonstrated autonomous anti-infective efficacy in vivo in a clinically relevant abscess infection mouse model. In summary, our motors combine navigation, catalytic conversion, and bactericidal capacity to deliver antimicrobial payloads to specific infection sites. This technology represents a much-needed tool to direct therapeutics to their target to help combat drug-resistant infections.

JTD Keywords: antibiotic-resistance, antimicrobial peptides, autonomous treatment, bacterial infection, delivery, ll-37, nanoparticles, peptide, self-propulsion, tissue, vitro, wasp venom, Antibiotic-resistance, Antimicrobial peptides, Autonomous treatment, Bacterial infection, Delivery, Ll-37, Mesoporous silica nanoparticles, Nanomotors, Nanoparticles, Peptide, Self-propulsion, Tissue, Vitro, Wasp venom


De Chiara, Francesco, Ferret-Miñana, Ainhoa, Fernández-Costa, Juan M., Senni, Alice, Jalan, Rajiv, Ramón-Azcón, Javier, (2022). Fatty Hepatocytes Induce Skeletal Muscle Atrophy In Vitro: A New 3D Platform to Study the Protective Effect of Albumin in Non-Alcoholic Fatty Liver Biomedicines 10, 958

The liver neutralizes endogenous and exogenous toxins and metabolites, being metabolically interconnected with many organs. Numerous clinical and experimental studies show a strong association between Non-alcoholic fatty liver disease (NAFLD) and loss of skeletal muscle mass known as sarcopenia. Liver transplantation solves the hepatic-related insufficiencies, but it is unable to revert sarcopenia. Knowing the mechanism(s) by which different organs communicate with each other is crucial to improve the drug development that still relies on the two-dimensional models. However, those models fail to mimic the pathological features of the disease. Here, both liver and skeletal muscle cells were encapsulated in gelatin methacryloyl and carboxymethylcellulose to recreate the disease’s phenotype in vitro. The 3D hepatocytes were challenged with non-esterified fatty acids (NEFAs) inducing features of Non-alcoholic fatty liver (NAFL) such as lipid accumulation, metabolic activity impairment and apoptosis. The 3D skeletal muscle tissues incubated with supernatant from fatty hepatocytes displayed loss of maturation and atrophy. This study demonstrates the connection between the liver and the skeletal muscle in NAFL, narrowing down the players for potential treatments. The tool herein presented was employed as a customizable 3D in vitro platform to assess the protective effect of albumin on both hepatocytes and myotubes.

JTD Keywords: 3r, ammonia, cirrhosis, disease, expression, myostatin, nefas, sarcopenia, tissue engineering, Crosstalk, Nuclear factor 4-alpha


Karkali, K, Tiwari, P, Singh, A, Tlili, S, Jorba, I, Navajas, D, Munoz, JJ, Saunders, TE, Martin-Blanco, E, (2022). Condensation of the Drosophila nerve cord is oscillatory and depends on coordinated mechanical interactions Developmental Cell 57, 867-882

During development, organs reach precise shapes and sizes. Organ morphology is not always obtained through growth; a classic counterexample is the condensation of the nervous system during Drosophila embryogenesis. The mechanics underlying such condensation remain poorly understood. Here, we characterize the condensation of the embryonic ventral nerve cord (VNC) at both subcellular and tissue scales. This analysis reveals that condensation is not a unidirectional continuous process but instead occurs through oscillatory contractions. The VNC mechanical properties spatially and temporally vary, and forces along its longitudinal axis are spatially heterogeneous. We demonstrate that the process of VNC condensation is dependent on the coordinated mechanical activities of neurons and glia. These outcomes are consistent with a viscoelastic model of condensation, which incorporates time delays and effective frictional interactions. In summary, we have defined the progressive mechanics driving VNC condensation, providing insights into how a highly viscous tissue can autonomously change shape and size.

JTD Keywords: Collagen-iv, Contraction, Forces, Gene, Glial-cells, Migration, Morphogenesis, Quantification, System, Tissue


Blanco-Fernandez G, Blanco-Fernandez B, Fernández-Ferreiro A, Otero-Espinar F, (2022). Bringing lipidic lyotropic liquid crystal technology into biomedicine Trends In Pharmacological Sciences 2022

Liquid crystals (LCs), discovered more than 130 years ago, are now emerging in the field of biomedicine. This article highlights the recent uses of lipid lyotropic LCs in therapeutics delivery, imaging, and tissue engineering and invites the scientific community to continue exploring the design of more complex LCs. © 2022 Elsevier Ltd

JTD Keywords: Biomedicine, Drug delivery, Glycerol monooleate, Imaging, tissue engineering, Lyotropic liquid crystals


Narciso M, Ulldemolins A, Júnior C, Otero J, Navajas D, Farré R, Gavara N, Almendros I, (2022). Novel Decellularization Method for Tissue Slices Frontiers In Bioengineering And Biotechnology 10, 832178

Decellularization procedures have been developed and optimized for the entire organ or tissue blocks, by either perfusion of decellularizing agents through the tissue’s vasculature or submerging large sections in decellularizing solutions. However, some research aims require the analysis of native as well as decellularized tissue slices side by side, but an optimal protocol has not yet been established to address this need. Thus, the main goal of this work was to develop a fast and efficient decellularization method for tissue slices—with an emphasis on lung—while attached to a glass slide. To this end, different decellularizing agents were compared for their effectiveness in cellular removal while preserving the extracellular matrix. The intensity of DNA staining was taken as an indicator of remaining cells and compared to untreated sections. The presence of collagen, elastin and laminin were quantified using immunostaining and signal quantification. Scaffolds resulting from the optimized protocol were mechanically characterized using atomic force microscopy. Lung scaffolds were recellularized with mesenchymal stromal cells to assess their biocompatibility. Some decellularization agents (CHAPS, triton, and ammonia hydroxide) did not achieve sufficient cell removal. Sodium dodecyl sulfate (SDS) was effective in cell removal (1% remaining DNA signal), but its sharp reduction of elastin signal (only 6% remained) plus lower attachment ratio (32%) singled out sodium deoxycholate (SD) as the optimal treatment for this application (6.5% remaining DNA signal), due to its higher elastin retention (34%) and higher attachment ratio (60%). Laminin and collagen were fully preserved in all treatments. The SD decellularization protocol was also successful for porcine and murine (mice and rat) lungs as well as for other tissues such as the heart, kidney, and bladder. No significant mechanical differences were found before and after sample decellularization. The resulting acellular lung scaffolds were shown to be biocompatible (98% cell survival after 72 h of culture). This novel method to decellularize tissue slices opens up new methodological possibilities to better understand the role of the extracellular matrix in the context of several diseases as well as tissue engineering research and can be easily adapted for scarce samples like clinical biopsies. Copyright © 2022 Narciso, Ulldemolins, Júnior, Otero, Navajas, Farré, Gavara and Almendros.

JTD Keywords: biocompatibility, bioscaffold recellularization, extracellular matrix, flow, impact, lung, scaffolds, tissue slices, Ammonia, Bio-scaffolds, Biocompatibility, Biological organs, Bioscaffold recellularization, Cell removal, Cells, Collagen, Cytology, Decellularization, Dna, Dna signals, Elastin, Extracellular matrices, Extracellular matrix, Extracellular-matrix, Glycoproteins, Laminin, Lung, Mammals, Recellularization, Scaffolds (biology), Sodium deoxycholate, Sulfur compounds, Tissue, Tissue slice, Tissue slices


Aydin, Onur, Passaro, Austin P., Raman, Ritu, Spellicy, Samantha E., Weinberg, Robert P., Kamm, Roger D., Sample, Matthew, Truskey, George A., Zartman, Jeremiah, Dar, Roy D., Palacios, Sebastian, Wang, Jason, Tordoff, Jesse, Montserrat, Nuria, Bashir, Rashid, Saif, MTaher A., Weiss, Ron, (2022). Principles for the design of multicellular engineered living systems Apl Bioengineering 6, 10903

Remarkable progress in bioengineering over the past two decades has enabled the formulation of fundamental design principles for a variety of medical and non-medical applications. These advancements have laid the foundation for building multicellular engineered living systems (M-CELS) from biological parts, forming functional modules integrated into living machines. These cognizant design principles for living systems encompass novel genetic circuit manipulation, self-assembly, cell–cell/matrix communication, and artificial tissues/organs enabled through systems biology, bioinformatics, computational biology, genetic engineering, and microfluidics. Here, we introduce design principles and a blueprint for forward production of robust and standardized M-CELS, which may undergo variable reiterations through the classic design-build-test-debug cycle. This Review provides practical and theoretical frameworks to forward-design, control, and optimize novel M-CELS. Potential applications include biopharmaceuticals, bioreactor factories, biofuels, environmental bioremediation, cellular computing, biohybrid digital technology, and experimental investigations into mechanisms of multicellular organisms normally hidden inside the “black box” of living cells.

JTD Keywords: cell-fate specification, endothelial-cells, escherichia-coli, extracellular-matrix, gene-expression noise, nuclear hormone-receptors, pluripotent stem-cells, primitive endoderm, transcription factors, Artificial tissues, Assembly cells, Biological parts, Biological systems, Bioremediation, Blood-brain-barrier, Cell engineering, Cell/matrix communication, Design principles, Environmental technology, Functional modules, Fundamental design, Genetic circuits, Genetic engineering, Living machines, Living systems, Medical applications, Molecular biology, Synthetic biology


Pérez-González, Carlos, Ceada, Gerardo, Matejcic, Marija, Trepat, Xavier, (2022). Digesting the mechanobiology of the intestinal epithelium Current Opinion In Genetics & Development 72, 82-90

The dizzying life of the homeostatic intestinal epithelium is governed by a complex interplay between fate, form, force and function. This interplay is beginning to be elucidated thanks to advances in intravital and ex vivo imaging, organoid culture, and biomechanical measurements. Recent discoveries have untangled the intricate organization of the forces that fold the monolayer into crypts and villi, compartmentalize cell types, direct cell migration, and regulate cell identity, proliferation and death. These findings revealed that the dynamic equilibrium of the healthy intestinal epithelium relies on its ability to precisely coordinate tractions and tensions in space and time. In this review, we discuss recent findings in intestinal mechanobiology, and highlight some of the many fascinating questions that remain to be addressed in this emerging field.Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.

JTD Keywords: crypt fission, designer matrices, differentiation, growth, gut, migration, model, scaffold, tissue mechanics, Cell migration, Cell proliferation, Ex vivo study, Human tissue, Intestine epithelium, Monolayer culture, Organoid, Review, Stem-cell, Tension, Traction therapy


Raymond, Yago, Johansson, Linh, Thorel, Emilie, Ginebra, Maria-Pau, (2022). Translation of three-dimensional printing of ceramics in bone tissue engineering and drug delivery Mrs Bulletin 47, 59-69

Beltrán G, Navajas D, García-Aznar JM, (2022). Mechanical modeling of lung alveoli: From macroscopic behaviour to cell mechano-sensing at microscopic level Journal Of The Mechanical Behavior Of Biomedical Materials 126, 105043

The mechanical signals sensed by the alveolar cells through the changes in the local matrix stiffness of the extracellular matrix (ECM) are determinant for regulating cellular functions. Therefore, the study of the mechanical response of lung tissue becomes a fundamental aspect in order to further understand the mechanosensing signals perceived by the cells in the alveoli. This study is focused on the development of a finite element (FE) model of a decellularized rat lung tissue strip, which reproduces accurately the mechanical behaviour observed in the experiments by means of a tensile test. For simulating the complex structure of the lung parenchyma, which consists of a heterogeneous and non-uniform network of thin-walled alveoli, a 3D model based on a Voronoi tessellation is developed. This Voronoi-based model is considered very suitable for recreating the geometry of cellular materials with randomly distributed polygons like in the lung tissue. The material model used in the mechanical simulations of the lung tissue was characterized experimentally by means of AFM tests in order to evaluate the lung tissue stiffness on the micro scale. Thus, in this study, the micro (AFM test) and the macro scale (tensile test) mechanical behaviour are linked through the mechanical simulation with the 3D FE model based on Voronoi tessellation. Finally, a micro-mechanical FE-based model is generated from the Voronoi diagram for studying the stiffness sensed by the alveolar cells in function of two independent factors: the stretch level of the lung tissue and the geometrical position of the cells on the extracellular matrix (ECM), distinguishing between pneumocyte type I and type II. We conclude that the position of the cells within the alveolus has a great influence on the local stiffness perceived by the cells. Alveolar cells located at the corners of the alveolus, mainly type II pneumocytes, perceive a much higher stiffness than those located in the flat areas of the alveoli, which correspond to type I pneumocytes. However, the high stiffness, due to the macroscopic lung tissue stretch, affects both cells in a very similar form, thus no significant differences between them have been observed. © 2021 The Authors

JTD Keywords: rat, scaffolds, stiffness, Afm, Animal cell, Animal experiment, Animal model, Animal tissue, Article, Biological organs, Cell function, Cells, Computational geometry, Cytology, Extracellular matrices, Extracellular matrix, Extracellular-matrix, Geometry, High stiffness, Human, Lung alveolus cell type 1, Lung alveolus cell type 2, Lung parenchyma, Lung tissue, Male, Mechanical behavior, Mechanical modeling, Mechanical simulations, Mechanosensing, Model-based opc, Nonhuman, Physical model, Rat, Rigidity, Stiffness, Stiffness matrix, Tensile testing, Thin walled structures, Three dimensional finite element analysis, Tissue, Type ii, Voronoi tessellations


Boda, SK, Aparicio, C, (2022). Dual keratinocyte-attachment and anti-inflammatory coatings for soft tissue sealing around transmucosal oral implants Biomaterials Science 10, 665-677

Unlike the attachment of soft epithelial skin tissue to penetrating solid natural structures like fingernails and teeth, sealing around percutaneous/permucosal devices such as dental implants is hindered by inflammation and epidermal down growth. Here, we employed a dual keratinocyte-adhesive peptide and anti-inflammatory biomolecule coating on titanium to promote oral epithelial tissue attachment. For minimizing inflammation-triggered epidermal down growth, we coated pristine and oxygen plasma pre-treated polished titanium (pTi) with conjugated linoleic acid (CLA). Further, in order to aid in soft tissue attachment via the formation of hemidesmosomes, adhesive structures by oral keratinocytes, we coated the anionic linoleic acid (LA) adsorbed titanium with cationic cell adhesive peptides (CAP), LamLG3, a peptide derived from Laminin 332, the major extracellular matrix component of the basement membrane in skin tissue and Net1, derived from Netrin-1, a neural chemoattractant capable of epithelial cell attachment via alpha 6 beta 4 integrins. The dual CLA-CAP coatings on pTi were characterized by X-ray photoelectron spectroscopy and dynamic water contact angle measurements. The proliferation of human oral keratinocytes (TERT-2/OKF6) was accelerated on the peptide coated titanium while also promoting the expression of Col XVII and beta-4 integrin, two markers for hemidesmosomes. Simultaneously, CLA coating suppressed the production of inducible nitric oxide synthase (anti-iNOS); a pro-inflammatory M1 marker expressed in lipopolysaccharide (LPS) stimulated murine macrophages (RAW 264.7) and elevated expression of anti-CD206, associated to an anti-inflammatory M2 macrophage phenotype. Taken together, the dual keratinocyte-adhesive peptide and anti-inflammatory biomolecule coating on titanium can help reduce inflammation and promote permucosal/peri-implant soft tissue sealing.

JTD Keywords: Adhesives, Animal, Animals, Anti-inflammatories, Anti-inflammatory agents, Antiinflammatory agent, Biomolecules, Bone, Cell adhesion, Cell-adhesives, Coatings, Conjugated linoleic acid, Conjugated linoleic-acid, Contact angle, Hemidesmosome, Hemidesmosomes, Human, Humans, Hydroxyapatite, Inflammation, Integrins, Keratinocyte, Keratinocytes, Linoleic acid, Macrophages, Mice, Mouse, Nitric oxide, Oral implants, Pathology, Peptides, Skin tissue, Soft tissue, Supplementation, Surface properties, Surface property, Tissue, Titania, Titanium, X ray photoelectron spectroscopy


Gawish R, Starkl P, Pimenov L, Hladik A, Lakovits K, Oberndorfer F, Cronin SJF, Ohradanova-Repic A, Wirnsberger G, Agerer B, Endler L, Capraz T, Perthold JW, Cikes D, Koglgruber R, Hagelkruys A, Montserrat N, Mirazimi A, Boon L, Stockinger H, Bergthaler A, Oostenbrink C, Penninger JM, Knapp S, (2022). ACE2 is the critical in vivo receptor for SARS-CoV-2 in a novel COVID-19 mouse model with TNF-and IFNy-driven immunopathology Elife 11, e74623

Despite tremendous progress in the understanding of COVID-19, mechanistic insight into immunological, disease-driving factors remains limited. We generated maVie16, a mouse-adapted SARS-CoV-2, by serial passaging of a human isolate. In silico modeling revealed how only three Spike mutations of maVie16 enhanced interaction with murine ACE2. maVie16 induced profound pathology in BALB/c and C57BL/6 mice, and the resulting mouse COVID-19 (mCOVID-19) replicated critical aspects of human disease, including early lymphopenia, pulmonary immune cell infiltration, pneumonia, and specific adaptive immunity. Inhibition of the proinflammatory cyto-kines IFN? and TNF substantially reduced immunopathology. Importantly, genetic ACE2-deficiency completely prevented mCOVID-19 development. Finally, inhalation therapy with recombinant ACE2 fully protected mice from mCOVID-19, revealing a novel and efficient treatment. Thus, we here present maVie16 as a new tool to model COVID-19 for the discovery of new therapies and show that disease severity is determined by cytokine-driven immunopathology and critically dependent on ACE2 in vivo. © Gawish et al.

JTD Keywords: covid-19 mouse model, covid-19 therapy, cytokine storm, mavie16, mouse, program, recombinant soluble ace2, tmprss2, Adaptive immunity, Angiotensin converting enzyme 2, Angiotensin-converting enzyme 2, Animal, Animal cell, Animal experiment, Animal model, Animal tissue, Animals, Apoptosis, Article, Bagg albino mouse, Breathing rate, Bronchoalveolar lavage fluid, C57bl mouse, Cell composition, Cell infiltration, Controlled study, Coronavirus disease 2019, Coronavirus spike glycoprotein, Covid-19, Cytokeratin 18, Cytokine production, Dipeptidyl carboxypeptidase, Disease model, Disease models, animal, Disease severity, Drosophila-melanogaster, Enzyme linked immunosorbent assay, Expression vector, Flow cytometry, Gamma interferon, Gene editing, Gene expression, Gene mutation, Genetic engineering, Genetics, Glycosylation, High mobility group b1 protein, Histology, Histopathology, Immune response, Immunocompetent cell, Immunology, Immunopathology, Interferon-gamma, Interleukin 2, Metabolism, Mice, inbred balb c, Mice, inbred c57bl, Mouse-adapted sars-cov-2, Myeloperoxidase, Neuropilin 1, Nonhuman, Nucleocapsid protein, Pathogenicity, Peptidyl-dipeptidase a, Pyroptosis, Renin angiotensin aldosterone system, Rna extraction, Rna isolation, Sars-cov-2, Severe acute respiratory syndrome coronavirus 2, Spike glycoprotein, coronavirus, T lymphocyte activation, Trabecular meshwork, Tumor necrosis factor, Virology, Virus load, Virus replication, Virus transmission, Virus virulence


Tejo-Otero A, Fenollosa-Artés F, Achaerandio I, Rey-Vinolas S, Buj-Corral I, Mateos-Timoneda MÁ, Engel E, (2022). Soft-Tissue-Mimicking Using Hydrogels for the Development of Phantoms Gels 8, 40

With the currently available materials and technologies it is difficult to mimic the mechanical properties of soft living tissues. Additionally, another significant problem is the lack of information about the mechanical properties of these tissues. Alternatively, the use of phantoms offers a promising solution to simulate biological bodies. For this reason, to advance in the state-of-the-art a wide range of organs (e.g., liver, heart, kidney as well as brain) and hydrogels (e.g., agarose, polyvinyl alcohol –PVA–, Phytagel –PHY– and methacrylate gelatine –GelMA–) were tested regarding their mechanical properties. For that, viscoelastic behavior, hardness, as well as a non-linear elastic mechanical response were measured. It was seen that there was a significant difference among the results for the different mentioned soft tissues. Some of them appear to be more elastic than viscous as well as being softer or harder. With all this information in mind, a correlation between the mechanical properties of the organs and the different materials was performed. The next conclusions were drawn: (1) to mimic the liver, the best material is 1% wt agarose; (2) to mimic the heart, the best material is 2% wt agarose; (3) to mimic the kidney, the best material is 4% wt GelMA; and (4) to mimic the brain, the best materials are 4% wt GelMA and 1% wt agarose. Neither PVA nor PHY was selected to mimic any of the studied tissues. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.

JTD Keywords: brain, composite hydrogel, elastography, hardness, hydrogels, in-vitro, liver, materials, mechanical-properties, mimicking, soft tissues, tissue scaffolding, viscoelasticity, warner-braztler shear test, Dynamic mechanical analysis, Hardness, Hydrogels, Materials, Mimicking, Soft tissues, Tissue scaffolding, Viscoelastic characterization, Viscoelasticity, Warner–braztler shear test


Dulay, S, Rivas, L, Pla, L, Berdun, S, Eixarch, E, Gratacos, E, Illa, M, Mir, M, Samitier, J, (2021). Fetal ischemia monitoring with in vivo implanted electrochemical multiparametric microsensors Journal Of Biological Engineering 15, 28

Under intrauterine growth restriction (IUGR), abnormal attainment of the nutrients and oxygen by the fetus restricts the normal evolution of the prenatal causing in many cases high morbidity being one of the top-ten causes of neonatal death. The current gold standards in hospitals to detect this relevant problem is the clinical observation by echography, cardiotocography and Doppler. These qualitative techniques are not conclusive and requires risky invasive fetal scalp blood testing and/or amniocentesis. We developed micro-implantable multiparametric electrochemical sensors for measuring ischemia in real time in fetal tissue and vascular. This implantable technology is designed to continuous monitoring for an early detection of ischemia to avoid potential fetal injury. Two miniaturized electrochemical sensors were developed based on oxygen and pH detection. The sensors were optimized in vitro under controlled concentration, to assess the selectivity and sensitivity required. The sensors were then validated in vivo in the ewe fetus model, by means of their insertion in the muscle leg and inside the iliac artery of the fetus. Ischemia was achieved by gradually obstructing the umbilical cord to regulate the amount of blood reaching the fetus. An important challenge in fetal monitoring is the detection of low levels of oxygen and pH changes under ischemic conditions, requiring high sensitivity sensors. Significant differences were observed in both; pH and pO(2) sensors under changes from normoxia to hypoxia states in the fetus tissue and vascular with both sensors. Herein, we demonstrate the feasibility of the developed sensors for future fetal monitoring in medical applications.

JTD Keywords: electrochemical biosensor, implantable sensor, in vivo validation, ischemia detection, tissue and vascular monitoring, Animal experiment, Animal model, Animal tissue, Article, Blood-gases, Brain, Classification, Controlled study, Diagnosis, Doppler, Early diagnosis, Electrochemical analysis, Electrochemical biosensor, Ewe, Feasibility study, Female, Fetus, Fetus disease, Fetus monitoring, Gestational age, Hypoxemia, Iliac artery, Implantable sensor, In vivo validation, Intrauterine growth restriction, Intrauterine growth retardation, Ischemia detection, Leg muscle, Management, Nonhuman, Oxygen consumption, Ph, Ph and oxygen detection, Ph measurement, Process optimization, Sheep, Tissue and vascular monitoring, Umbilical-cord occlusion


Sans, Jordi, Sanz, Vanesa, Turon, Pau, Alemán, Carlos, (2021). Enhanced CO2 Conversion into Ethanol by Permanently Polarized Hydroxyapatite through C-C Coupling Chemcatchem 13, 5025-5033

Lopez-Muñoz, Gerardo A, Fernández-Costa, Juan M, Ortega, Maria Alejandra, Balaguer-Trias, Jordina, Martin-Lasierra, Eduard, Ramón-Azcón, Javier, (2021). Plasmonic nanocrystals on polycarbonate substrates for direct and label-free biodetection of Interleukin-6 in bioengineered 3D skeletal muscles Nanophotonics 10, 4477-4488

Abstract The development of nanostructured plasmonic biosensors has been widely widespread in the last years, motivated by the potential benefits they can offer in integration, miniaturization, multiplexing opportunities, and enhanced performance label-free biodetection in a wide field of applications. Between them, engineering tissues represent a novel, challenging, and prolific application field for nanostructured plasmonic biosensors considering the previously described benefits and the low levels of secreted biomarkers (?pM–nM) to detect. Here, we present an integrated plasmonic nanocrystals-based biosensor using high throughput nanostructured polycarbonate substrates. Metallic film thickness and incident angle of light for reflectance measurements were optimized to enhance the detection of antibody–antigen biorecognition events using numerical simulations. We achieved an enhancement in biodetection up to 3× as the incident angle of light decreases, which can be related to shorter evanescent decay lengths. We achieved a high reproducibility between channels with a coefficient of variation below 2% in bulk refractive index measurements, demonstrating a high potential for multiplexed sensing. Finally, biosensing potential was demonstrated by the direct and label-free detection of interleukin-6 biomarker in undiluted cell culture media supernatants from bioengineered 3D skeletal muscle tissues stimulated with different concentrations of endotoxins achieving a limit of detection (LOD) of ? 0.03 ng/mL (1.4 pM).

JTD Keywords: assay, crystals, drug, label-free biosensing, molecules, plasmonic nanostructures, sensors, skeletal muscle, tissue engineering, Biodetection, Biomarkers, Biosensors, Cell culture, Cells, Chemical detection, Histology, Interleukin-6, Interleukin6 (il6), Label free, Label-free biosensing, Muscle, Nano-structured, Nanocrystals, Plasmonic nanocrystals, Plasmonic nanostructures, Plasmonics, Polycarbonate substrates, Polycarbonates, Refractive index, Sensitivity, Skeletal muscle, Tissue engineering, Tissues engineerings


Brennan M, Monahan DS, Brulin B, Gallinetti S, Humbert P, Tringides C, Canal C, Ginebra MP, Layrolle P, (2021). Biomimetic versus sintered macroporous calcium phosphate scaffolds enhanced bone regeneration and human mesenchymal stromal cell engraftment in calvarial defects Acta Biomaterialia 135, 689-704

In contrast to sintered calcium phosphates (CaPs) commonly employed as scaffolds to deliver mesenchymal stromal cells (MSCs) targeting bone repair, low temperature setting conditions of calcium deficient hydroxyapatite (CDHA) yield biomimetic topology with high specific surface area. In this study, the healing capacity of CDHA administering MSCs to bone defects is evaluated for the first time and compared with sintered beta-tricalcium phosphate (β-TCP) constructs sharing the same interconnected macroporosity. Xeno-free expanded human bone marrow MSCs attached to the surface of the hydrophobic β-TCP constructs, while infiltrating the pores of the hydrophilic CDHA. Implantation of MSCs on CaPs for 8 weeks in calvaria defects of nude mice exhibited complete healing, with bone formation aligned along the periphery of β-TCP, and conversely distributed within the pores of CDHA. Human monocyte-osteoclast differentiation was inhibited in vitro by direct culture on CDHA compared to β-TCP biomaterials and indirectly by administration of MSC-conditioned media generated on CDHA, while MSCs increased osteoclastogenesis in both CaPs in vivo. MSC engraftment was significantly higher in CDHA constructs, and also correlated positively with bone in-growth in scaffolds. These findings demonstrate that biomimetic CDHA are favorable carriers for MSC therapies and should be explored further towards clinical bone regeneration strategies. Statement of significance: Delivery of mesenchymal stromal cells (MSCs) on calcium phosphate (CaP) biomaterials enhances reconstruction of bone defects. Traditional CaPs are produced at high temperature, but calcium deficient hydroxyapatite (CDHA) prepared at room temperature yields a surface structure more similar to native bone mineral. The objective of this study was to compare the capacity of biomimetic CDHA scaffolds with sintered β-TCP scaffolds for bone repair mediated by MSCs for the first time. In vitro, greater cell infiltration occurred in CDHA scaffolds and following 8 weeks in vivo, MSC engraftment was higher in CDHA compared to β-TCP, as was bone in-growth. These findings demonstrate the impact of material features such as surface structure, and highlight that CDHA should be explored towards clinical bone regeneration strategies.

JTD Keywords: beta-tricalcium phosphate, bone regeneration, calcium deficient hydroxyapatite, differentiation, engraftment, human bone marrow mesenchymal stromal cells, hydroxyapatite scaffolds, in-vitro, inhibition, osteogenesis, osteoinduction, stem-cells, surface-topography, tissue, Beta-tricalcium phosphate, Bone regeneration, Calcium deficient hydroxyapatite, Engraftment, Human bone marrow mesenchymal stromal cells


Konka J, Buxadera-Palomero J, Espanol M, Ginebra M-P, (2021). 3D printing of hierarchical porous biomimetic hydroxyapatite scaffolds: Adding concavities to the convex filaments Acta Biomaterialia 134, 744-759

Porosity plays a key role on the osteogenic performance of bone scaffolds. Direct Ink Writing (DIW) allows the design of customized synthetic bone grafts with patient-specific architecture and controlled macroporosity. Being an extrusion-based technique, the scaffolds obtained are formed by arrays of cylindrical filaments, and therefore have convex surfaces. This may represent a serious limitation, as the role of surface curvature and more specifically the stimulating role of concave surfaces in osteoinduction and bone growth has been recently highlighted. Hence the need to design strategies that allow the introduction of concave pores in DIW scaffolds. In the current study, we propose to add gelatin microspheres as a sacrificial material in a self-setting calcium phosphate ink. Neither the phase transformation responsible for the hardening of the scaffold nor the formation of characteristic network of needle-like hydroxyapatite crystals was affected by the addition of gelatin microspheres. The partial dissolution of the gelatin resulted in the creation of spherical pores throughout the filaments and exposed on the surface, increasing filament porosity from 0.2 % to 67.9 %. Moreover, the presence of retained gelatin proved to have a significant effect on the mechanical properties, reducing the strength but simultaneously giving the scaffolds an elastic behavior, despite the high content of ceramic as a continuous phase. Notwithstanding the inherent difficulty of in vitro cultures with this highly reactive material an enhancement of MG-63 cell proliferation, as well as better spreading of hMSCs was recorded on the developed scaffolds. Statement of significance: Recent studies have stressed the role that concave surfaces play in tissue regeneration and, more specifically, in osteoinduction and osteogenesis. Direct ink writing enables the production of patient-specific bone grafts with controlled architecture. However, besides many advantages, it has the serious limitation that the surfaces obtained are convex. In this article, for the first time we develop a strategy to introduce concave pores in the printed filaments of biomimetic hydroxyapatite by incorporation and partial dissolution of gelatin microspheres. The retention of part of the gelatin results in a more elastic behavior compared to the brittleness of hydroxyapatite scaffolds, while the needle-shaped nanostructure of biomimetic hydroxyapatite is maintained and gelatin-coated concave pores on the surface of the filaments enhance cell spreading. © 2021 The Authors

JTD Keywords: 3d printing, bioceramics, biomimetic, bone, bone regeneration, concavity, concavity, bone regeneration, gelatin, hydrogel, hydroxyapatite, microspheres, osteoinduction, porosity, porous filament, substitutes, tissue-growth, 3d printing, Biomimetic, Calcium-phosphate scaffolds, Concavity, bone regeneration, Gelatin, Hydroxyapatite, Porous filament


Illa M, Pla L, Berdún S, Mir M, Rivas L, Dulay S, Picard-Hagen N, Samitier J, Gratacós E, Eixarch E, (2021). Miniaturized electrochemical sensors to monitor fetal hypoxia and acidosis in a pregnant sheep model Biomedicines 9,

Perinatal asphyxia is a major cause of severe brain damage and death. For its prenatal identification, Doppler ultrasound has been used as a surrogate marker of fetal hypoxia. However, Doppler evaluation cannot be performed continuously. We have evaluated the performance of a miniaturized multiparametric sensor aiming to evaluate tissular oxygen and pH changes continuously in an umbilical cord occlusion (UCO) sheep model. The electrochemical sensors were inserted in fetal hindlimb skeletal muscle and electrochemical signals were recorded. Fetal hemodynamic changes and metabolic status were also monitored during the experiment. Additionally, histological assessment of the tissue surrounding the sensors was performed. Both electrochemical sensors detected the pO2 and pH changes induced by the UCO and these changes were correlated with hemodynamic parameters as well as with pH and oxygen content in the blood. Finally, histological assessment revealed no signs of alteration on the same day of insertion. This study provides the first evidence showing the application of miniaturized multiparametric electrochemical sensors detecting changes in oxygen and pH in skeletal muscular tissue in a fetal sheep model.

JTD Keywords: continuous monitoring of acid-base status, diagnosis, doppler, electrochemical sensors, growth restriction, high-risk pregnancies, human-fetus, management, responses, tissue ph, Continuous monitoring of acid-base status, Electrochemical sensors, High-risk pregnancies, Umbilical cord occlusion, Umbilical-cord occlusion


Nyga, Agata, Muñoz, Jose J., Dercksen, Suze, Fornabaio, Giulia, Uroz, Marina, Trepat, Xavier, Baum, Buzz, Matthews, Helen K., Conte, Vito, (2021). Oncogenic RAS instructs morphological transformation of human epithelia via differential tissue mechanics Science Advances 7, eabg6467

Rubí-Sans G, Nyga A, Rebollo E, Pérez-Amodio S, Otero J, Navajas D, Mateos-Timoneda MA, Engel E, (2021). Development of Cell-Derived Matrices for Three-Dimensional in Vitro Cancer Cell Models Acs Applied Materials & Interfaces 13, 44108-44123

Most morphogenetic and pathological processes are driven by cells responding to the surrounding matrix, such as its composition, architecture, and mechanical properties. Despite increasing evidence for the role of extracellular matrix (ECM) in tissue and disease development, many in vitro substitutes still fail to effectively mimic the native microenvironment. We established a novel method to produce macroscale (>1 cm) mesenchymal cell-derived matrices (CDMs) aimed to mimic the fibrotic tumor microenvironment surrounding epithelial cancer cells. CDMs are produced by human adipose mesenchymal stem cells cultured in sacrificial 3D scaffold templates of fibronectin-coated poly-lactic acid microcarriers (MCs) in the presence of macromolecular crowders. We showed that decellularized CDMs closely mimic the fibrillar protein composition, architecture, and mechanical properties of human fibrotic ECM from cancer masses. CDMs had highly reproducible composition made of collagen types I and III and fibronectin ECM with tunable mechanical properties. Moreover, decellularized and MC-free CDMs were successfully repopulated with cancer cells throughout their 3D structure, and following chemotherapeutic treatment, cancer cells showed greater doxorubicin resistance compared to 3D culture in collagen hydrogels. Collectively, these results support the use of CDMs as a reproducible and tunable tool for developing 3D in vitro cancer models.

JTD Keywords: 3d cell-derived matrices, adipose mesenchymal stem cells, collagen matrix, colorectal adenocarcinoma, cytotoxicity assay, deposition, expansion, extracellular microenvironment, extracellular-matrix, fibronectin, growth, macromolecular crowders, microcarriers, scaffolds, tissue, 3d cell-derived matrices, Adipose mesenchymal stem cells, Cytotoxicity assay, Extracellular microenvironment, Macromolecular crowders, Mesenchymal stem-cells, Microcarriers


Rial-Hermida MI, Rey-Rico A, Blanco-Fernandez B, Carballo-Pedrares N, Byrne EM, Mano JF, (2021). Recent Progress on Polysaccharide-Based Hydrogels for Controlled Delivery of Therapeutic Biomolecules Acs Biomaterials Science & Engineering 7, 4102-4127

A plethora of applications using polysaccharides have been developed in recent years due to their availability as well as their frequent nontoxicity and biodegradability. These polymers are usually obtained from renewable sources or are byproducts of industrial processes, thus, their use is collaborative in waste management and shows promise for an enhanced sustainable circular economy. Regarding the development of novel delivery systems for biotherapeutics, the potential of polysaccharides is attractive for the previously mentioned properties and also for the possibility of chemical modification of their structures, their ability to form matrixes of diverse architectures and mechanical properties, as well as for their ability to maintain bioactivity following incorporation of the biomolecules into the matrix. Biotherapeutics, such as proteins, growth factors, gene vectors, enzymes, hormones, DNA/RNA, and antibodies are currently in use as major therapeutics in a wide range of pathologies. In the present review, we summarize recent progress in the development of polysaccharide-based hydrogels of diverse nature, alone or in combination with other polymers or drug delivery systems, which have been implemented in the delivery of biotherapeutics in the pharmaceutical and biomedical fields. © 2021 American Chemical Society.

JTD Keywords: biodegradable dextran hydrogels, biotherapeutics, bone morphogenetic protein-2, carrageenan-based hydrogels, chitosan-based hydrogels, controlled delivery, controlled-release, cross-linked hydrogels, growth-factor delivery, hydrogels, in-vitro characterization, polysaccharides, self-healing hydrogel, stimuli-responsiveness, tissue engineering, Antibodies, Bioactivity, Biodegradability, Biomedical fields, Biomolecules, Biotherapeutics, Chemical modification, Circular economy, Controlled delivery, Controlled drug delivery, Delivery systems, Drug delivery system, Functional polymers, Hyaluronic-acid hydrogels, Hydrogels, Industrial processs, Polysaccharides, Recent progress, Renewable sources, Stimuli-responsiveness, Targeted drug delivery, Tissue engineering, Waste management


Falcones B, Sanz-Fraile H, Marhuenda E, Mendizábal I, Cabrera-Aguilera I, Malandain N, Uriarte JJ, Almendros I, Navajas D, Weiss DJ, Farré R, Otero J, (2021). Bioprintable lung extracellular matrix hydrogel scaffolds for 3d culture of mesenchymal stromal cells Polymers 13, 2350

Mesenchymal stromal cell (MSC)-based cell therapy in acute respiratory diseases is based on MSC secretion of paracrine factors. Several strategies have proposed to improve this are being explored including pre-conditioning the MSCs prior to administration. We here propose a strategy for improving the therapeutic efficacy of MSCs based on cell preconditioning by growing them in native extracellular matrix (ECM) derived from the lung. To this end, a bioink with tunable stiffness based on decellularized porcine lung ECM hydrogels was developed and characterized. The bioink was suitable for 3D culturing of lung-resident MSCs without the need for additional chemical or physical crosslinking. MSCs showed good viability, and contraction assays showed the existence of cell–matrix interactions in the bioprinted scaffolds. Adhesion capacity and length of the focal adhesions formed were increased for the cells cultured within the lung hydrogel scaffolds. Also, there was more than a 20-fold increase of the expression of the CXCR4 receptor in the 3D-cultured cells compared to the cells cultured in plastic. Secretion of cytokines when cultured in an in vitro model of lung injury showed a decreased secretion of pro-inflammatory mediators for the cells cultured in the 3D scaffolds. Moreover, the morphology of the harvested cells was markedly different with respect to conventionally (2D) cultured MSCs. In conclusion, the developed bioink can be used to bioprint structures aimed to improve preconditioning MSCs for therapeutic purposes.

JTD Keywords: 3d bioprinting, acute lung injury, adhesion, collagen, differentiation, dimension, elastic properties, extracellular matrix, hydrogels, in-vitro, mechanical-properties, mesenchymal stromal cells, microenvironment, potentiate, tissue engineering, 3d bioprinting, Acute lung injury, Extracellular matrix, Hydrogels, Mesenchymal stromal cells, Stem-cells, Tissue engineering


Fernández-Garibay X, Ortega MA, Cerro-Herreros E, Comelles J, Martínez E, Artero R, Fernández-Costa JM, Ramón-Azcón J, (2021). Bioengineered in vitro 3D model of myotonic dystrophy type 1 human skeletal muscle Biofabrication 13, 35035

Myotonic dystrophy type 1 (DM1) is the most common hereditary myopathy in the adult population. The disease is characterized by progressive skeletal muscle degeneration that produces severe disability. At present, there is still no effective treatment for DM1 patients, but the breakthroughs in understanding the molecular pathogenic mechanisms in DM1 have allowed the testing of new therapeutic strategies. Animal models and in vitro two-dimensional cell cultures have been essential for these advances. However, serious concerns exist regarding how faithfully these models reproduce the biological complexity of the disease. Biofabrication tools can be applied to engineer human three-dimensional (3D) culture systems that complement current preclinical research models. Here, we describe the development of the first in vitro 3D model of DM1 human skeletal muscle. Transdifferentiated myoblasts from patient-derived fibroblasts were encapsulated in micromolded gelatin methacryloyl-carboxymethyl cellulose methacrylate hydrogels through photomold patterning on functionalized glass coverslips. These hydrogels present a microstructured topography that promotes myoblasts alignment and differentiation resulting in highly aligned myotubes from both healthy and DM1 cells in a long-lasting cell culture. The DM1 3D microtissues recapitulate the molecular alterations detected in patient biopsies. Importantly, fusion index analyses demonstrate that 3D micropatterning significantly improved DM1 cell differentiation into multinucleated myotubes compared to standard cell cultures. Moreover, the characterization of the 3D cultures of DM1 myotubes detects phenotypes as the reduced thickness of myotubes that can be used for drug testing. Finally, we evaluated the therapeutic effect of antagomiR-23b administration on bioengineered DM1 skeletal muscle microtissues. AntagomiR-23b treatment rescues both molecular DM1 hallmarks and structural phenotype, restoring myotube diameter to healthy control sizes. Overall, these new microtissues represent an improvement over conventional cell culture models and can be used as biomimetic platforms to establish preclinical studies for myotonic dystrophy.

JTD Keywords: 3d cell culture, hydrogel micropatterning, myotonic dystrophy, skeletal muscle, tissue engineering, 3d cell culture, Hydrogel micropatterning, Myotonic dystrophy, Skeletal muscle, Tissue engineering


López-Canosa A, Perez-Amodio S, Yanac-Huertas E, Ordoño J, Rodriguez-Trujillo R, Samitier J, Castaño O, Engel E, (2021). A microphysiological system combining electrospun fibers and electrical stimulation for the maturation of highly anisotropic cardiac tissue Biofabrication 13, 35047

The creation of cardiac tissue models for preclinical testing is still a non-solved problem in drug discovery, due to the limitations related to thein vitroreplication of cardiac tissue complexity. Among these limitations, the difficulty of mimicking the functional properties of the myocardium due to the immaturity of the used cells hampers the obtention of reliable results that could be translated into human patients.In vivomodels are the current gold standard to test new treatments, although it is widely acknowledged that the used animals are unable to fully recapitulate human physiology, which often leads to failures during clinical trials. In the present work, we present a microfluidic platform that aims to provide a range of signaling cues to immature cardiac cells to drive them towards an adult phenotype. The device combines topographical electrospun nanofibers with electrical stimulation in a microfabricated system. We validated our platform using a co-culture of neonatal mouse cardiomyocytes and cardiac fibroblasts, showing that it allows us to control the degree of anisotropy of the cardiac tissue inside the microdevice in a cost-effective way. Moreover, a 3D computational model of the electrical field was created and validated to demonstrate that our platform is able to closely match the distribution obtained with the gold standard (planar electrode technology) using inexpensive rod-shaped biocompatible stainless-steel electrodes. The functionality of the electrical stimulation was shown to induce a higher expression of the tight junction protein Cx-43, as well as the upregulation of several key genes involved in conductive and structural cardiac properties. These results validate our platform as a powerful tool for the tissue engineering community due to its low cost, high imaging compatibility, versatility, and high-throughput configuration capabilities.

JTD Keywords: bioreactor, cardiac tissue engineering, cardiomyocytes, electrospinning, fabrication, fibroblasts, heart-on-a-chip, heart-tissue, in vitro models, myocardium, orientation, platform, scaffolds, Cardiac tissue engineering, Electrospinning, Field stimulation, Heart-on-a-chip, In vitro models, Microphysiological system


Velasco-Mallorqui, F, Rodriguez-Comas, J, Ramon-Azcon, J, (2021). Cellulose-based scaffolds enhance pseudoislets formation and functionality Biofabrication 13, 35044

In vitro research for the study of type 2 diabetes (T2D) is frequently limited by the availability of a functional model for islets of Langerhans. To overcome the limitations of obtaining pancreatic islets from different sources, such as animal models or human donors, immortalized cell lines as the insulin-producing INS1E beta-cells have appeared as a valid alternative to model insulin-related diseases. However, immortalized cell lines are mainly used in flat surfaces or monolayer distributions, not resembling the spheroid-like architecture of the pancreatic islets. To generate islet-like structures, the use of scaffolds appeared as a valid tool to promote cell aggregations. Traditionally-used hydrogel encapsulation methods do not accomplish all the requisites for pancreatic tissue engineering, as its poor nutrient and oxygen diffusion induces cell death. Here, we use cryogelation technology to develop a more resemblance scaffold with the mechanical and physical properties needed to engineer pancreatic tissue. This study shows that carboxymethyl cellulose (CMC) cryogels prompted cells to generate beta-cell clusters in comparison to gelatin-based scaffolds, that did not induce this cell organization. Moreover, the high porosity achieved with CMC cryogels allowed us to create specific range pseudoislets. Pseudoislets formed within CMC-scaffolds showed cell viability for up to 7 d and a better response to glucose over conventional monolayer cultures. Overall, our results demonstrate that CMC-scaffolds can be used to control the organization and function of insulin-producing beta-cells, representing a suitable technique to generate beta-cell clusters to study pancreatic islet function.

JTD Keywords: biomaterial, cryogel, pancreatic islets, scaffold, tissue engineering, ?-cell, Architecture, Beta-cell, Beta-cell heterogeneity, Biomaterial, Carboxymethyl cellulose, Cell culture, Cell death, Cell engineering, Cell organization, Cells, Cellulose, Cryogel, Cryogels, Cytoarchitecture, Delivery, Encapsulation methods, Gelation, Gene-expression, Immortalized cells, Insulin, Insulin secretory responses, Islets of langerhans, Mechanical and physical properties, Monolayer culture, Monolayers, Pancreatic islets, Pancreatic tissue, Pancreatic-islets, Proliferation, Scaffold, Scaffolds, Scaffolds (biology), Size, Tissue, Tissue engineering


Minguela J, Müller DW, Mücklich F, Llanes L, Ginebra MP, Roa JJ, Mas-Moruno C, (2021). Peptidic biofunctionalization of laser patterned dental zirconia: A biochemical-topographical approach Materials Science & Engineering C-Materials For Biological Applications 125, 112096

A dual approach employing peptidic biofunctionalization and laser micro-patterns on dental zirconia was explored, with the aim of providing a flexible tool to improve tissue integration of restorations. Direct laser interference patterning with a femtosecond Ti:Sapphire laser was employed, and two periodic grooved patterns were produced with a periodicity of 3 and 10 μm. A platform containing the cell-adhesive RGD and the osteogenic DWIVA peptides was used to functionalize the grooved surfaces. Topography and surface damage were characterized by confocal laser scanning (CLSM), scanning electron and scanning transmission electron microscopy techniques. The surface patterns exhibited a high homogeneity and subsurface damage was found in the form of nano-cracks and nano-pores, at the bottom of the valleys. Accelerated tests in water steam were carried out to assess hydrothermal degradation resistance, which slightly decreased after the laser treatment. Interestingly, the detrimental effects of the laser modification were reverted by a post-laser thermal treatment. The attachment of the molecule was verified trough fluorescence CLSM and X-ray photoelectron spectroscopy. Finally, the biological properties of the surfaces were studied in human mesenchymal stem cells. Cell adhesion, morphology, migration and differentiation were investigated. Cells on grooved surfaces displayed an elongated morphology and aligned along the patterns. On these surfaces, migration was greatly enhanced along the grooves, but also highly restricted in the perpendicular direction as compared to flat specimens. After biofunctionalization, cell number and cell area increased and well-developed cell cytoskeletons were observed. However, no effects on cell migration were found for the peptidic platform. Although some osteogenic potential was found in specimens grooved with a periodicity of 10 μm, the largest effects were observed from the biomolecule, which favored upregulation of several genes related to osteoblastic differentiation in all the surfaces.

JTD Keywords: alumina toughened zirconia, cell alignment, grain-size, implants, interference, laser patterning, osteogenic differentiation, osteointegration, peptides, surface functionalization, surface-topography, tissue, titanium surface, Laser patterning, Low-temperature degradation, Osteointegration, Peptides, Surface functionalization, Zirconia


Elosegui-Artola A, (2021). The extracellular matrix viscoelasticity as a regulator of cell and tissue dynamics Current Opinion In Cell Biology 72, 10-18

The extracellular matrix mechanical properties regulate processes in development, cancer, and fibrosis. Among the distinct mechanical properties, the vast majority of research has focused on the extracellular matrix's elasticity as the primary determinant of cell and tissue behavior. However, both cells and the extracellular matrix are not only elastic but also viscous. Despite viscoelasticity being a universal feature of living tissues, our knowledge of the influence of the extracellular matrix's viscoelasticity in cell behavior is limited. This mini-review describes some of the recent findings that have highlighted the role of the extracellular matrix's viscoelasticity in cell and tissue dynamics.

JTD Keywords: behavior, cell adhesion, cell mechanics, cell migration, deformability, extracellular matrix, extracellular matrix mechanics, fluidity, forces, hydrogels, mechanobiology, mechanotransduction, tissue mechanics, viscoelasticity, viscosity, Cell adhesion, Cell mechanics, Cell migration, Extracellular matrix, Extracellular matrix mechanics, Fluidity, Mechanobiology, Mechanotransduction, Migration, Tissue mechanics, Viscoelasticity, Viscosity


Dulay S, Rivas L, Miserere S, Pla L, Berdún S, Parra J, Eixarch E, Gratacós E, Illa M, Mir M, Samitier J, (2021). in vivo Monitoring with micro-implantable hypoxia sensor based on tissue acidosis Talanta 226, 122045

© 2020 Elsevier B.V. Hypoxia is a common medical problem, sometimes difficult to detect and caused by different situations. Control of hypoxia is of great medical importance and early detection is essential to prevent life threatening complications. However, the few current methods are invasive, expensive, and risky. Thus, the development of reliable and accurate sensors for the continuous monitoring of hypoxia is of vital importance for clinical monitoring. Herein, we report an implantable sensor to address these needs. The developed device is a low-cost, miniaturised implantable electrochemical sensor for monitoring hypoxia in tissue by means of pH detection. This technology is based on protonation/deprotonation of polypyrrole conductive polymer. The sensor was optimized in vitro and tested in vivo intramuscularly and ex vivo in blood in adult rabbits with respiration-induced hypoxia and correlated with the standard device ePOCTM. The sensor demonstrated excellent sensitivity and reproducibility; 46.4 ± 0.4 mV/pH in the pH range of 4–9 and the selectivity coefficient exhibited low interference activity in vitro. The device was linear (R2 = 0.925) with a low dispersion of the values (n = 11) with a cut-off of 7.1 for hypoxia in vivo and ex vivo. Statistics with one-way ANOVA (α = 0.05), shows statistical differences between hypoxia and normoxia states and the good performance of the pH sensor, which demonstrated good agreement with the standard device. The sensor was stable and functional after 18 months. The excellent results demonstrated the feasibility of the sensors in real-time monitoring of intramuscular tissue and blood for medical applications.

JTD Keywords: biocompatibility, blood-flow, clinical monitoring, electrochemical biosensor, electrodes, hypoxia, implantable sensor, in vivo tissue monitoring, ischemia, lactate, ph, ph sensor, rabbits, responses, vitro, Clinical monitoring, Dual signal outputs, Hypoxia, Implantable sensor, In vivo tissue monitoring, Ischemia, Ph sensor


Olmo C, Franco L, Vidal A, Del Valle LJ, Puiggalí J, (2021). Ultrasound micromolding of porous polylactide/hydroxyapatite scaffolds Express Polymer Letters 15, 389-403

© BME-PT. Ultrasound micromolding (USM) preparation of hybrid scaffolds based on polylactide (PLA) and hydroxyapatite (HAp) particles has been evaluated. PLA was stable under the applied ultrasound source since a minimum degradation was detected. Porous materials were achieved using polyethylene glycol (PEG) and NaCl salts to the initial PLA and the subsequent leaching of the micromolded specimens. To avoid cavitation and decomposition problems during micromolding, it was necessary to use HAp free of typical synthesis impurities like carbonate and nitrate compounds. Compact PLA/HAp pieces allowed a maximum HAp load of 60 wt%, while porous specimens could be obtained with a maximum load of 38 wt%. Physical characterization of new scaffolds was performed by X-ray diffraction, spectroscopic and calorimetric techniques, stress-strain tests and contact angle measurements. Results indicated that a degree of porosity of 35% and relatively good mechanical properties could be achieved (i.e., 580 MPa, 4%, and 15.6 MPa for the Young modulus, elongation at break, and tensile strength, respectively). Scaffolds showed the positive effect of HAp and porosity on cell proliferation; this latter was 40% higher than that detected for non-porous PLA specimens.

JTD Keywords: apatite, conformation, fabrication, hydroxyapatite, micropieces, polymers, porous scaffolds, proliferation, tissue, ultrasound micromolding, vibration, Composite scaffolds, Hydroxyapatite, Micropieces, Porous scaffolds, Processing technologies, Ultrasound micromolding


Guix M, Mestre R, Patiño T, de Corato M, Fuentes J, Zarpellon G, Sánchez S, (2021). Biohybrid soft robots with self-stimulating skeletons Science Robotics 6, eabe7577

Bioinspired hybrid soft robots that combine living and synthetic components are an emerging field in the development of advanced actuators and other robotic platforms (i.e., swimmers, crawlers, and walkers). The integration of biological components offers unique characteristics that artificial materials cannot precisely replicate, such as adaptability and response to external stimuli. Here, we present a skeletal muscle–based swimming biobot with a three-dimensional (3D)–printed serpentine spring skeleton that provides mechanical integrity and self-stimulation during the cell maturation process. The restoring force inherent to the spring system allows a dynamic skeleton compliance upon spontaneous muscle contraction, leading to a cyclic mechanical stimulation process that improves the muscle force output without external stimuli. Optimization of the 3D-printed skeletons is carried out by studying the geometrical stiffnesses of different designs via finite element analysis. Upon electrical actuation of the muscle tissue, two types of motion mechanisms are experimentally observed: directional swimming when the biobot is at the liquid-air interface and coasting motion when it is near the bottom surface. The integrated compliant skeleton provides both the mechanical self-stimulation and the required asymmetry for directional motion, displaying its maximum velocity at 5 hertz (800 micrometers per second, 3 body lengths per second). This skeletal muscle–based biohybrid swimmer attains speeds comparable with those of cardiac-based biohybrid robots and outperforms other muscle-based swimmers. The integration of serpentine-like structures in hybrid robotic systems allows self-stimulation processes that could lead to higher force outputs in current and future biomimetic robotic platforms. Copyright © 2021 The Authors, some rights reserved;

JTD Keywords: actuators, design, fabrication, mechanics, mems, myotubes, platform, tissue, 3d printers, Agricultural robots, Biological components, Biomimetic processes, Electrical actuation, Geometrical stiffness, Intelligent robots, Liquefied gases, Liquid-air interface, Mechanical integrity, Mechanical stimulation, Muscle, Muscle contractions, Phase interfaces, Robotics, Serpentine, Springs (components), Threedimensional (3-d)


Zeinali R, Del Valle LJ, Torras J, Puiggalí J, (2021). Recent progress on biodegradable tissue engineering scaffolds prepared by thermally-induced phase separation (Tips) International Journal Of Molecular Sciences 22, 3504

Porous biodegradable scaffolds provide a physical substrate for cells allowing them to attach, proliferate and guide the formation of new tissues. A variety of techniques have been developed to fabricate tissue engineering (TE) scaffolds, among them the most relevant is the thermally-induced phase separation (TIPS). This technique has been widely used in recent years to fabricate three-dimensional (3D) TE scaffolds. Low production cost, simple experimental procedure and easy processability together with the capability to produce highly porous scaffolds with controllable architecture justify the popularity of TIPS. This paper provides a general overview of the TIPS methodology applied for the preparation of 3D porous TE scaffolds. The recent advances in the fabrication of porous scaffolds through this technique, in terms of technology and material selection, have been reviewed. In addition, how properties can be effectively modified to serve as ideal substrates for specific target cells has been specifically addressed. Additionally, examples are offered with re-spect to changes of TIPS procedure parameters, the combination of TIPS with other techniques and innovations in polymer or filler selection.

JTD Keywords: biodegradable polymer, composites, morphology, pore structure, porosity, processing parameters, thermally induced phase separation, Biodegradable polymer, Composites, Morphology, Pore structure, Porosity, Processing parameters, Thermally induced phase separation, Tissue engineering scaffold


Vera D, García-Díaz M, Torras N, Álvarez M, Villa R, Martinez E, (2021). Engineering Tissue Barrier Models on Hydrogel Microfluidic Platforms Acs Applied Materials & Interfaces 13, 13920-13933

Tissue barriers play a crucial role in human physiology by establishing tissue compartmentalization and regulating organ homeostasis. At the interface between the extracellular matrix (ECM) and flowing fluids, epithelial and endothelial barriers are responsible for solute and gas exchange. In the past decade, microfluidic technologies and organ-on-chip devices became popular as in vitro models able to recapitulate these biological barriers. However, in conventional microfluidic devices, cell barriers are primarily grown on hard polymeric membranes within polydimethylsiloxane (PDMS) channels that do not mimic the cell-ECM interactions nor allow the incorporation of other cellular compartments such as stromal tissue or vascular structures. To develop models that accurately account for the different cellular and acellular compartments of tissue barriers, researchers have integrated hydrogels into microfluidic setups for tissue barrier-on-chips, either as cell substrates inside the chip, or as self-contained devices. These biomaterials provide the soft mechanical properties of tissue barriers and allow the embedding of stromal cells. Combining hydrogels with microfluidics technology provides unique opportunities to better recreate in vitro the tissue barrier models including the cellular components and the functionality of the in vivo tissues. Such platforms have the potential of greatly improving the predictive capacities of the in vitro systems in applications such as drug development, or disease modeling. Nevertheless, their development is not without challenges in their microfabrication. In this review, we will discuss the recent advances driving the fabrication of hydrogel microfluidic platforms and their applications in multiple tissue barrier models.

JTD Keywords: hydrogel, microfabrication, microfluidics, organ-on-chip, tissue barrier, Hydrogel, Microfabrication, Microfluidics, Organ-on-chip, Tissue barrier


Rubi-Sans, G, Cano-Torres, I, Perez-Amodio, S, Blanco-Fernandez, B, Mateos-Timoneda, MA, Engel, E, (2021). Development and Angiogenic Potential of Cell-Derived Microtissues Using Microcarrier-Template Biomedicines 9, 232

Tissue engineering and regenerative medicine approaches use biomaterials in combination with cells to regenerate lost functions of tissues and organs to prevent organ transplantation. However, most of the current strategies fail in mimicking the tissue's extracellular matrix properties. In order to mimic native tissue conditions, we developed cell-derived matrix (CDM) microtissues (MT). Our methodology uses poly-lactic acid (PLA) and Cultispher(R) S microcarriers' (MCs') as scaffold templates, which are seeded with rat bone marrow mesenchymal stem cells (rBM-MSCs). The scaffold template allows cells to generate an extracellular matrix, which is then extracted for downstream use. The newly formed CDM provides cells with a complex physical (MT architecture) and biochemical (deposited ECM proteins) environment, also showing spontaneous angiogenic potential. Our results suggest that MTs generated from the combination of these two MCs (mixed MTs) are excellent candidates for tissue vascularization. Overall, this study provides a methodology for in-house fabrication of microtissues with angiogenic potential for downstream use in various tissue regenerative strategies.

JTD Keywords: angiogenesis, cell-derived matrix, cultispher® s, microtissue, poly-lactic acid microcarriers, Angiogenesis, Cell-derived matrix, Cultispher (r) s, Microtissue, Poly-lactic acid microcarriers, Rat bone marrow mesenchymal stem cells


Puiggalí-Jou A, Ordoño J, del Valle LJ, Pérez-Amodio S, Engel E, Alemán C, (2021). Tuning multilayered polymeric self-standing films for controlled release of L-lactate by electrical stimulation Journal Of Controlled Release 330, 669-683

© 2020 Elsevier B.V. We examine different approaches for the controlled release of L-lactate, which is a signaling molecule that participates in tissue remodeling and regeneration, such as cardiac and muscle tissue. Robust, flexible, and self-supported 3-layers films made of two spin-coated poly(lactic acid) (PLA) layers separated by an electropolymerized poly(3,4-ethylenedioxythiophene) (PEDOT) layer, are used as loading and delivery systems. Films with outer layers prepared using homochiral PLA and with nanoperforations of diameter 146 ± 70 experience more bulk erosion, which also contributes to the release of L-lactic acid, than those obtained using heterochiral PLA and with nanoperforations of diameter 66 ± 24. Moreover, the release of L-lactic acid as degradation product is accelerated by applying biphasic electrical pulses. The four approaches used for loading extra L-lactate in the 3-layered films were: incorporation of L-lactate at the intermediate PEDOT layer as primary dopant agent using (1) organic or (2) basic water solutions as reaction media; (3) substitution at the PEDOT layer of the ClO4− dopant by L-lactate using de-doping and re-doping processes; and (4) loading of L-lactate at the outer PLA layers during the spin-coating process. Electrical stimuli were applied considering biphasic voltage pulses and constant voltages (both negative and positive). Results indicate that the approach used to load the L-lactate has a very significant influence in the release regulation process, affecting the concentration of released L-lactate up to two orders of magnitude. Among the tested approaches, the one based on the utilization of the outer layers for loading, approach (4), can be proposed for situations requiring prolonged and sustained L-lactate release over time. The biocompatibility and suitability of the engineered films for cardiac tissue engineering has also been confirmed using cardiac cells.

JTD Keywords: biphasic voltage pulse, cardiac tissue regeneration, cardiomyocytes proliferation, conducting polymer, nanoperforated films, sustained delivery, Biphasic voltage pulse, Cardiac tissue regeneration, Cardiomyocytes proliferation, Conducting polymer, Nanoperforated films, Sustained delivery


Garreta, Elena, Kamm, Roger D., Chuva de Sousa Lopes, Susana M., Lancaster, Madeline A., Weiss, Ron, Trepat, Xavier, Hyun, Insoo, Montserrat, Nuria, (2021). Rethinking organoid technology through bioengineering Nature Materials 20, 145-155

In recent years considerable progress has been made in the development of faithful procedures for the differentiation of human pluripotent stem cells (hPSCs). An important step in this direction has also been the derivation of organoids. This technology generally relies on traditional three-dimensional culture techniques that exploit cell-autonomous self-organization responses of hPSCs with minimal control over the external inputs supplied to the system. The convergence of stem cell biology and bioengineering offers the possibility to provide these stimuli in a controlled fashion, resulting in the development of naturally inspired approaches to overcome major limitations of this nascent technology. Based on the current developments, we emphasize the achievements and ongoing challenges of bringing together hPSC organoid differentiation, bioengineering and ethics. This Review underlines the need for providing engineering solutions to gain control of self-organization and functionality of hPSC-derived organoids. We expect that this knowledge will guide the community to generate higher-grade hPSC-derived organoids for further applications in developmental biology, drug screening, disease modelling and personalized medicine. This Review provides an overview of bioengineering technologies that can be harnessed to facilitate the culture, self-organization and functionality of human pluripotent stem cell-derived organoids.

JTD Keywords: Differentiation, Embryonic-tissues, Extracellular-matrix, In-vitro, Kidney organoids, Model, Neural-tube, Pluripotent stem-cells, Reconstitution, Self-organization


Blanco-Fernandez B, Cano-Torres I, Garrido C, Rubi-Sans G, Sanchez-Cid L, Guerra-Rebollo M, Rubio N, Blanco J, Perez-Amodio S, Mateos-Timoneda MA, Engel E, (2021). Engineered microtissues for the bystander therapy against cancer Materials Science & Engineering C-Materials For Biological Applications 121, 111854

© 2021 Elsevier B.V. Thymidine kinase expressing human adipose mesenchymal stem cells (TK-hAMSCs) in combination with ganciclovir (GCV) are an effective platform for antitumor bystander therapy in mice models. However, this strategy requires multiple TK-hAMSCs administrations and a substantial number of cells. Therefore, for clinical translation, it is necessary to find a biocompatible scaffold providing TK-hAMSCs retention in the implantation site against their rapid wash-out. We have developed a microtissue (MT) composed by TKhAMSCs and a scaffold made of polylactic acid microparticles and cell-derived extracellular matrix deposited by hAMSCs. The efficacy of these MTs as vehicles for TK-hAMSCs/GCV bystander therapy was evaluated in a rodent model of human prostate cancer. Subcutaneously implanted MTs were integrated in the surrounding tissue, allowing neovascularization and maintenance of TK-hAMSCs viability. Furthermore, MTs implanted beside tumors allowed TK-hAMSCs migration towards tumor cells and, after GCV administration, inhibited tumor growth. These results indicate that TK-hAMSCs-MTs are promising cell reservoirs for clinical use of therapeutic MSCs in bystander therapies.

JTD Keywords: adipose mesenchymal stem cells, bioluminescence, bystander therapy, cancer, Adipose mesenchymal stem cells, Bioluminescence, Bystander therapy, Cancer, Self-assembled cell-based microtissues


Fernández-Costa JM, Fernández-Garibay X, Velasco-Mallorquí F, Ramón-Azcón J, (2021). Bioengineered in vitro skeletal muscles as new tools for muscular dystrophies preclinical studies Journal Of Tissue Engineering 12, 2041731420981339

© The Author(s) 2021. Muscular dystrophies are a group of highly disabling disorders that share degenerative muscle weakness and wasting as common symptoms. To date, there is not an effective cure for these diseases. In the last years, bioengineered tissues have emerged as powerful tools for preclinical studies. In this review, we summarize the recent technological advances in skeletal muscle tissue engineering. We identify several ground-breaking techniques to fabricate in vitro bioartificial muscles. Accumulating evidence shows that scaffold-based tissue engineering provides topographical cues that enhance the viability and maturation of skeletal muscle. Functional bioartificial muscles have been developed using human myoblasts. These tissues accurately responded to electrical and biological stimulation. Moreover, advanced drug screening tools can be fabricated integrating these tissues in electrical stimulation platforms. However, more work introducing patient-derived cells and integrating these tissues in microdevices is needed to promote the clinical translation of bioengineered skeletal muscle as preclinical tools for muscular dystrophies.

JTD Keywords: biomaterials, drug screening platforms, muscular dystrophy, skeletal muscle, tissue engineering, Biomaterials, Drug screening platforms, Muscular dystrophy, Skeletal muscle, Tissue engineering


Moya-Andérico L, Admella J, Torrents E, (2021). A clearing protocol for Galleria mellonella larvae: Visualization of internalized fluorescent nanoparticles New Biotechnology 60, 20-26

© 2020 Elsevier B.V. Light scattering is a challenge for imaging three-dimensional organisms. A number of new tissue clearing methodologies have been described in recent years, increasing the utilities of clearing techniques to obtain transparent samples. Here, we describe the optimization of a suitable and novel protocol for clearing Galleria mellonella larvae, an alternative infection animal model with a promising potential for the toxicological evaluation of different molecules and materials. This has allowed the visualization of internalised fluorescent nanoparticles using confocal microscopy, opening the door to a wide range of different applications.

JTD Keywords: galleria mellonella, nanotoxicology, Galleria mellonella, Nanotoxicology, Tissue clearance


Ben Hamouda S, Vargas A, Boivin R, Miglino MA, da Palma RK, Lavoie JP, (2021). Recellularization of Bronchial Extracellular Matrix With Primary Bronchial Smooth Muscle Cells Journal Of Equine Veterinary Science 96, 103313

© 2020 Elsevier Inc. Severe asthma is associated with an increased airway smooth muscle (ASM) mass and altered composition of the extracellular matrix (ECM). Studies have indicated that ECM-ASM cell interactions contribute to this remodeling and its limited reversibility with current therapy. Three-dimensional matrices allow the study of complex cellular responses to different stimuli in an almost natural environment. Our goal was to obtain acellular bronchial matrices and then develop a recellularization protocol with ASM cells. We studied equine bronchi as horses spontaneously develop a human asthma-like disease. The bronchi were decellularized using Triton/Sodium Deoxycholate. The obtained scaffolds retained their anatomical and histological properties. Using immunohistochemistry and a semi-quantitative score to compare native bronchi to scaffolds revealed no significant variation for matrixial proteins. DNA quantification and electrophoresis revealed that most DNA was 29.6 ng/mg of tissue ± 5.6, with remaining fragments of less than 100 bp. Primary ASM cells were seeded on the scaffolds. Histological analysis of the recellularizations showed that ASM cells migrated and proliferated primarily in the decellularized smooth muscle matrix, suggesting a chemotactic effect of the scaffolds. This is the first report of primary ASM cells preferentially repopulating the smooth muscle matrix layer in bronchial matrices. This protocol is now being used to study the molecular interactions occurring between the asthmatic ECMs and ASM to identify effectors of asthmatic bronchial remodeling.

JTD Keywords: 2d, airway smooth muscle cells, asthma, decellularization, disease, elastin, extracellular matrix, lung scaffolds, migration, peptide, recellularization, tissues, Airway smooth muscle cells, Asthma, Culture-systems, Decellularization, Extracellular matrix, Recellularization


Selfa IL, Gallo M, Montserrat N, Garreta E, (2021). Directed Differentiation of Human Pluripotent Stem Cells for the Generation of High-Order Kidney Organoids Methods In Molecular Biology 2258, 171-192

© 2021, The Editor(s) (if applicable) and The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature. Our understanding in the inherent properties of human pluripotent stem cells (hPSCs) have made possible the development of differentiation procedures to generate three-dimensional tissue-like cultures, so-called organoids. Here we detail a stepwise methodology to generate kidney organoids from hPSCs. This is achieved through direct differentiation of hPSCs in two-dimensional monolayer culture toward the posterior primitive streak fate, followed by induction of intermediate mesoderm-committed cells, which are further aggregated and cultured in three-dimensions to generate kidney organoids containing segmented nephron-like structures in a process that lasts 20 days. We also provide a concise description on how to assess renal commitment during the time course of kidney organoid generation. This includes the use of flow cytometry and immunocytochemistry analyses for the detection of specific renal differentiation markers.

JTD Keywords: 2d monolayer, 3d organotypic culture, differentiation, flow cytometry, human pluripotent stem cells, immunocytochemistry, intermediate mesoderm, kidney organoid, nephron progenitor cells, nephrons, primitive streak, 2d monolayer, 3d organotypic culture, Differentiation, Flow cytometry, Human pluripotent stem cells, Immunocytochemistry, Intermediate mesoderm, Kidney organoid, Nephron progenitor cells, Nephrons, Primitive streak, Tissue


Watt, AC, Cejas, P, DeCristo, MJ, Metzger, O, Lam, EYN, Qiu, XT, BrinJones, H, Kesten, N, Coulson, R, Font-Tello, A, Lim, K, Vadhi, R, Daniels, VW, Montero, J, Taing, L, Meyer, CA, Gilan, O, Bell, CC, Korthauer, KD, Giambartolomei, C, Pasaniuc, B, Seo, JH, Freedman, ML, Ma, CT, Ellis, MJ, Krop, I, Winer, E, Letai, A, Brown, M, Dawson, MA, Long, HW, Zhao, JJ, Goel, S, (2021). CDK4/6 inhibition reprograms the breast cancer enhancer landscape by stimulating AP-1 transcriptional activity Nature Cancer 2, 34-48

Goel and colleagues show that CDK4/6 inhibition induces global chromatin changes mediated by AP-1 factors, which mediate key biological and clinical effects in breast cancer. Pharmacologic inhibitors of cyclin-dependent kinases 4 and 6 (CDK4/6) were designed to induce cancer cell cycle arrest. Recent studies have suggested that these agents also exert other effects, influencing cancer cell immunogenicity, apoptotic responses and differentiation. Using cell-based and mouse models of breast cancer together with clinical specimens, we show that CDK4/6 inhibitors induce remodeling of cancer cell chromatin characterized by widespread enhancer activation, and that this explains many of these effects. The newly activated enhancers include classical super-enhancers that drive luminal differentiation and apoptotic evasion, as well as a set of enhancers overlying endogenous retroviral elements that are enriched for proximity to interferon-driven genes. Mechanistically, CDK4/6 inhibition increases the level of several activator protein-1 transcription factor proteins, which are in turn implicated in the activity of many of the new enhancers. Our findings offer insights into CDK4/6 pathway biology and should inform the future development of CDK4/6 inhibitors.

JTD Keywords: Abemaciclib, Androgen receptor, Animal experiment, Animal model, Animal tissue, Apoptosis, Article, Breast cancer, C-jun, Cancer cell, Carcinoembryonic antigen related cell adhesion molecule 1, Caspase 3, Cell cycle arrest, Cells, Chromatin, Chromatin immunoprecipitation, Controlled study, Cyclin dependent kinase 4, Cyclin dependent kinase 6, Dna damage, Epidermal growth factor receptor 2, Estrogen receptor, Female, Flow cytometry, Fulvestrant, Hla drb1 antigen, Human, Human cell, Immunoblotting, Immunogenicity, Immunoprecipitation, Interferon, Luciferase assay, Mcf-7 cell line, Mda-mb-231 cell line, Microarray analysis, Morphogenesis, Mouse, Nonhuman, Palbociclib, Protein, Protein expression, Rb, Resistance, Rna polymerase ii, Rna sequence, Selective-inhibition, Senescence, Short tandem repeat, Signal transduction, Tamoxifen, Transcription elongation, Transcription factor, Transcription factor ap 1, Transcriptome, Tumor biopsy, Tumor differentiation, Tumor spheroid, Tumor xenograft, Vinculin, Whole exome sequencing


Park, D., Wershof, E., Boeing, S., Labernadie, A., Jenkins, R. P., George, S., Trepat, X., Bates, P. A., Sahai, E., (2020). Extracellular matrix anisotropy is determined by TFAP2C-dependent regulation of cell collisions Nature Materials 19, 227-238

The isotropic or anisotropic organization of biological extracellular matrices has important consequences for tissue function. We study emergent anisotropy using fibroblasts that generate varying degrees of matrix alignment from uniform starting conditions. This reveals that the early migratory paths of fibroblasts are correlated with subsequent matrix organization. Combined experimentation and adaptation of Vicsek modelling demonstrates that the reorientation of cells relative to each other following collision plays a role in generating matrix anisotropy. We term this behaviour ‘cell collision guidance’. The transcription factor TFAP2C regulates cell collision guidance in part by controlling the expression of RND3. RND3 localizes to cell–cell collision zones where it downregulates actomyosin activity. Cell collision guidance fails without this mechanism in place, leading to isotropic matrix generation. The cross-referencing of alignment and TFAP2C gene expression signatures against existing datasets enables the identification and validation of several classes of pharmacological agents that disrupt matrix anisotropy.

JTD Keywords: Biomaterials – cells, Cell migration, Self-assembly, Tissues


Prat-Vidal, C., Rodríguez-Gómez, L., Aylagas, M., Nieto-Nicolau, N., Gastelurrutia, P., Agustí, E., Gálvez-Montón, C., Jorba, I., Teis, A., Monguió-Tortajada, M., Roura, S., Vives, J., Torrents-Zapata, S., Coca, M. I., Reales, L., Cámara-Rosell, M. L., Cediel, G., Coll, R., Farré, R., Navajas, D., Vilarrodona, A., García-López, J., Muñoz-Guijosa, C., Querol, S., Bayes-Genis, A., (2020). First-in-human PeriCord cardiac bioimplant: Scalability and GMP manufacturing of an allogeneic engineered tissue graft EBioMedicine 54, 102729

Background Small cardiac tissue engineering constructs show promise for limiting post-infarct sequelae in animal models. This study sought to scale-up a 2-cm2 preclinical construct into a human-size advanced therapy medicinal product (ATMP; PeriCord), and to test it in a first-in-human implantation. Methods The PeriCord is a clinical-size (12–16 cm2) decellularised pericardial matrix colonised with human viable Wharton's jelly-derived mesenchymal stromal cells (WJ-MSCs). WJ-MSCs expanded following good manufacturing practices (GMP) met safety and quality standards regarding the number of cumulative population doublings, genomic stability, and sterility. Human decellularised pericardial scaffolds were tested for DNA content, matrix stiffness, pore size, and absence of microbiological growth. Findings PeriCord implantation was surgically performed on a large non-revascularisable scar in the inferior wall of a 63-year-old male patient. Coronary artery bypass grafting was concomitantly performed in the non-infarcted area. At implantation, the 16-cm2 pericardial scaffold contained 12·5 × 106 viable WJ-MSCs (85·4% cell viability; <0·51 endotoxin units (EU)/mL). Intraoperative PeriCord delivery was expeditious, and secured with surgical glue. The post-operative course showed non-adverse reaction to the PeriCord, without requiring host immunosuppression. The three-month clinical follow-up was uneventful, and three-month cardiac magnetic resonance imaging showed ~9% reduction in scar mass in the treated area. Interpretation This preliminary report describes the development of a scalable clinical-size allogeneic PeriCord cardiac bioimplant, and its first-in-human implantation. Funding La Marató de TV3 Foundation, Government of Catalonia, Catalan Society of Cardiology, “La Caixa” Banking Foundation, Spanish Ministry of Science, Innovation and Universities, Institute of Health Carlos III, and the European Regional Development Fund.

JTD Keywords: Advanced therapy medicinal product (ATMP), Biofabrication, Cardiac tissue engineering, Myocardial infarction, Scaffold, Wharton's jelly-derived mesenchymal stromal cells (WJ-MSCs)


Altay, Gizem, Batlle, Eduard, Fernández-Majada, Vanesa, Martínez, Elena, (2020). In vitro self-organized mouse small intestinal epithelial monolayer protocol Bio-protocol 10, (3), e3514

Developing protocols to obtain intestinal epithelial monolayers that recapitulate in vivo physiology to overcome the limitations of the organoids’ closed geometry has become of great interest during the last few years. Most of the developed culture models showed physiological-relevant cell composition but did not prove self-renewing capacities. Here, we show a simple method to obtain mouse small intestine-derived epithelial monolayers organized into proliferative crypt-like domains, containing stem cells, and differentiated villus-like regions, closely resembling the in vivo cell composition and distribution. In addition, we adapted our model to a tissue culture format compatible with functional studies and prove close to physiological barrier properties of our in vitro epithelial monolayers. Thus, we have set-up a protocol to generate physiologically relevant intestinal epithelial monolayers to be employed in assays where independent access to both luminal and basolateral compartments is needed, such as drug absorption, intracellular trafficking and microbiome-epithelium interaction assays.

JTD Keywords: Mouse intestinal organoids, Adult intestinal stem cells, Matrigel, Intestinal epithelial monolayer, In vitro intestinal epithelial model, Tissue-like functionality, TEER


Alert, R., Trepat, X., (2020). Physical models of collective cell migration Annual Review of Condensed Matter Physics 11, 77-101

Collective cell migration is a key driver of embryonic development, wound healing, and some types of cancer invasion. Here, we provide a physical perspective of the mechanisms underlying collective cell migration. We begin with a catalog of the cell-cell and cell-substrate interactions that govern cell migration, which we classify into positional and orientational interactions. We then review the physical models that have been developed to explain how these interactions give rise to collective cellular movement. These models span the subcellular to the supracellular scales, and they include lattice models, phase-field models, active network models, particle models, and continuum models. For each type of model, we discuss its formulation, its limitations, and the main emergent phenomena that it has successfully explained. These phenomena include flocking and fluid-solid transitions, as well as wetting, fingering, and mechanical waves in spreading epithelial monolayers. We close by outlining remaining challenges and future directions in the physics of collective cell migration.

JTD Keywords: Active network models, Cellular Potts models, Continuum models, Particle models, Phase-field models, Tissue biophysics


Otero, J., Navajas, D., Alcaraz, J., (2020). Characterization of the elastic properties of extracellular matrix models by atomic force microscopy Methods in Cell Biology (ed. Caballero, David, Kundu, Subhas C., Reis, Rui L.), Academic Press (Cambridge, USA) 156, 59-83

Tissue elasticity is a critical regulator of cell behavior in normal and diseased conditions like fibrosis and cancer. Since the extracellular matrix (ECM) is a major regulator of tissue elasticity and function, several ECM-based models have emerged in the last decades, including in vitro endogenous ECM, decellularized tissue ECM and ECM hydrogels. The development of such models has urged the need to quantify their elastic properties particularly at the nanometer scale, which is the relevant length scale for cell-ECM interactions. For this purpose, the versatility of atomic force microscopy (AFM) to quantify the nanomechanical properties of soft biomaterials like ECM models has emerged as a very suitable technique. In this chapter we provide a detailed protocol on how to assess the Young's elastic modulus of ECM models by AFM, discuss some of the critical issues, and provide troubleshooting guidelines as well as illustrative examples of AFM measurements, particularly in the context of cancer.

JTD Keywords: 3D ECM hydrogels, Atomic force microscopy, Decellularized tissue, Elastic modulus, Endogenous ECM, Extracellular matrix


Hernández-Albors, Alejandro, Castaño, Albert G., Fernández-Garibay, Xiomara, Ortega, María Alejandra, Balaguer, Jordina, Ramón-Azcón, Javier, (2019). Microphysiological sensing platform for an in-situ detection of tissue-secreted cytokines Biosensors and Bioelectronics: X 2, 100025

Understanding the protein-secretion dynamics from single, specific tissues is critical toward the advancement of disease detection and treatments. However, such secretion dynamics remain difficult to measure in vivo due to the uncontrolled contributions from other tissue populations. Here, we describe an integrated platform designed for the reliable, near real-time measurements of cytokines secreted from an in vitro single-tissue model. In our setup, we grow 3D biomimetic tissues to discretize cytokine source, and we separate them from a magnetic microbead-based biosensing system using a Transwell insert. This design integrates physiochemically controlled biological activity, high-sensitivity protein detection (LOD < 20 pg mL−1), and rapid protein diffusion to enable non-invasive, near real-time measurements. To showcase the specificity and sensitivity of the system, we use our setup to probe the inflammatory process related to the protein Interleukine 6 (IL-6) and to the Tumor Necrosis Factor (TNF-α). We show that our setup can monitor the time-dependence profile of IL-6 and TNF-α secretion that results from the electrical and chemical stimulation of 3D skeletal muscle tissues. We demonstrate a novel and affordable methodology for discretizing the secretion kinetics of specific tissues for advancing metabolic-disorder studies and drug-screening applications.

JTD Keywords: Microphysiological tissues, Tissue engineering, Electrochemical, biosensors, Magnetic particles, Skeletal muscle, Electric stimulation


Valls-Margarit, M., Iglesias-García, O., Di Guglielmo, C., Sarlabous, L., Tadevosyan, K., Paoli, R., Comelles, J., Blanco-Almazán, D., Jiménez-Delgado, S., Castillo-Fernández, O., Samitier, J., Jané, R., Martínez, Elena, Raya, Á., (2019). Engineered macroscale cardiac constructs elicit human myocardial tissue-like functionality Stem Cell Reports 13, (1), 207-220

In vitro surrogate models of human cardiac tissue hold great promise in disease modeling, cardiotoxicity testing, and future applications in regenerative medicine. However, the generation of engineered human cardiac constructs with tissue-like functionality is currently thwarted by difficulties in achieving efficient maturation at the cellular and/or tissular level. Here, we report on the design and implementation of a platform for the production of engineered cardiac macrotissues from human pluripotent stem cells (PSCs), which we term “CardioSlice.” PSC-derived cardiomyocytes, together with human fibroblasts, are seeded into large 3D porous scaffolds and cultured using a parallelized perfusion bioreactor with custom-made culture chambers. Continuous electrical stimulation for 2 weeks promotes cardiomyocyte alignment and synchronization, and the emergence of cardiac tissue-like properties. These include electrocardiogram-like signals that can be readily measured on the surface of CardioSlice constructs, and a response to proarrhythmic drugs that is predictive of their effect in human patients.

JTD Keywords: Cardiac tissue engineering, CardioSlice, ECG-like signals, Electrical stimulation, Heart physiology, Human induced pluripotent stem cells, Perfusion bioreactor, Tissue-like properties


de la Mata, Ana, Mateos-Timoneda, Miguel A., Nieto-Miguel, Teresa, Galindo, Sara, López-Paniagua, Marina, Planell, Josep A., Engel, Elisabeth, Calonge, Margarita, (2019). Poly-l/dl-lactic acid films functionalized with collagen IV as carrier substrata for corneal epithelial stem cells Colloids and Surfaces B: Biointerfaces 177, 121-129

Limbal epithelial stem cells (LESCs) are responsible for the renewal of corneal epithelium. Cultivated limbal epithelial transplantation is the current treatment of choice for restoring the loss or dysfunction of LESCs. To perform this procedure, a substratum is necessary for in vitro culturing of limbal epithelial cells and their subsequent transplantation onto the ocular surface. In this work, we evaluated poly-L/DL-lactic acid 70:30 (PLA) films functionalized with type IV collagen (col IV) as potential in vitro carrier substrata for LESCs. We first demonstrated that PLA-col IV films were biocompatible and suitable for the proliferation of human corneal epithelial cells. Subsequently, limbal epithelial cell suspensions, isolated from human limbal rings, were cultivated using culture medium that did not contain animal components. The cells adhered significantly faster to PLA-col IV films than to tissue culture plastic (TCP). The mRNA expression levels for the LESC specific markers, K15, P63α and ABCG2 were similar or greater (significantly in the case of K15) in limbal epithelial cells cultured on PLA-col IV films than limbal epithelial cells cultured on TCP. The percentage of cells expressing the corneal (K3, K12) and the LESC (P63α, ABCG2) specific markers was similar for both substrata. These results suggest that the PLA-col IV films promoted LESC attachment and helped to maintain their undifferentiated stem cell phenotype. Consequently, these substrata offer an alternative for the transplantation of limbal cells onto the ocular surface.

JTD Keywords: Corneal epithelium, Collagen IV, Limbal stem cells, Polylactic acid, Tissue engineering


Cofiño, C., Perez-Amodio, S., Semino, C. E., Engel, E., Mateos-Timoneda, M. A., (2019). Development of a self-assembled peptide/methylcellulose-based bioink for 3D bioprinting Macromolecular Materials and Engineering 304, (11), 1900353

The introduction of 3D bioprinting to fabricate living constructs with tailored architecture has provided a new paradigm for biofabrication, with the potential to overcome several drawbacks of conventional scaffold-based tissue regeneration strategies. Hydrogel-based materials are suitable candidates regarding cell biocompatibility but often display poor mechanical properties. Self-assembling peptides are a promising source of biomaterials to be used as 3D scaffolds based on their similarity to extracellular matrices (structurally and mechanically). In this study, an advanced bioink for biofabrication is presented based on the optimization of a RAD16-I-based biomaterial. The strategy followed to build 3D predefined structures by 3D printing is based on an enhancement of bioink viscosity by adding methylcellulose (MC) to a RAD16-I solution. The resultant constructs display high shape fidelity and stability and embedded human mesenchymal stem cells present high viability after 7 days of culture. Moreover, cells are also able to differentiate to the adipogenic lineage, suggesting the suitability of this novel biomaterial for soft tissue engineering applications.

JTD Keywords: 3D bioprinting, Biofabrication, Bioinks, Self-assembling peptides, Tissue engineering


Gil, V., Del Río, J. A., (2019). Generation of 3-d collagen-based hydrogels to analyze axonal growth and behavior during nervous system development Journal of Visualized Experiments , (148), e59481

This protocol uses natural type I collagen to generate three-dimensional (3-D) hydrogel for monitoring and analyzing the axonal growth. The protocol is centered on culturing small pieces of embryonic or early postnatal rodent brains inside a 3-D hydrogel formed by the rat tail tendon-derived type I collagen with specific porosity. Tissue pieces are cultured inside the hydrogel and confronted to specific brain fragments or genetically-modified cell aggregates to produce and secrete molecules suitable for creating a gradient inside the porous matrix. The steps of this protocol are simple and reproducible but include critical steps to be considered carefully during its development. Moreover, the behavior of growing axons can be monitored and analyzed directly using a phase-contrast microscope or mono/multiphoton fluorescence microscope after fixation by immunocytochemical methods.

JTD Keywords: 3-D hydrogel cultures, Axonal growth, Cell transfection, Chemoattraction, Chemorepulsion, Embryonic nervous system, Issue 148, Neuroscience, Tissue explants


Fuentes-Mera, L., Camacho, A., Engel, E., Pérez-Silos, V., Lara-Arias, J., Marino-Martínez, I., Peña-Martínez, V., (2019). Therapeutic potential of articular cartilage regeneration using tissue engineering based on multiphase designs Cartilage Tissue Engineering and Regeneration Techniques (ed. Nikolopoulos, Dimitrios D., Safos, George K., Dimitrios, Kalpaxis), IntechOpen (Budapest, Hungary) , 331-359

Articular cartilage tissue possesses poor ability to regenerate; as the lesion progresses, it extends to the underlying subchondral bone and an osteochondral (OC) defect appears complicating the therapeutic approaches. Cartilage tissue engineering has become a very active research area capable of contributing to medical technology innovation. In this regard, the development of new biomaterials in combination with cells represents one of the best alternatives for the treatment of OC injuries. In the last decades, the strategies have been designed without considering the cartilage as a complex tissue with a functionally stratified three-dimensional structure. Today, efforts are focused on creating a starting point in the process of cartilage formation with the development of a multiphase implants that recapitulates the cartilage as an OC unit, which improves its integration. This chapter will focus on a review of tissue engineering based on multiphase designs for cartilage and OC injuries, highlighting the importance of the biomaterial selection, and also the relevance of a biomimetic approach to reach a suitable microenvironment for the differentiation and maturation of the chondral tissue.

JTD Keywords: Osteochondral regeneration, Cartilage tissue engineering, Multiphasic designs, Biofunctionalization, Vascularization


Castaño, O., Pérez-Amodio, S., Navarro, C., Mateos-Timoneda, M.A., Engel, E., (2018). Instructive microenvironments in skin wound healing: Biomaterials as signal releasing platforms Advanced Drug Delivery Reviews 129, 95-117

Skin wound healing aims to repair and restore tissue through a multistage process that involves different cells and signalling molecules that regulate the cellular response and the dynamic remodelling of the extracellular matrix. Nowadays, several therapies that combine biomolecule signals (growth factors and cytokines) and cells are being proposed. However, a lack of reliable evidence of their efficacy, together with associated issues such as high costs, a lack of standardization, no scalable processes, and storage and regulatory issues, are hampering their application. In situ tissue regeneration appears to be a feasible strategy that uses the body's own capacity for regeneration by mobilizing host endogenous stem cells or tissue-specific progenitor cells to the wound site to promote repair and regeneration. The aim is to engineer instructive systems to regulate the spatio-temporal delivery of proper signalling based on the biological mechanisms of the different events that occur in the host microenvironment. This review describes the current state of the different signal cues used in wound healing and skin regeneration, and their combination with biomaterial supports to create instructive microenvironments for wound healing.

JTD Keywords: Instructive biomaterials, Skin regeneration, Wound healing, Signalling release, In situ tissue engineering


Alcaraz, J., Otero, J., Jorba, I., Navajas, D., (2018). Bidirectional mechanobiology between cells and their local extracellular matrix probed by atomic force microscopy Seminars in Cell and Developmental Biology 73, 71-81

There is growing recognition that the mechanical interactions between cells and their local extracellular matrix (ECM) are central regulators of tissue development, homeostasis, repair and disease progression. The unique ability of atomic force microscopy (AFM) to probe quantitatively mechanical properties and forces at the nanometer or micrometer scales in all kinds of biological samples has been instrumental in the recent advances in cell and tissue mechanics. In this review we illustrate how AFM has provided important insights on our current understanding of the mechanobiology of cells, ECM and cell-ECM bidirectional interactions, particularly in the context of soft acinar tissues like the mammary gland or pulmonary tissue. AFM measurements have revealed that intrinsic cell micromechanics is cell-type specific, and have underscored the prominent role of β1 integrin/FAK(Y397) signaling and the actomyosin cytoskeleton in the mechanoresponses of both parenchymal and stromal cells. Moreover AFM has unveiled that the micromechanics of the ECM obtained by tissue decellularization is unique for each anatomical compartment, which may support both its specific function and cell differentiation. AFM has also enabled identifying critical mechanoregulatory proteins involved in branching morphogenesis (MMP14) and acinar differentiation (α3β1 integrin), and has clarified the role of altered tissue mechanics and architecture in a variety of pathologic conditions. Critical technical issues of AFM mechanical measurements like tip geometry effects are also discussed.

JTD Keywords: Atomic force microscopy, Beta1 integrin, Elastic modulus, Extracellular matrix, Morphogenesis, Tissue decellularization


Torras, N., García-Díaz, M., Fernández-Majada, V., Martínez, Elena, (2018). Mimicking epithelial tissues in three-dimensional cell culture models Frontiers in Bioengineering and Biotechnology 6, Article 197

Epithelial tissues are composed of layers of tightly connected cells shaped into complex three-dimensional (3D) structures such as cysts, tubules, or invaginations. These complex 3D structures are important for organ-specific functions and often create biochemical gradients that guide cell positioning and compartmentalization within the organ. One of the main functions of epithelia is to act as physical barriers that protect the underlying tissues from external insults. In vitro, epithelial barriers are usually mimicked by oversimplified models based on cell lines grown as monolayers on flat surfaces. While useful to answer certain questions, these models cannot fully capture the in vivo organ physiology and often yield poor predictions. In order to progress further in basic and translational research, disease modeling, drug discovery, and regenerative medicine, it is essential to advance the development of new in vitro predictive models of epithelial tissues that are capable of representing the in vivo-like structures and organ functionality more accurately. Here, we review current strategies for obtaining biomimetic systems in the form of advanced in vitro models that allow for more reliable and safer preclinical tests. The current state of the art and potential applications of self-organized cell-based systems, organ-on-a-chip devices that incorporate sensors and monitoring capabilities, as well as microfabrication techniques including bioprinting and photolithography, are discussed. These techniques could be combined to help provide highly predictive drug tests for patient-specific conditions in the near future.

JTD Keywords: 3D cell culture models, Biofabrication, Disease modeling, Drug screening, Epithelial barriers, Microengineered tissues, Organ-on-a-chip, Organoids


Casanellas, Ignasi, García-Lizarribar, Andrea, Lagunas, Anna, Samitier, Josep, (2018). Producing 3D biomimetic nanomaterials for musculoskeletal system regeneration Frontiers in Bioengineering and Biotechnology 6, Article 128

The human musculoskeletal system is comprised mainly of connective tissues such as cartilage, tendon, ligaments, skeletal muscle and skeletal bone. These tissues support the structure of the body, hold and protect the organs, and are responsible of movement. Since it is subjected to continuous strain, the musculoskeletal system is prone to injury by excessive loading forces or aging, whereas currently available treatments are usually invasive and not always effective. Most of the musculoskeletal injuries require surgical intervention facing a limited post-surgery tissue regeneration, especially for widespread lesions. Therefore, many tissue engineering approaches have been developed tackling musculoskeletal tissue regeneration. Materials are designed to meet the chemical and mechanical requirements of the native tissue three-dimensional (3D) environment, thus facilitating implant integration while providing a good reabsorption rate. With biological systems operating at the nanoscale, nanoengineered materials have been developed to support and promote regeneration at the interprotein communication level. Such materials call for a great precision and architectural control in the production process fostering the development of new fabrication techniques. In this mini review, we would like to summarize the most recent advances in 3D nanoengineered biomaterials for musculoskeletal tissue regeneration, with especial emphasis on the different techniques used to produce them.

JTD Keywords: Nanofiber, 3D printing, Musculoskeletal, Regeneration, Scaffold, Tissue Engineering, Stimuli-responsive


Farré, Ramon, Otero, Jordi, Almendros, Isaac, Navajas, Daniel, (2018). Bioengineered lungs: A challenge and an opportunity Archivos de Bronconeumología 54, (1), 31-38

Lung biofabrication is a new tissue engineering and regenerative development aimed at providing organs for potential use in transplantation. Lung biofabrication is based on seeding cells into an acellular organ scaffold and on culturing them in an especial purpose bioreactor. The acellular lung scaffold is obtained by decellularizing a non-transplantable donor lung by means of conventional procedures based on application of physical, enzymatic and detergent agents. To avoid immune recipient's rejection of the transplanted bioengineered lung, autologous bone marrow/adipose tissue-derived mesenchymal stem cells, lung progenitor cells or induced pluripotent stem cells are used for biofabricating the bioengineered lung. The bioreactor applies circulatory perfusion and mechanical ventilation with physiological parameters to the lung during biofabrication. These physical stimuli to the organ are translated into the stem cell local microenvironment - e.g. shear stress and cyclic stretch - so that cells sense the physiological conditions in normally functioning mature lungs. After seminal proof of concept in a rodent model was published in 2010, the hypothesis that lungs can be biofabricated is accepted and intense research efforts are being devoted to the topic. The current experimental evidence obtained so far in animal tests and in ex vivo human bioengineered lungs suggests that the date of first clinical tests, although not immediate, is coming. Lung bioengineering is a disrupting concept that poses a challenge for improving our basic science knowledge and is also an opportunity for facilitating lung transplantation in future clinical translation.

JTD Keywords: Tissue engineering, Regenerative medicine, Lung transplantation, Lung repair, Lung regeneration


Farré, N., Jorba, I., Torres, M., Falcones, B., Martí-Almor, J., Farré, R., Almendros, I., Navajas, D., (2018). Passive stiffness of left ventricular myocardial tissue is reduced by ovariectomy in a post-menopause mouse model Frontiers in Physiology 9, Article 1545

Background: Heart failure (HF) – a very prevalent disease with high morbidity and mortality – usually presents with diastolic dysfunction. Although post-menopause women are at increased risk of HF and diastolic dysfunction, poor attention has been paid to clinically and experimentally investigate this group of patients. Specifically, whether myocardial stiffness is affected by menopause is unknown. Aim: To investigate whether loss of female sexual hormones modifies the Young’s modulus (E) of left ventricular (LV) myocardial tissue in a mouse model of menopause induced by ovariectomy (OVX). Methods: After 6 months of bilateral OVX, eight mice were sacrificed, fresh LV myocardial strips were prepared (∼8 × 1 × 1 mm), and their passive stress–stretch relationship was measured. E was computed by exponential fitting of the stress–stretch relationship. Subsequently, to assess the relative role of cellular and extracellular matrix components in determining OVX-induced changes in E, the tissues strips were decellularized and subjected to the same stretching protocol to measure E. A control group of eight sham-OVX mice was simultaneously studied. Results: E (kPa; m ± SE) in OVX mice was ∼twofold lower than in controls (11.7 ± 1.8 and 22.1 ± 4.4, respectively; p < 0.05). No significant difference between groups was found in E of the decellularized tissue (31.4 ± 12.05 and 40.9 ± 11.5, respectively; p = 0.58). Conclusion: Loss of female sexual hormones in an OVX model induces a reduction in the passive stiffness of myocardial tissue, suggesting that active relaxation should play a counterbalancing role in diastolic dysfunction in post-menopausal women with HF.

JTD Keywords: Decellularized tissue, Female hormones, Heart tissue, Ovariectomy, Stress-strain


Garreta, E., González, F., Montserrat, N., (2018). Studying kidney disease using tissue and genome engineering in human pluripotent stem cells Nephron 138, 48-59

Kidney morphogenesis and patterning have been extensively studied in animal models such as the mouse and zebrafish. These seminal studies have been key to define the molecular mechanisms underlying this complex multistep process. Based on this knowledge, the last 3 years have witnessed the development of a cohort of protocols allowing efficient differentiation of human pluripotent stem cells (hPSCs) towards defined kidney progenitor populations using two-dimensional (2D) culture systems or through generating organoids. Kidney organoids are three-dimensional (3D) kidney-like tissues, which are able to partially recapitulate kidney structure and function in vitro. The current possibility to combine state-of-the art tissue engineering with clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated systems 9 (Cas9)-mediated genome engineering provides an unprecedented opportunity for studying kidney disease with hPSCs. Recently, hPSCs with genetic mutations introduced through CRISPR/Cas9-mediated genome engineering have shown to produce kidney organoids able to recapitulate phenotypes of polycystic kidney disease and glomerulopathies. This mini review provides an overview of the most recent advances in differentiation of hPSCs into kidney lineages, and the latest implementation of the CRISPR/Cas9 technology in the organoid setting, as promising platforms to study human kidney development and disease.

JTD Keywords: Clustered regularly interspaced short palindromic repeats/CRISPR-associated systems 9, Disease modeling, Gene editing, Human pluripotent stem cells, Kidney genetics, Tissue engineering


Gugutkov, D., Gustavsson, J., Cantini, M., Salmeron-Sánchez, M., Altankov, G., (2017). Electrospun fibrinogen-PLA nanofibres for vascular tissue engineering Journal of Tissue Engineering and Regenerative Medicine 11, (10), 2774-2784

Here we report on the development of a new type of hybrid fibrinogen-polylactic acid (FBG-PLA) nanofibres (NFs) with improved stiffness, combining the good mechanical properties of PLA with the excellent cell recognition properties of native FBG. We were particularly interested in the dorsal and ventral cell response to the nanofibres' organization (random or aligned), using human umbilical endothelial cells (HUVECs) as a model system. Upon ventral contact with random NFs, the cells developed a stellate-like morphology with multiple projections. The well-developed focal adhesion complexes suggested a successful cellular interaction. However, time-lapse analysis shows significantly lowered cell movements, resulting in the cells traversing a relatively short distance in multiple directions. Conversely, an elongated cell shape and significantly increased cell mobility were observed in aligned NFs. To follow the dorsal cell response, artificial wounds were created on confluent cell layers previously grown on glass slides and covered with either random or aligned NFs. Time-lapse analysis showed significantly faster wound coverage (within 12 h) of HUVECs on aligned samples vs. almost absent directional migration on random ones. However, nitric oxide (NO) release shows that endothelial cells possess lowered functionality on aligned NFs compared to random ones, where significantly higher NO production was found. Collectively, our studies show that randomly organized NFs could support the endothelization of implants while aligned NFs would rather direct cell locomotion for guided neovascularization.

JTD Keywords: Electrospun nanofibers, Endothelial cells, Fibrinogen, Guided cellular behavior, Polylactic acid, Vascular tissue engineering


Planell, J. A., Navarro, M., Engel, E., (2017). Developing targeted biocomposites in tissue engineering and regenerative medicine Biomedical Composites (ed. Ambrosio, L.), Woodhead Publishing (Duxfor, UK) Biomaterials, 569-587

Regenerative medicine is a relatively new field with new requirements for smart materials, where composites will have a strong role to play. The new paradigm of regenerative medicine and tissue engineering requires biomaterials with high specificity, where physical and chemical properties are duly tailored and combined with appropriate mechanical and degradation features in order to trigger specific cell events and functions involved in the regenerative process. In this chapter, the chemical, physical, and biological elements that have to be targeted by biocomposites in regenerative medicine are described.

JTD Keywords: Biocomposite, Regenerative medicine, Tissue engineering, Scaffolds, Cell/material interactions


Obregón, R., Ramón-Azcón, J., Ahadian, S., (2017). Nanofiber composites in blood vessel tissue engineering Nanofiber Composites for Biomedical Applications (ed. Ramalingam, M., Ramakrishna, S.), Elsevier (Duxford, UK) Woodhead Publishing Series in Biomaterials, 483-506

Tissue engineering (TE) aims to restore function or replace damaged tissue through biological principles and engineering. Nanofibers are attractive substrates for tissue regeneration applications because they structurally mimic the native extracellular matrix. Composite nanofibers, which are hybrid nanofibers blended from natural and synthetic polymers, represent a major advancement in TE and regenerative medicine, since they take advantage of the physical properties of the synthetic polymer and the bioactivity of the natural polymer while minimizing the disadvantages of both. Although various nanofibrous matrices have been applied to almost all the areas of TE, in this chapter we will focus on nanofiber composites scaffolds for vascular TE.

JTD Keywords: Blood vessels, Nanofiber composite, Tissue engineering, Vascularized tissue


Vila, M., García, A., Girotti, A., Alonso, M., Rodríguez-Cabello, J. C., González-Vázquez, A., Planell, J. A., Engel, E., Buján, J., Garcíaa-Honduvilla, N., Vallet-Regí, M., (2016). 3D silicon doped hydroxyapatite scaffolds decorated with Elastin-like Recombinamers for bone regenerative medicine Acta Biomaterialia 45, 349-356

The current study reports on the manufacturing by rapid prototyping technique of three-dimensional (3D) scaffolds based on silicon substituted hydroxyapatite with Elastin-like Recombinamers (ELRs) functionalized surfaces. Silicon doped hydroxyapatite (Si-HA), with Ca10(PO4)5.7(SiO4)0.3(OH)1.7h0.3 nominal formula, was surface functionalized with two different types of polymers designed by genetic engineering: ELR-RGD that contain cell attachment specific sequences and ELR-SNA15/RGD with both hydroxyapatite and cells domains that interact with the inorganic phase and with the cells, respectively. These hybrid materials were subjected to in vitro assays in order to clarify if the ELRs coating improved the well-known biocompatible and bone regeneration properties of calcium phosphates materials. The in vitro tests showed that there was a total and homogeneous colonization of the 3D scaffolds by Bone marrow Mesenchymal Stromal Cells (BMSCs). In addition, the BMSCs were viable and able to proliferate and differentiate into osteoblasts. Statement of Significance Bone tissue engineering is an area of increasing interest because its main applications are directly related to the rising life expectancy of the population, which promotes higher rates of several bone pathologies, so innovative strategies are needed for bone tissue regeneration therapies. Here we use the rapid prototyping technology to allow moulding ceramic 3D scaffolds and we use different bio-polymers for the functionalization of their surfaces in order to enhance the biological response. Combining the ceramic material (silicon doped hydroxyapatite, Si-HA) and the Elastin like Recombinamers (ELRs) polymers with the presence of the integrin-mediate adhesion domain alone or in combination with SNA15 peptide that possess high affinity for hydroxyapatite, provided an improved Bone marrow Mesenchymal Stromal Cells (BMSCs) differentiation into osteoblastic linkage.

JTD Keywords: Bone marrow Mesenchymal Stromal Cells (BMSCs), Bone repair, Elastin-like Recombinamers (ELRs), Rapid prototyped 3D scaffolds, Silicon doped hydroxyapatite (Si-HA), Tissue engineering


Wills, C. R., Malandrino, A., Van Rijsbergen, M., Lacroix, D., Ito, K., Noailly, J., (2016). Simulating the sensitivity of cell nutritive environment to composition changes within the intervertebral disc Journal of the Mechanics and Physics of Solids 90, 108-123

Altered nutrition in the intervertebral disc affects cell viability and can generate catabolic cascades contributing to extracellular matrix (ECM) degradation. Such degradation is expected to affect couplings between disc mechanics and nutrition, contributing to accelerate degenerative processes. However, the relation of ECM changes to major biophysical events within the loaded disc remains unclear. A L4-L5 disc finite element model including the nucleus (NP), annulus (AF) and endplates was used and coupled to a transport-cell viability model. Solute concentrations and cell viability were evaluated along the mid-sagittal plane path. A design of experiment (DOE) was performed. DOE parameters corresponded to AF and NP biochemical tissue measurements in discs with different degeneration grades. Cell viability was not affected by any parameter combinations defined. Nonetheless, the initial water content was the parameter that affected the most the solute contents, especially glucose. Calculations showed that altered NP composition could negatively affect AF cell nutrition. Results suggested that AF and NP tissue degeneration are not critical to nutrition-related cell viability at early-stage of disc degeneration. However, small ECM degenerative changes may alter significantly disc nutrition under mechanical loads. Coupling disc mechano-transport simulations and enzyme expression studies could allow identifying spatiotemporal sequences related to tissue catabolism.

JTD Keywords: Cell nutrition, Finite element analysis, Intervertebral disc degeneration, Multiphysics, Tissue composition


Valero, C., Navarro, B., Navajas, D., García-Aznar, J. M., (2016). Finite element simulation for the mechanical characterization of soft biological materials by atomic force microscopy Journal of the Mechanical Behavior of Biomedical Materials , 62, 222-235

The characterization of the mechanical properties of soft materials has been traditionally performed through uniaxial tensile tests. Nevertheless, this method cannot be applied to certain extremely soft materials, such as biological tissues or cells that cannot be properly subjected to these tests. Alternative non-destructive tests have been designed in recent years to determine the mechanical properties of soft biological tissues. One of these techniques is based on the use of atomic force microscopy (AFM) to perform nanoindentation tests. In this work, we investigated the mechanical response of soft biological materials to nanoindentation with spherical indenters using finite element simulations. We studied the responses of three different material constitutive laws (elastic, isotropic hyperelastic and anisotropic hyperelastic) under the same process and analyzed the differences thereof. Whereas linear elastic and isotropic hyperelastic materials can be studied using an axisymmetric simplification, anisotropic hyperelastic materials require three-dimensional analyses. Moreover, we established the limiting sample size required to determine the mechanical properties of soft materials while avoiding boundary effects. Finally, we compared the results obtained by simulation with an estimate obtained from Hertz theory. Hertz theory does not distinguish between the different material constitutive laws, and thus, we proposed corrections to improve the quantitative measurement of specific material properties by nanoindentation experiments.

JTD Keywords: AFM, Cell mechanics, FEM, Nanoindentation, Soft-tissue


González, F., (2016). CRISPR/Cas9 genome editing in human pluripotent stem cells: Harnessing human genetics in a dish Developmental Dynamics , 245, (7), 788-806

Abstract: Because of their extraordinary differentiation potential, human pluripotent stem cells (hPSCs) can differentiate into virtually any cell type of the human body, providing a powerful platform not only for generating relevant cell types useful for cell replacement therapies, but also for modeling human development and disease. Expanding this potential, structures resembling human organs, termed organoids, have been recently obtained from hPSCs through tissue engineering. Organoids exhibit multiple cell types self-organizing into structures recapitulating in part the physiology and the cellular interactions observed in the organ in vivo, offering unprecedented opportunities for human disease modeling. To fulfill this promise, tissue engineering in hPSCs needs to be supported by robust and scalable genome editing technologies. With the advent of the CRISPR/Cas9 technology, manipulating the genome of hPSCs has now become an easy task, allowing modifying their genome with superior precision, speed, and throughput. Here we review current and potential applications of the CRISPR/Cas9 technology in hPSCs and how they contribute to establish hPSCs as a model of choice for studying human genetics.

JTD Keywords: CRISPR/Cas9, Disease modeling, Human genetics, Human pluripotent stem cells, Tissue and genome engineering


Sánchez-Ferrero, Aitor, Mata, Álvaro, Mateos-Timoneda, Miguel A., Rodríguez-Cabello, José C., Alonso, Matilde, Planell, Josep, Engel, Elisabeth, (2015). Development of tailored and self-mineralizing citric acid-crosslinked hydrogels for in situ bone regeneration Biomaterials 68, 42-53

Bone tissue engineering demands alternatives overcoming the limitations of traditional approaches in the context of a constantly aging global population. In the present study, elastin-like recombinamers hydrogels were produced by means of carbodiimide-catalyzed crosslinking with citric acid, a molecule suggested to be essential for bone nanostructure. By systematically studying the effect of the relative abundance of reactive species on gelation and hydrogel properties such as functional groups content, degradation and structure, we were able to understand and to control the crosslinking reaction to achieve hydrogels mimicking the fibrillary nature of the extracellular matrix. By studying the effect of polymer concentration on scaffold mechanical properties, we were able to produce hydrogels with a stiffness value of 36.13 ± 10.72 kPa, previously suggested to be osteoinductive. Microstructured and mechanically-tailored hydrogels supported the growth of human mesenchymal stem cells and led to higher osteopontin expression in comparison to their non-tailored counterparts. Additionally, tailored hydrogels were able to rapidly self-mineralize in biomimetic conditions, evidencing that citric acid was successfully used both as a crosslinker and a bioactive molecule providing polymers with calcium phosphate nucleation capacity.

JTD Keywords: Biomimetic material, Biomineralisation, Bone tissue engineering, Cross-linking, Hydrogel, Mesenchymal stem cell


Perea-Gil, I., Uriarte, J. J., Prat-Vidal, C., Gálvez-Montón, C., Roura, S., Llucià-Valldeperas, A., Soler-Botija, C., Farré, R., Navajas, D., Bayes-Genis, A., (2015). In vitro comparative study of two decellularization protocols in search of an optimal myocardial scaffold for recellularization American Journal of Translational Research , 7, (3), 558-573

Introduction. Selection of a biomaterial-based scaffold that mimics native myocardial extracellular matrix (ECM) architecture can facilitate functional cell attachment and differentiation. Although decellularized myocardial ECM accomplishes these premises, decellularization processes may variably distort or degrade ECM structure. Materials and methods. Two decellularization protocols (DP) were tested on porcine heart samples (epicardium, mid myocardium and endocardium). One protocol, DP1, was detergent-based (SDS and Triton X-100), followed by DNase I treatment. The other protocol, DP2, was focused in trypsin and acid with Triton X-100 treatments. Decellularized myocardial scaffolds were reseeded by embedding them in RAD16-I peptidic hydrogel with adipose tissue-derived progenitor cells (ATDPCs). Results. Both protocols yielded acellular myocardial scaffolds (~82% and ~94% DNA reduction for DP1 and DP2, respectively). Ultramicroscopic assessment of scaffolds was similar for both protocols and showed filamentous ECM with preserved fiber disposition and structure. DP1 resulted in more biodegradable scaffolds (P = 0.04). Atomic force microscopy revealed no substantial ECM stiffness changes post-decellularization compared to native tissue. The Young’s modulus did not differ between heart layers (P = 0.69) or decellularization protocols (P = 0.15). After one week, recellularized DP1 scaffolds contained higher cell density (236 ± 106 and 98 ± 56 cells/mm2 for recellularized DP1 and DP2 scaffolds, respectively; P = 0.04). ATDPCs in both DP1 and DP2 scaffolds expressed the endothelial marker isolectin B4, but only in the DP1 scaffold ATDPCs expressed the cardiac markers GATA4, connexin43 and cardiac troponin T. Conclusions. In our hands, DP1 produced myocardial scaffolds with higher cell repopulation and promotes ATDPCs expression of endothelial and cardiomyogenic markers.

JTD Keywords: Acellular myocardial scaffold, Adipose tissue-derived progenitor cells, Decellularization protocols, Extracellular matrix, Myocardial infarction, Recellularization


da Palma, R. K., Campillo, N., Uriarte, J. J., Oliveira, L. V. F., Navajas, D., Farré, R., (2015). Pressure- and flow-controlled media perfusion differently modify vascular mechanics in lung decellularization Journal of the Mechanical Behavior of Biomedical Materials , 49, 69-79

Organ biofabrication is a potential future alternative for obtaining viable organs for transplantation. Achieving intact scaffolds to be recellularized is a key step in lung bioengineering. Perfusion of decellularizing media through the pulmonary artery has shown to be effective. How vascular perfusion pressure and flow vary throughout lung decellularization, which is not well known, is important for optimizing the process (minimizing time) while ensuring scaffold integrity (no barotrauma). This work was aimed at characterizing the pressure/flow relationship at the pulmonary vasculature and at how effective vascular resistance depends on pressure- and flow-controlled variables when applying different methods of media perfusion for lung decellularization. Lungs from 43 healthy mice (C57BL/6; 7-8 weeks old) were investigated. After excision and tracheal cannulation, lungs were inflated at 10cmH2O airway pressure and subjected to conventional decellularization with a solution of 1% sodium dodecyl sulfate (SDS). Pressure (PPA) and flow (V'PA) at the pulmonary artery were continuously measured. Decellularization media was perfused through the pulmonary artery: (a) at constant PPA=20cmH2O or (b) at constant V'PA=0.5 and 0.2ml/min. Effective vascular resistance was computed as Rv=PPA/V'PA. Rv (in cmH2O/(ml/min)); mean±SE) considerably varied throughout lung decellularization, particularly for pressure-controlled perfusion (from 29.1±3.0 in baseline to a maximum of 664.1±164.3 (p<0.05), as compared with flow-controlled perfusion (from 49.9±3.3 and 79.5±5.1 in baseline to a maximum of 114.4±13.9 and 211.7±70.5 (p<0.05, both), for V'PA of 0.5 and 0.2ml/min respectively. Most of the media infused to the pulmonary artery throughout decellularization circulated to the airways compartment across the alveolar-capillary membrane. This study shows that monitoring perfusion mechanics throughout decellularization provides information relevant for optimizing the process time while ensuring that vascular pressure is kept within a safety range to preserve the organ scaffold integrity.

JTD Keywords: Acellular lung, Fluid mechanics, Lung bioengineering, Lung scaffold, Organ biofabrication, Tissue engineering, Vascular resistance


Won, J. E., Mateos-Timoneda, M. A., Castaño, O., Planell, J. A., Seo, S. J., Lee, E. J., Han, C. M., Kim, H. W., (2015). Fibronectin immobilization on to robotic-dispensed nanobioactive glass/polycaprolactone scaffolds for bone tissue engineering Biotechnology Letters , 37, (4), 935-342

Bioactive nanocomposite scaffolds with cell-adhesive surface have excellent bone regeneration capacities. Fibronectin (FN)-immobilized nanobioactive glass (nBG)/polycaprolactone (PCL) (FN-nBG/PCL) scaffolds with an open pore architecture were generated by a robotic-dispensing technique. The surface immobilization level of FN was significantly higher on the nBG/PCL scaffolds than on the PCL scaffolds, mainly due to the incorporated nBG that provided hydrophilic chemical-linking sites. FN-nBG/PCL scaffolds significantly improved cell responses, including initial anchorage and subsequent cell proliferation. Although further in-depth studies on cell differentiation and the in vivo animal responses are required, bioactive nanocomposite scaffolds with cell-favoring surface are considered to provide promising three-dimensional substrate for bone regeneration.

JTD Keywords: Bone scaffolds, Cell response, Fibronectin, Nanobioactive glass, Nanocomposites, Polycaprolactone, Bone, Cell proliferation, Cells, Cytology, Glass, Nanocomposites, Polycaprolactone, Robotics, Bone scaffolds, Bone tissue engineering, Cell response, Fibronectin, Fibronectin immobilizations, Nano bioactive glass, Nanocomposite scaffolds, Three-dimensional substrates, Scaffolds (biology)


Pérez-Madrigal, M. M., Giannotti, M. I., Del Valle, L. J., Franco, L., Armelin, E., Puiggalí, J., Sanz, F., Alemán, C., (2014). Thermoplastic polyurethane:polythiophene nanomembranes for biomedical and biotechnological applications ACS Applied Materials & Interfaces 6, (12), 9719-9732

Nanomembranes have been prepared by spin-coating mixtures of a polythiophene (P3TMA) derivative and thermoplastic polyurethane (TPU) using 20:80, 40:60, and 60:40 TPU:P3TMA weight ratios. After structural, topographical, electrochemical, and thermal characterization, properties typically related with biomedical applications have been investigated: swelling, resistance to both hydrolytic and enzymatic degradation, biocompatibility, and adsorption of type I collagen, which is an extra cellular matrix protein that binds fibronectin favoring cell adhesion processes. The swelling ability and the hydrolytic and enzymatic degradability of TPU:P3TMA membranes increases with the concentration of P3TMA. Moreover, the degradation of the blends is considerably promoted by the presence of enzymes in the hydrolytic medium, TPU:P3TMA blends behaving as biodegradable materials. On the other hand, TPU:P3TMA nanomembranes behave as bioactive platforms stimulating cell adhesion and, especially, cell viability. Type I collagen adsorption largely depends on the substrate employed to support the nanomembrane, whereas it is practically independent of the chemical nature of the polymeric material used to fabricate the nanomembrane. However, detailed microscopy study of the morphology and topography of adsorbed collagen evidence the formation of different organizations, which range from fibrils to pseudoregular honeycomb networks depending on the composition of the nanomembrane that is in contact with the protein. Scaffolds made of electroactive TPU:P3TMA nanomembranes are potential candidates for tissue engineering biomedical applications.

JTD Keywords: Bioactive platform, Biodegradable blend, Collaged adsorption, Scaffolds, Tissue engineering, Ultrathin films


Tahirbegi, I. B., Mir, M., Schostek, S., Schurr, M., Samitier, J., (2014). In vivo ischemia monitoring array for endoscopic surgery Biosensors and Bioelectronics 61, 124-130

An array with all-solid-state, potentiometric, miniaturized sensors for pH and potassium was developed to be introduced into the stomach or other sectors of the digestive tract by means of flexible endoscopy. These sensors perform continuous and simultaneous measurement of extracellular pH and potassium. This detection seeks to sense ischemia in the gastric mucosa inside the stomach, an event indicative of local microvascular perfusion and tissue oxygenation status. Our array is proposed as a medical tool to identify the occurrence of the ischemia after gastrointestinal or gastroesophageal anastomosis. The stability and feasibility of the miniaturized working and reference electrodes integrated in the array were studied under in vitro conditions, and the behavior of the potassium and pH ion-selective membranes were optimized to work under acidic gastric conditions with high concentrations of HCl. The array was tested in vivo in pigs to measure the ischemia produced by clamping the blood flow into the stomach. Our results indicate that ischemic and reperfusion states can be sensed in vivo and that information on tissue damage can be collected by this sensor array. The device described here provides a miniaturized, inexpensive, and mass producible sensor array for detecting local ischemia caused by unfavorable anastomotic perfusion and will thus contribute to preventing anastomotic leakage and failure caused by tissue necrosis.

JTD Keywords: Endoscopy, Surgery, Tissue, Gastric anastomosis, Gastric conditions, Ion selective sensors, Ischemia, pH detection, Reference electrodes, Simultaneous measurement, Tissue oxygenation, Sensors


Dalmases, M., Torres, M., Márquez-Kisinousky, L., Almendros, I., Planas, A. M., Embid, C., Martínez-Garcia, M. A., Navajas, D., Farré, R., Montserrat, J. M., (2014). Brain tissue hypoxia and oxidative stress induced by obstructive apneas is different in young and aged rats Sleep , 37, (7), 1249-1256

Study Objectives: To test the hypotheses that brain oxygen partial pressure (PtO2) in response to obstructive apneas changes with age and that it might lead to different levels of cerebral tissue oxidative stress. Design: Prospective controlled animal study. Setting: University laboratory. Participants: Sixty-four male Wistar rats: 32 young (3 mo old) and 32 aged (18 mo). Interventions: Protocol 1: Twenty-four animals were subjected to obstructive apneas (50 apneas/h, lasting 15 sec each) or to sham procedure for 50 min. Protocol 2: Forty rats were subjected to obstructive apneas or sham procedure for 4 h. Measurements and Results: Protocol 1: Real-time PtO2 measurements were performed using a fast-response oxygen microelectrode. During successive apneas cerebral cortex PtO2 presented a different pattern in the two age groups; there was a fast increase in young rats, whereas it remained without significant changes between the beginning and the end of the protocol in the aged group. Protocol 2: Brain oxidative stress assessed by lipid peroxidation increased after apneas in young rats (1.34 ± 0.17 nmol/mg of protein) compared to old ones (0.63 ± 0.03 nmol/mg), where a higher expression of antioxidant enzymes was observed. Conclusions: The results suggest that brain oxidative stress in aged rats is lower than in young rats in response to recurrent apneas, mimicking obstructive sleep apnea. This could be due to the different PtO2 response observed between age groups and the increased antioxidant expression in aged rats.

JTD Keywords: Aging, Animal model, Obstructive apnea, Oxidative stress, Tissue oxygenation, antioxidant, glutathione disulfide, aged, animal experiment, animal model, animal tissue, apnea, arterial oxygen saturation, article, brain cortex, brain oxygen tension, brain tissue, controlled study, groups by age, hypoxia, lipid peroxidation, male, nonhuman, oxidative stress, pressure, priority journal, rat


Melo, E., Cárdenes, N., Garreta, E., Luque, T., Rojas, M., Navajas, D., Farré, R., (2014). Inhomogeneity of local stiffness in the extracellular matrix scaffold of fibrotic mouse lungs Journal of the Mechanical Behavior of Biomedical Materials , 37, 186-195

Lung disease models are useful to study how cell engraftment, proliferation and differentiation are modulated in lung bioengineering. The aim of this work was to characterize the local stiffness of decellularized lungs in aged and fibrotic mice. Mice (2- and 24-month old; 14 of each) with lung fibrosis (N=20) and healthy controls (N=8) were euthanized after 11 days of intratracheal bleomycin (fibrosis) or saline (controls) infusion. The lungs were excised, decellularized by a conventional detergent-based (sodium-dodecyl sulfate) procedure and slices of the acellular lungs were prepared to measure the local stiffness by means of atomic force microscopy. The local stiffness of the different sites in acellular fibrotic lungs was very inhomogeneous within the lung and increased according to the degree of the structural fibrotic lesion. Local stiffness of the acellular lungs did not show statistically significant differences caused by age. The group of mice most affected by fibrosis exhibited local stiffness that were ~2-fold higher than in the control mice: from 27.2±1.64 to 64.8±7.1. kPa in the alveolar septa, from 56.6±4.6 to 99.9±11.7. kPa in the visceral pleura, from 41.1±8.0 to 105.2±13.6. kPa in the tunica adventitia, and from 79.3±7.2 to 146.6±28.8. kPa in the tunica intima. Since acellular lungs from mice with bleomycin-induced fibrosis present considerable micromechanical inhomogeneity, this model can be a useful tool to better investigate how different degrees of extracellular matrix lesion modulate cell fate in the process of organ bioengineering from decellularized lungs.

JTD Keywords: Ageing, Atomic force microscopy, Decellularization, Lung fibrosis, Tissue engineering, Atomic force microscopy, Biological organs, Peptides, Sodium dodecyl sulfate, Sodium sulfate, Tissue engineering, Ageing, Decellularization, Extracellular matrices, Healthy controls, Inhomogeneities, Lung fibrosis, Micro-mechanical, Statistically significant difference, Mammals, bleomycin, adventitia, animal experiment, animal model, article, atomic force microscopy, bleomycin-induced pulmonary fibrosis, cell fate, controlled study, extracellular matrix, female, intima, lung alveolus, lung fibrosis, lung mechanics, mechanical probe, microenvironment, mouse, nonhuman, pleura, priority journal, rigidity, tissue engineering


Uriarte, J. J., Nonaka, P. N., Campillo, N., Palma, R. K., Melo, E., de Oliveira, L. V. F., Navajas, D., Farré, R., (2014). Mechanical properties of acellular mouse lungs after sterilization by gamma irradiation Journal of the Mechanical Behavior of Biomedical Materials , 40, 168-177

Lung bioengineering using decellularized organ scaffolds is a potential alternative for lung transplantation. Clinical application will require donor scaffold sterilization. As gamma-irradiation is a conventional method for sterilizing tissue preparations for clinical application, the aim of this study was to evaluate the effects of lung scaffold sterilization by gamma irradiation on the mechanical properties of the acellular lung when subjected to the artificial ventilation maneuvers typical within bioreactors. Twenty-six mouse lungs were decellularized by a sodium dodecyl sulfate detergent protocol. Eight lungs were used as controls and 18 of them were submitted to a 31kGy gamma irradiation sterilization process (9 kept frozen in dry ice and 9 at room temperature). Mechanical properties of acellular lungs were measured before and after irradiation. Lung resistance (RL) and elastance (EL) were computed by linear regression fitting of recorded signals during mechanical ventilation (tracheal pressure, flow and volume). Static (Est) and dynamic (Edyn) elastances were obtained by the end-inspiratory occlusion method. After irradiation lungs presented higher values of resistance and elastance than before irradiation: RL increased by 41.1% (room temperature irradiation) and 32.8% (frozen irradiation) and EL increased by 41.8% (room temperature irradiation) and 31.8% (frozen irradiation). Similar increases were induced by irradiation in Est and Edyn. Scanning electron microscopy showed slight structural changes after irradiation, particularly those kept frozen. Sterilization by gamma irradiation at a conventional dose to ensure sterilization modifies acellular lung mechanics, with potential implications for lung bioengineering.

JTD Keywords: Gamma irradiation, Lung bioengineering, Lung decellularization, Organ scaffold, Pulmonary mechanics, Decellularization, Gamma irradiation, Mouse lung, Pulmonary mechanics, dodecyl sulfate sodium, animal tissue, Article, artificial ventilation, bioengineering, bioreactor, compliance (physical), controlled study, freezing, gamma irradiation, lung, lung mechanics, lung resistance, male, mouse, nonhuman, room temperature, scanning electron microscopy, tissue scaffold, trachea pressure


Sanzana, E. S., Navarro, M., Ginebra, M. P., Planell, J. A., Ojeda, A. C., Montecinos, H. A., (2014). Role of porosity and pore architecture in the in vivo bone regeneration capacity of biodegradable glass scaffolds Journal of Biomedical Materials Research - Part A , 102, (6), 1767-1773

The aim of this work is to shed light on the role of porosity and pore architecture in the in vivo bone regeneration capacity of biodegradable glass scaffolds. A calcium phosphate glass in the system P2O5-CaO-Na2O-TiO2 was foamed using two different porogens, namely albumen and hydrogen peroxide (H2O2); the resulting three-dimensional porous structures were characterized and implanted in New Zealand rabbits to study their in vivo behavior. Scaffolds foamed with albumen displayed a monomodal pore size distribution centered around 150 μm and a porosity of 82%, whereas scaffolds foamed with H2O2 showed lower porosity (37%), with larger elongated pores, and multimodal size distribution. After 12 weeks of implantation, histology results revealed a good osteointegration for both types of scaffolds. The quantitative morphometric analysis showed the substitution of the biomaterial by new bone in the case of glasses foamed with albumen. In contrast, bone neoformation and material resorption were significantly lower in the defects filled with the scaffolds foamed with H2O2. The results obtained in this study showed that both calcium phosphate glass scaffolds were osteoconductive, biocompatible, and biodegradable materials. However, differences in porosity, pore architecture, and microstructure led to substantially different in vivo response.

JTD Keywords: Bone substitutes, Calcium phosphate glasses, in vivo, Scaffolds, Tissue engineering


Rajzer, I., Menaszek, E., Kwiatkowski, R., Planell, J. A., Castaño, O., (2014). Electrospun gelatin/poly(ε-caprolactone) fibrous scaffold modified with calcium phosphate for bone tissue engineering Materials Science and Engineering: C 44, 183-190

In this study gelatin (Gel) modified with calcium phosphate nanoparticles (SG5) and polycaprolactone (PCL) were used to prepare a 3D bi-layer scaffold by collecting electrospun PCL and gelatin/SG5 fibers separately in the same collector. The objective of this study was to combine the desired properties of PCL and Gel/SG5 in the same scaffold in order to enhance mineralization, thus improving the ability of the scaffold to bond to the bone tissue. The scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and the wide angle X-ray diffraction (WAXD) measurements confirmed that SG5 nanoparticles were successfully incorporated into the fibrous gelatin matrix. The composite Gel/SG5/PCL scaffold exhibited more enhanced mechanical properties than individual Gel and Gel/SG5 scaffolds. The presence of SG5 nanoparticles accelerated the nucleation and growth of apatite crystals on the surface of the composite Gel/SG5/PCL scaffold in simulated body fluid (SBF). The osteoblast response in vitro to developed electrospun scaffolds (PCL and Gel/SG5/PCL) was investigated by using normal human primary NHOst cell lines. NHOst cell culture studies showed that higher alkaline phosphatase (ALP) activity and better mineralization were obtained in the case of composite materials than in pure PCL scaffolds. The mechanically strong PCL scaffold served as a skeleton, while the Gel/SG5 fibers facilitated cell spreading and mineralization of the scaffold.

JTD Keywords: Bilayer fibrous scaffold, Ceramic nanoparticles, Electrospinning, Gelatin, Polycaprolactone, Biomechanics, Bone, Calcium phosphate, Cell culture, Electrospinning, Fourier transform infrared spectroscopy, Mechanical properties, Mineralogy, Nanoparticles, Phosphatases, Polycaprolactone, Scanning electron microscopy, X ray diffraction, Polycaprolactone, Alkaline phosphatase activity, Bone tissue engineering, Calcium phosphate nanoparticles, Ceramic nanoparticles, Fibrous scaffolds, Gelatin, Simulated body fluids, Wide-angle x-ray diffraction, Electrospuns, Scaffolds (biology), Electrospinning


Malandrino, A., Noailly, J., Lacroix, D., (2014). Numerical exploration of the combined effect of nutrient supply, tissue condition and deformation in the intervertebral disc Journal of Biomechanics 47, (6), 1520-1525

Novel strategies to heal discogenic low back pain could highly benefit from comprehensive biophysical studies that consider both mechanical and biological factors involved in intervertebral disc degeneration. A decrease in nutrient availability at the bone-disc interface has been indicated as a relevant risk factor and as a possible initiator of cell death processes. Mechanical behaviour of both healthy and degenerated discs could highly interact with cell death in these compromised situations. In the present study, a mechano-transport finite element model was used to investigate the nature of mechanical effects on cell death processes via load-induced metabolic transport variations. Cycles of static sustained compression were chosen to simulate daily human activity. Healthy and degenerated cases were simulated as well as a reduced supply of solutes and an increase in solute exchange area at the bone-disc interface. Results showed that a reduction in metabolite concentrations at the bone-disc boundaries induced cell death, even when the increased exchange area was simulated. Slight local mechanical enhancements of glucose in the disc centre were capable of decelerating cell death but occurred only with healthy mechanical properties. However, mechanical deformations were responsible for a worsening in terms of cell death in the inner annulus, a disadvantaged zone far from the boundary supply with both an increased cell demand and a strain-dependent decrease of diffusivity. Such adverse mechanical effects were more accentuated when degenerative properties were simulated. Overall, this study paves the way for the use of biophysical models for a more integrated understanding of intervertebral disc pathophysiology.

JTD Keywords: Boundary conditions, Cell nutrition, Cell viability, Computational analysis, Intervertebraldisc, Softtissuebiomechanics


Nonaka, P. N., Uriarte, J. J., Campillo, N., Melo, E., Navajas, D., Farré, R., Oliveira, L. V. F., (2014). Mechanical properties of mouse lungs along organ decellularization by sodium dodecyl sulfate Respiratory Physiology & Neurobiology , 200, 1-5

Lung decellularization is based on the use of physical, chemical, or enzymatic methods to break down the integrity of the cells followed by a treatment to extract the cellular material from the lung scaffold. The aim of this study was to characterize the mechanical changes throughout the different steps of lung decellularization process. Four lungs from mice (C57BL/6) were decellularized by using a conventional protocol based on sodium dodecyl sulfate. Lungs resistance (RL) and elastance (EL) were measured along decellularization steps and were computed by linear regression fitting of tracheal pressure, flow, and volume during mechanical ventilation. Transients differences found were more distinct in an intermediate step after the lungs were rinsed with deionized water and treated with 1% SDS, whereupon the percentage of variation reached approximately 80% for resistance values and 30% for elastance values. In conclusion, although a variation in extracellular matrix stiffness was observed during the decellularization process, this variation can be considered negligible overall because the resistance and elastance returned to basal values at the final decellularization step.

JTD Keywords: Lung bioengineering, Lung decellularization, Organ scaffold, dodecyl sulfate sodium, animal tissue, article, artificial ventilation, compliance (physical), controlled study, enzyme chemistry, extracellular matrix, female, flow, lung, lung decellularization, lung pressure, lung resistance, mouse, nonhuman, positive end expiratory pressure, priority journal, rigidity, tissue engineering, trachea pressure


Pérez-Amodio, Soledad, Engel, Elisabeth, (2014). Bone biology and Regeneration Bio-Ceramics with Clinical Applications (ed. Vallet-Regí, M.), John Wiley & Sons, Ltd (Chichester, UK) , 315-342

Each bone of the skeleton constantly undergoes modeling during life to help it to adapt to changing biomechanical forces as well as remodeling to remove old bone and replace it with new, mechanically stronger bone to help preserve bone strength. Bone remodeling involves the removal of mineralized bone by osteoclasts, followed by the formation of bone matrix through the osteoblasts that subsequently become mineralized. All these assets make bone a suitable model for regeneration. Bone tissue can be grossly divided into inorganic mineral material (mostly HA), and organic material from cells and the extracellular matrix. This chapter outlines some of the bone diseases such as osteoporosis and Paget's disease. Bone can be considered as a biphasic composite material, with two phases: one the mineral and the other collagen. This combination confers better mechanical properties on the tissue than each component itself.

JTD Keywords: Bone biology, Bone cells, Bone diseases, Bone extracellular matrix, Bone mechanics, Bone remodeling, Bone tissue regeneration, Skeleton


Noailly, J., Malandrino, A., Galbusera, F., Jin, Zhongmin, (2014). Computational modelling of spinal implants Computational Modelling of Biomechanics and Biotribology in the Musculoskeletal System (ed. Jin, Z.), Woodhead Publishing (Cambridge, UK) Biomaterials and Tissues, 447-484

This chapter focuses on the use of the finite element method in the design and exploration of spinal implants. Following an introduction to biomechanical alterations of the spine in disease and to spine finite element modelling, focus is placed on different models developed for spine treatment simulations. Despite the hindrance of working thorough representations of in vivo situations, predictions of load transfer within both the implants and the tissues simulated allow improved interpretations of known clinical outcomes, and permit the educated design of new implants. The potential of probabilistic modelling is also discussed in relation to model validation and patient-specific analyses. Finally, the latest developments in the multiphysical modelling of intervertebral discs are presented, revealing a strong potential for the study of implant-based strategies that aim to restore the functional biophysics of the spine.

JTD Keywords: Spinal implant, Finite element modelling, Spine surgery, Spine biomechanics, Tissue mechanobiology


Gustavsson, J., Planell, J., Engel, E., (2013). Ion-selective electrodes to monitor osteoblast-like cellular influence on the extracellular concentration of calcium Journal of Tissue Engineering and Regenerative Medicine 7, (8), 609-620

In bone tissue engineering, the composition of the ionic extracellular environment (IEE) can determine both cellular fate and a biomaterial's development and performance. Therefore, precise control of the IEE and a perfect understanding of the dynamic changes that it can be subject to due to cellular activity is highly desired. To achieve this, we initially monitored how two standard osteoblast-like cell models that expressed either high or low alkaline phosphatase activity - SAOS-2 and MG63 cells, respectively - affected the extracellular concentrations of calcium and phosphate during long-term cultures. It was observed that cellular influence on the IEE varied greatly between the two models and could be linked to the capacity of cells to deposit calcium in the extracellular matrix. Miniaturized ion-selective electrodes that could allow for real-time monitoring of calcium in a minimally invasive way were then constructed. The electrodes were characterized in standard in vitro cell culture environments, prior to being successfully applied for periods of 24h, to record the dynamics of cell-induced deposition of calcium in the extracellular matrix, while using osteogenic media of either high or low concentrations of phosphate. As a result, this study provides the background and technological means for the non-destructive evaluation of the IEE in vitro and allows for the optimization and development of better models of bone tissue construction.

JTD Keywords: Extracellular ions, Ion-selective electrode, MG63, Mineralization, Osteoblasts, Saos-2, Sensor, Tissue engineering


Gil, V., Del Río, J. A., (2012). Analysis of axonal growth and cell migration in 3D hydrogel cultures of embryonic mouse CNS tissue Nature Protocols 7, (2), 268-280

This protocol uses rat tail-derived type I collagen hydrogels to analyze key processes in developmental neurobiology, such as chemorepulsion and chemoattraction. The method is based on culturing small pieces of brain tissue from embryonic or early perinatal mice inside a 3D hydrogel formed by rat tail-derived type I collagen or, alternatively, by commercial Matrigel. The neural tissue is placed in the hydrogel with other brain tissue pieces or cell aggregates genetically modified to secrete a particular molecule that can generate a gradient inside the hydrogel. The present method is uncomplicated and generally reproducible, and only a few specific details need to be considered during its preparation. Moreover, the degree and behavior of axonal growth or neural migration can be observed directly using phase-contrast, fluorescence microscopy or immunocytochemical methods. This protocol can be carried out in 4 weeks.

JTD Keywords: Cell biology, Cell culture, Developmental biology, Imaging, Model organisms, Neuroscience, Tissue culture


Tejeda-Montes, E., Smith, K. H., Poch, M., López-Bosque, M. J., Martín, L., Alonso, M., Engel, E., Mata, Alvaro., (2012). Engineering membrane scaffolds with both physical and biomolecular signaling Acta Biomaterialia 8, (3), 998-1009

We report on the combination of a top-down and bottom-up approach to develop thin bioactive membrane scaffolds based on functional elastin-like polymers (ELPs). Our strategy combines ELP cross-linking and assembly, and a variety of standard and novel micro/nanofabrication techniques to create self-supporting membranes down to ∼500 nm thick that incorporate both physical and biomolecular signals, which can be easily tailored for a specific application. In this study we used an ELP that included the cell-binding motif arginine-glycine-aspartic acid-serine (RGDS). Furthermore, fabrication processes were developed to create membranes that exhibited topographical patterns with features down to 200 nm in lateral dimensions and up to 10 μm in height on either one or both sides, uniform and well-defined pores, or multiple ELP layers. A variety of processing parameters were tested in order to optimize membrane fabrication, including ELP and cross-linker concentration, temperature, reaction time and ambient humidity. Membrane micro/nanopatterning, swelling and stiffness were characterized by atomic force microscopy, nanoindentation tests and scanning electron microscopy. Upon immersion in phosphate-buffered saline and an increase in temperature from 25 to 40°C, membranes exhibited a significant increase in surface stiffness, with the reduced Young's modulus increasing with temperature. Finally, rat mesenchymal stem cells were cultured on thin RGDS-containing membranes, which allowed cell adhesion, qualitatively enhanced spreading compared to membranes without RGDS epitopes and permitted proliferation. Furthermore, cell morphology was drastically affected by topographical patterns on the surface of the membranes.

JTD Keywords: Elastin-like polymers, Membranes, Nanotechnology, Scaffolds, Tissue engineering


Malandrino, A., Fritsch, A., Lahayne, O., Kropik, K., Redl, H., Noailly, J., Lacroix, D., Hellmich, C., (2012). Anisotropic tissue elasticity in human lumbar vertebra, by means of a coupled ultrasound-micromechanics approach Materials Letters , 78, 154-158

The extremely fine structure of vertebral cortex challenges reliable determination of the tissue's anisotropic elasticity, which is important for the spine's load carrying patterns often causing pain in patients. As a potential remedy, we here propose a combined experimental (ultrasonic) and modeling (micromechanics) approach. Longitudinal acoustic waves are sent in longitudinal (superior-inferior, axial) as well as transverse (circumferential) direction through millimeter-sized samples containing this vertebral cortex, and corresponding wave velocities agree very well with recently identified 'universal' compositional and acoustic characteristics (J Theor Biol 287:115, 2011), which are valid for a large data base comprising different bones from different species and different organs. This provides evidence that the 'universal' organization patterns inherent to all the bone tissues of the aforementioned data base also hold for vertebral bone. Consequently, an experimentally validated model covering the mechanical effects of this organization patterns (J Theor Biol 244:597, 2007, J Theor Biol 260:230, 2009) gives access to the complete elasticity tensor of human lumbar vertebral bone tissue, as a valuable input for structural analyses aiming at patient-specific fracture risk assessment, e.g. based on the Finite Element Method.

JTD Keywords: Human vertebra, Micromechanics, Tissue elasticity, Ultrasonics


Serra, T., Navarro, M., Planell, J. A., (2012). Fabrication and characterization of biodegradable composite scaffolds for tissue engineering Innovative Developments in Virtual and Physical Prototyping 5th International Conference on Advanced Research and Rapid Prototyping (ed. Margarida, T., Ferreira, D.), Taylor & Francis (Leiria, Portugal) VR@P, 67-72

In this study, polylactic acid (PLA) and polyethylene glycol (PEG) were combined with soluble CaP glass particles and processed by rapid prototyping to obtain fully biodegradable structures for Tissue Engineering applications. The obtained 3D biodegradable structures were characterized in terms of their architecture and mechanical properties. The scaffold morphology, internal micro-architecture and mechanical properties were evaluated using Scanning Electron Microscopy (SEM), micro-computed tomography (micro-CT) and mechanical testing, respectively. Well defined structures with pore size of 350-400μm (in the axial view), struts width of approximately 70-80μm, and a porosity ranging between 60-65% were obtained. The combination RP and PLA/PEG/CaP glass turned into promising fully degradable, mechanically stable, bioactive and biocompatible composite scaffolds for TE.

JTD Keywords: Axial view, Biodegradable composites, Composite scaffolds, Glass particles, Mechanically stable, Micro architectures, Micro computed tomography (micro-CT), Poly lactic acid, Scaffold morphology, Tissue engineering applications, Well-defined structures, Bioactive glass, Mechanical properties, Mechanical testing, Polyethylene glycols, Polymer blends, Rapid prototyping, Scaffolds (biology), Scanning electron microscopy, Computerized tomography


Castaño, O., Eltohamy, M., Kim, H. W., (2012). Electrospinning technology in tissue regeneration Nanotechnology in Regenerative Medicine - Methods and Protocols (Methods in Molecular Biology) (ed. Navarro, M., Planell, J. A.), Springer (New York, USA) 811, 127-140

Electrospinning is one of the most versatile and effective tools to produce nanostructured fibers in the biomedical science fields. The nanofibrous structure with diameters from tens to hundreds of nanometers largely mimics the native extracellular matrix (ECM) of many tissues. Thus far, a range of compositions including polymers and ceramics and their composites/hybrids have been successfully applied for generating electrospun nanofibers. Different processing tools in electrospinning set-ups and assemblies are currently developed to tune the morphology and properties of nanofibers. Herein, we demonstrate the electrospinning process and the electrospun biomaterials for specific use in tissue regeneration with some examples, involving different material combinations and fiber morphologies.

JTD Keywords: Ceramic, Composites, Electrospinning, Nanofi bers, Nanostructured fi bers, Polymer, Tissue regeneration


Navarro, M., Planell, J. A., (2012). Is nanotechnology the key to unravel and engineer biological processes? Nanotechnology in Regenerative Medicine - Methods and Protocols (Methods in Molecular Biology) (ed. Navarro, M., Planell, J. A.), Springer (New York, USA) 811, 1-16

Regenerative medicine is an emerging field aiming to the development of new reparative strategies to treat degenerative diseases, injury, and trauma through developmental pathways in order to rebuild the architecture of the original injured organ and take over its functionality. Most of the processes and interactions involved in the regenerative process take place at subcellular scale. Nanotechnology provides the tools and technology not only to detect, to measure, or to image the interactions between the different biomolecules and biological entities, but also to control and guide the regenerative process. The relevance of nanotechnology for the development of regenerative medicine as well as an overview of the different tools that contribute to unravel and engineer biological systems are presented in this chapter. In addition, general data about the social impact and global investment in nanotechnology are provided.

JTD Keywords: Nanotechnology, Regenerative medicine, Tissue engineering


Almendros, Isaac, Farre, Ramon, Planas, Anna M., Torres, Marta, Bonsignore, Maria R., Navajas, Daniel, Montserrat, Josep M., (2011). Tissue oxygenation in brain, muscle, and fat in a rat model of sleep apnea: Differential effect of obstructive apneas and intermittent hypoxia Sleep , 34, (8), 1127-1133

Study Objectives: To test the hypotheses that the dynamic changes in brain oxygen partial pressure (PtO(2)) in response to obstructive apneas or to intermittent hypoxia differ from those in other organs and that the changes in brain PtO(2) in response to obstructive apneas is a source of oxidative stress. Design: Prospective controlled animal study. Setting: University laboratory. Participants: 98 Sprague-Dawley rats. Interventions: Cerebral cortex, skeletal muscle, or visceral fat tissues were exposed in anesthetized animals subjected to either obstructive apneas or intermittent hypoxia (apneic and hypoxic events of 15 s each and 60 events/h) for 1 h. Measurements and Results: Arterial oxygen saturation (spO(2)) presented a stable pattern, with similar desaturations during both stimuli. The PtO(2) was measured by a microelectrode. During obstructive apneas, a fast increase in cerebral PtO(2) was observed (38.2 +/- 3.4 vs. 54.8 +/- 5.9 mm Hg) but not in the rest of tissues. This particular cerebral response was not found during intermittent hypoxia. The cerebral content of reduced glutathione was decreased after obstructive apneas (46.2% +/- 15.2%) compared to controls (100.0% +/- 14.7%), but not after intermittent hypoxia. This antioxidant consumption after obstructive apneas was accompanied by increased cerebral lipid peroxidation under this condition. No changes were observed for these markers in the other tissues. Conclusions: These results suggest the cerebral cortex could be protected in some way from hypoxic periods caused by obstructive apneas. The increased cerebral PtO(2) during obstructive apneas may, however, cause harmful effects (oxidative stress). The obstructive apnea model appears to be more adequate than the intermittent hypoxia model for studying brain changes associated with OSA.

JTD Keywords: Tissue oxygenation, Obstructive apnea, Intermittent hypoxia, Animal model, Oxidative stress


Bohner, M., Loosli, Y., Baroud, G., Lacroix, D., (2011). Commentary: Deciphering the link between architecture and biological response of a bone graft substitute Acta Biomaterialia 7, (2), 478-484

Hundreds of studies have been devoted to the search for the ideal architecture for bone scaffold. Despite these efforts, results are often contradictory, and rules derived from these studies are accordingly vague. In fact, there is enough evidence to postulate that ideal scaffold architecture does not exist. The aim of this document is to explain this statement and review new approaches to decipher the existing but complex link between scaffold architecture and in vivo response.

JTD Keywords: Biomaterial, Bone, Tissue engineering, Resorbable, Graft


Perut, F., Montufar, E. B., Ciapetti, G., Santin, M., Salvage, J., Traykova, T., Planell, J. A., Ginebra, M. P., Baldini, N., (2011). Novel soybean/gelatine-based bioactive and injectable hydroxyapatite foam: Material properties and cell response Acta Biomaterialia 7, (4), 1780-1787

Despite their known osteoconductivity, clinical use of calcium phosphate cements is limited both by their relatively slow rate of resorption and by rheological properties incompatible with injectability. Bone in-growth and material resorption have been improved by the development of porous calcium phosphate cements. However, injectable formulations have so far only been obtained through the addition of relatively toxic surfactants. The present work describes the response of osteoblasts to a novel injectable foamed bone cement based on a composite formulation including the bioactive foaming agents soybean and gelatine. The foaming properties of both defatted soybean and gelatine gels were exploited to develop a self-hardening soy/gelatine/hydroxyapatite composite foam able to retain porosity upon injection. After setting, the foamed paste produced a calcium-deficient hydroxyapatite scaffold, showing good injectability and cohesion as well as interconnected porosity after injection. The intrinsic bioactivity of soybean and gelatine was shown to favour osteoblast adhesion and growth. These findings suggest that injectable, porous and bioactive calcium phosphate cements can be produced for bone regeneration through minimally invasive surgery.

JTD Keywords: Calcium phosphate cement, Composite, Bone tissue engineering, Cell viability, Bioactivity


Byrne, Damien P., Lacroix, Damien, Prendergast, Patrick J., (2011). Simulation of fracture healing in the tibia: Mechanoregulation of cell activity using a lattice modeling approach Journal of Orthopaedic Research , 29, (10), 1496-1503

In this study, a three-dimensional (3D) computational simulation of bone regeneration was performed in a human tibia under realistic muscle loading. The simulation was achieved using a discrete lattice modeling approach combined with a mechanoregulation algorithm to describe the cellular processes involved in the healing process namely proliferation, migration, apoptosis, and differentiation of cells. The main phases of fracture healing were predicted by the simulation, including the bone resorption phase, and there was a qualitative agreement between the temporal changes in interfragmentary strain and bending stiffness by comparison to experimental data and clinical results. Bone healing was simulated beyond the reparative phase by modeling the transition of woven bone into lamellar bone. Because the simulation has been shown to work with realistic anatomical 3D geometry and muscle loading, it demonstrates the potential of simulation tools for patient-specific pre-operative treatment planning.

JTD Keywords: Tissue differentiation, Computational analysis, Mechanical conditions, Bone regeneration, Weight-bearing, Proliferation, Osteoblast, Stiffness, Ingrowth, Scaffold


Lacroix, Damien, Ramirez Patino, Juan Fernando, (2011). Finite Element Analysis of Donning Procedure of a Prosthetic Transfemoral Socket Annals of Biomedical Engineering , 39, (12), 2972-2983

Lower limb amputation is a severe psychological and physical event in a patient. A prosthetic solution can be provided but should respond to a patient-specific need to accommodate for the geometrical and biomechanical specificities. A new approach to calculate the stress-strain state at the interaction between the socket and the stump of five transfemoral amputees is presented. In this study the socket donning procedure is modeled using an explicit finite element method based on the patient-specific geometry obtained from CT and laser scan data. Over stumps the mean maximum pressure is 4 kPa (SD 1.7) and the mean maximum shear stresses are 1.4 kPa (SD 0.6) and 0.6 kPa (SD 0.3) in longitudinal and circumferential directions, respectively. Locations of the maximum values are according to pressure zones at the sockets. The stress-strain states obtained in this study can be considered more reliable than others, since there are normal and tangential stresses associated to the socket donning procedure.

JTD Keywords: Trans-tibial prosthesis, Knee residual limb, Pressure distribution, Transtibial amputees, Stump/socket interface, Mechanical conditions, Load-transfer, Soft-tissues, Stresses, Contact


Perez, R. A., Del Valle, S., Altankov, G., Ginebra, M. P., (2011). Porous hydroxyapatite and gelatin/hydroxyapatite microspheres obtained by calcium phosphate cement emulsion Journal of Biomedical Materials Research - Part B: Applied Biomaterials , 97B, (1), 156-166

Hydroxyapatite and hybrid gelatine/hydroxyapatite microspheres were obtained through a water in oil emulsion of a calcium phosphate cement (CPC). The setting reaction of the CPC, in this case the hydrolysis of alpha-tricalcium phosphate, was responsible for the consolidation of the microspheres. After the setting reaction, the microspheres consisted of an entangled network of hydroxyapatite crystals, with a high porosity and pore sizes ranging between 0.5 and 5 mu m. The size of the microspheres was tailored by controlling the viscosity of the hydrophobic phase, the rotation speed, and the initial powder size of the CPC. The incorporation of gelatin increased the sphericity of the microspheres, as well as their size and size dispersion. To assess the feasibility of using the microspheres as cell microcarriers, Saos-2 cells were cultured on the microspheres. Fluorescent staining, SEM studies, and LDH quantification showed that the microspheres were able to sustain cell growth. Cell adhesion and proliferation was significantly improved in the hybrid gelatin/hydroxyapatite microspheres as compared to the hydroxyapatite ones.

JTD Keywords: Calcium phosphate(s), Bone graft, Microspheres, Composite/hard tissue, Hydroxy(1)lapatite


del Rio, Jose Antonio, Soriano, Eduardo, (2010). Regenerating cortical connections in a dish: the entorhino-hippocampal organotypic slice co-culture as tool for pharmacological screening of molecules promoting axon regeneration Nature Protocols 5, (2), 217-226

We present a method for using long-term organotypic slice co-cultures of the entorhino-hippocampal formation to analyze the axon-regenerative properties of a determined compound. The culture method is based on the membrane interphase method, which is easy to perform and is generally reproducible. The degree of axonal regeneration after treatment in lesioned cultures can be seen directly using green fluorescent protein (GFP) transgenic mice or by axon tracing and histological methods. Possible changes in cell morphology after pharmacological treatment can be determined easily by focal in vitro electroporation. The well-preserved cytoarchitectonics in the co-culture facilitate the analysis of identified cells or regenerating axons. The protocol takes up to a month.

JTD Keywords: Cajal-retzius cells, Green-fluorescent-protein, In-vitro model, Rat hippocampus, Nervous-tissue, Brain-slices, Dentate gyrus, Gene-transfer, Cultures, Damage


Santoro, R., Olivares, A. L., Brans, G., Wirz, D., Longinotti, C., Lacroix, D., Martin, I., Wendt, D., (2010). Bioreactor based engineering of large-scale human cartilage grafts for joint resurfacing Biomaterials 31, (34), 8946-8952

Apart from partial or total joint replacement, no surgical procedure is currently available to treat large and deep cartilage defects associated with advanced diseases such as osteoarthritis. In this work, we developed a perfusion bioreactor system to engineer human cartilage grafts in a size with clinical relevance for unicompartmental resurfacing of human knee joints (50 mm diameter x 3 mm thick). Computational fluid dynamics models were developed to optimize the flow profile when designing the perfusion chamber. Using the developed system, human chondrocytes could be seeded throughout large 50 mm diameter scaffolds with a uniform distribution. Following two weeks culture, tissues grown in the bioreactor were viable and homogeneously cartilaginous, with biomechanical properties approaching those of native cartilage. In contrast, tissues generated by conventional manual production procedures were highly inhomogeneous and contained large necrotic regions. The unprecedented engineering of human cartilage tissues in this large-scale opens the practical perspective of grafting functional biological substitutes for the clinical treatment for extensive cartilage defects, possibly in combination with surgical or pharmacological therapies to support durability of the implant. Ongoing efforts are aimed at integrating the up-scaled bioreactor based processes within a fully automated and closed manufacturing system for safe, standardized, and GMP compliant production of large-scale cartilage grafts.

JTD Keywords: Bioreactor, Cartilage repair, Computational fluid dynamics, Scale-up, Regenerative medicine, Tissue engineering


Sandino, C., Checa, S., Prendergast, P. J., Lacroix, D., (2010). Simulation of angiogenesis and cell differentiation in a CaP scaffold subjected to compressive strains using a lattice modeling approach Biomaterials 31, (8), 2446-2452

Mechanical stimuli are one of the factors that influence tissue differentiation. In the development of biomaterials for bone tissue engineering, mechanical stimuli and formation of a vascular network that transport oxygen to cells within the pores of the scaffolds are essential. Angiogenesis and cell differentiation have been simulated in scaffolds of regular porosity; however, the dynamics of differentiation can be different when the porosity is not uniform. The objective of this study was to investigate the effect of the mechanical stimuli and the capillary network formation on cell differentiation within a scaffold of irregular morphology. A porous scaffold of calcium phosphate based glass was used. The pores and the solid phase were discretized using micro computed tomography images. Cell activity was simulated within the interconnected pore domain of the scaffold using a lattice modeling approach. Compressive strains of 0.5 and 1% of total deformation were applied and two cases of mesenchymal stem cells initialization (in vitro seeding and in vivo) were simulated. Similar capillary networks were formed independently of the cell initialization mode and the magnitude of the mechanical strain applied. Most of vessels grew in the pores at the periphery of the scaffolds and were blocked by the walls of the scaffold. When 0.5% of strain was applied, 70% of the pore volume was affected by mechano-regulatory stimuli corresponding to bone formation; however, because of the lack of oxygen, only 40% of the volume was filled with osteoblasts. 40% of volume was filled with chondrocytes and 3% with fibroblasts. When the mechanical strain was increased to 1%, 11% of the pore volume was filled with osteoblasts, 59% with chondrocytes, and 8% with fibroblasts. This study has shown the dynamics of the correlation between mechanical load, angiogenesis and tissue differentiation within a scaffold with irregular morphology.

JTD Keywords: Tissue engineering, Calcium phosphates, Mechanoregulation, Micro computer tomography, Finite element modeling


Milan, J. L., Planell, J. A., Lacroix, D., (2010). Simulation of bone tissue formation within a porous scaffold under dynamic compression Biomechanics and Modeling in Mechanobiology 9, (5), 583-596

A computational model of mechanoregulation is proposed to predict bone tissue formation stimulated mechanically by overall dynamical compression within a porous polymeric scaffold rendered by micro-CT. Dynamic compressions of 0.5-5% at 0.0025-0.025 s(-1) were simulated. A force-controlled dynamic compression was also performed by imposing a ramp of force from 1 to 70 N. The model predicts homogeneous mature bone tissue formation under strain levels of 0.5-1% at strain rates of 0.0025-0.005 s(-1). Under higher levels of strain and strain rates, the scaffold shows heterogeneous mechanical behaviour which leads to the formation of a heterogeneous tissue with a mixture of mature bone and fibrous tissue. A fibrous tissue layer was also predicted under the force-controlled dynamic compression, although the same force magnitude was found promoting only mature bone during a strain-controlled compression. The model shows that the mechanical stimulation of bone tissue formation within a porous scaffold closely depends on the loading history and on the mechanical behaviour of the scaffold at local and global scales.

JTD Keywords: Bone tissue engineering, Scaffold, Tissue differentiation, Mechanoregulation, Finite element analysis


Koch, M. A., Vrij, E. J., Engel, E., Planell, J. A., Lacroix, D., (2010). Perfusion cell seeding on large porous PLA/calcium phosphate composite scaffolds in a perfusion bioreactor system under varying perfusion parameters Journal of Biomedical Materials Research - Part A , 95A, (4), 1011-1018

A promising approach to bone tissue engineering lies in the use of perfusion bioreactors where cells are seeded and cultured on scaffolds under conditions of enhanced nutrient supply and removal of metabolic products. Fluid flow alterations can stimulate cell activity, making the engineering of tissue more efficient. Most bioreactor systems are used to culture cells on thin scaffold discs. In clinical use, however, bone substitutes of large dimensions are needed. In this study, MG63 osteoblast-like cells were seeded on large porous PLA/glass scaffolds with a custom developed perfusion bioreactor system. Cells were seeded by oscillating perfusion of cell suspension through the scaffolds. Applicable perfusion parameters for successful cell seeding were determined by varying fluid flow velocity and perfusion cycle number. After perfusion, cell seeding, the cell distribution, and cell seeding efficiency were determined. A fluid flow velocity of 5 mm/s had to be exceeded to achieve a uniform cell distribution throughout the scaffold interior. Cell seeding efficiencies of up to 50% were achieved. Results suggested that perfusion cycle number influenced cell seeding efficiency rather than fluid flow velocities. The cell seeding conducted is a promising basis for further long term cell culture studies in large porous scaffolds.

JTD Keywords: Bioreactor, Bone tissue engineering, Scaffolds, In vitro


Salmeron-Sanchez, M., Altankov, G., (2010). Cell-Protein-Material interaction in tissue engineering Tissue Engineering (ed. Eberli, D.), Intech (Vukovar, Croatia) , 77-102

The initial cellular events that take place at the biomaterials interface mimic to a certain extent the natural adhesive interaction of cells with the extracellular matrix (ECM) (Spie, 2002; Griffin & Naughton, 2002; Grinnell, 1986). In fact, the living cells cannot interact directly with foreign materials, but they readily attach to the adsorbed layer of proteins (upon contact with physiological fluids in vivo or culture medium in vitro) such as fibronectin (FN), vitronectin (VN), fibrinogen (FG), representing the so-called soluble matrix proteins in the biological fluids (Grinnell 1986).

JTD Keywords: Tissue Engineering, Protein-material interaction, ECM, Biomaterials


Prendergast, P. J., Checa, S., Lacroix, D., (2010). Computational models of tissue differentiation Computational Modeling in Biomechanics (ed. Suvranu De, Farshid Guilak, Mohammad R. K. Mofrad), Springer-Verlag Berlin (Berlin) 3, 353-372

Readers of this chapter will learn about our approach to computer simulation of tissue differentiation in response to mechanical forces. It involves defining algorithms for mechanoregulation of each of following cell activities: proliferation, apoptosis, migration, and differentiation using a stimulus based on a combination of strain and fluid flow (Prendergast et al., J. Biomech., 1997) - algorithms are based on a lattice-modelling which also facilitates building algorithms for complex processes such as angiogenesis. The algorithms are designed to be collaboratable individually. They can be combined to create a computational simulation method for tissue differentiation, using finite element analysis to compute the mechanical stimuli in even quite complex biomechanical environments. Examples are presented of the simulation method in use.

JTD Keywords: Mechanobiology, Lattice modeling, Differentiation, Tissue engineering, Finite element modeling, Scaffolds


Jang, J. H., Castano, O., Kim, H. W., (2009). Electrospun materials as potential platforms for bone tissue engineering Advanced Drug Delivery Reviews 61, (12), 1065-1083

Nanofibrous materials produced by electrospinning processes have attracted considerable interest in tissue regeneration, including bone reconstruction. A range of novel materials and processing tools have been developed to mimic the native bone extracellular matrix for potential applications as tissue engineering scaffolds and ultimately to restore degenerated functions of the bone. Degradable polymers, bioactive inorganics and their nanocomposites/hybrids nanofibers with suitable mechanical properties and bone bioactivity for osteoblasts and progenitor/stem cells have been produced. The surface functionalization with apatite minerals and proteins/peptides as well as drug encapsulation within the nanofibers is a promising strategy for achieving therapeutic functions with nanofibrous materials. Recent attempts to endow a 3D scaffolding technique to the electrospinning regime have shown some promise for engineering 3D tissue constructs. With the improvement in knowledge and techniques of bone-targeted nanofibrous matrices, bone tissue engineering is expected to be realized in the near future.

JTD Keywords: Electrospun nanofiber, Bone tissue engineering, Biomimetic matrix, Bone bioactivity, 3D scaffolding


Milan, J. L., Planell, J. A., Lacroix, D., (2009). Computational modelling of the mechanical environment of osteogenesis within a polylactic acid-calcium phosphate glass scaffold Biomaterials 30, (25), 4219-4226

A computational model based on finite element method (FEM) and computational fluid dynamics (CFD) is developed to analyse the mechanical stimuli in a composite scaffold made of polylactic acid (PLA) matrix with calcium phosphate glass (Glass) particles. Different bioreactor loading conditions were simulated within the scaffold. In vitro perfusion conditions were reproduced in the model. Dynamic compression was also reproduced in an uncoupled fluid-structure scheme: deformation level was studied analyzing the mechanical response of scaffold alone under static compression while strain rate was studied considering the fluid flow induced by compression through fixed scaffold. Results of the model show that during perfusion test an inlet velocity of 25mum/s generates on scaffold surface a fluid flow shear stress which may stimulate osteogenesis. Dynamic compression of 5% applied on the PLA-Glass scaffold with a strain rate of 0.005s(-1) has the benefit to generate mechanical stimuli based on both solid shear strain and fluid flow shear stress on large scaffold surface area. Values of perfusion inlet velocity or compression strain rate one order of magnitude lower may promote cell proliferation while values one order of magnitude higher may be detrimental for cells. FEM-CFD scaffold models may help to determine loading conditions promoting bone formation and to interpret experimental results from a mechanical point of view.

JTD Keywords: Bone tissue engineering, Scaffold, Finite element analysis, Computational fluid dynamics, Mechanical stimuli


Olivares, A. L., Marshal, E., Planell, J. A., Lacroix, D., (2009). Finite element study of scaffold architecture design and culture conditions for tissue engineering Biomaterials 30, (30), 6142-6149

Tissue engineering scaffolds provide temporary mechanical support for tissue regeneration and transfer global mechanical load to mechanical stimuli to cells through its architecture. In this study the interactions between scaffold pore morphology, mechanical stimuli developed at the cell microscopic level, and culture conditions applied at the macroscopic scale are studied on two regular scaffold structures. Gyroid and hexagonal scaffolds of 55% and 70% porosity were modeled in a finite element analysis and were submitted to an inlet fluid flow or compressive strain. A mechanoregulation theory based on scaffold shear strain and fluid shear stress was applied for determining the influence of each structures on the mechanical stimuli on initial conditions. Results indicate that the distribution of shear stress induced by fluid perfusion is very dependent on pore distribution within the scaffold. Gyroid architectures provide a better accessibility of the fluid than hexagonal structures. Based on the mechanoregulation theory, the differentiation process in these structures was more sensitive to inlet fluid flow than axial strain of the scaffold. This study provides a computational approach to determine the mechanical stimuli at the cellular level when cells are cultured in a bioreactor and to relate mechanical stimuli with cell differentiation.

JTD Keywords: Tissue engineering, Scaffold, Rapid prototyping, Computational fluid dynamics, Finite element


Fernandez, Javier G., Mills, C. A., Samitier, J., (2009). Complex microstructured 3D surfaces using chitosan biopolymer Small 5, (5), 614-620

A technique for producing micrometer-scale structures over large, nonplanar chitosan surfaces is described. The technique makes use of the rheological characteristics (deformability) of the chitosan to create freestanding, three-dimensional scaffolds with controlled shapes, incorporating defined microtopography. The results of an investigation into the technical limits of molding different combinations of shapes and microtopographies are presented, highlighting the versatility of the technique when used irrespectively with inorganic or delicate organic moulds. The final, replicated scaffolds presented here are patterned with arrays of one-micrometer-tall microstructures over large areas. Structural integrity is characterized by the measurement of structural degradation. Human umbilical vein endothelial cells cultured on a tubular scaffold show that early cell growth is conditioned by the microtopography and indicate possible uses for the structures in biomedical applications. For those applications requiring improved chemical and mechanical resistance, the structures can be replicated in poly(dimethyl siloxane).

JTD Keywords: Biocompatible Materials/ chemistry, Cell Adhesion, Cell Culture Techniques/ methods, Cell Proliferation, Cells, Cultured, Chitosan/ chemistry, Crystallization/methods, Endothelial Cells/ cytology/ physiology, Humans, Materials Testing, Nanostructures/ chemistry/ ultrastructure, Nanotechnology/methods, Particle Size, Surface Properties, Tissue Engineering/methods


Lacroix, D., Planell, J. A., Prendergast, P. J., (2009). Computer-aided design and finite-element modelling of biomaterial scaffolds for bone tissue engineering Philosophical Transactions of the Royal Society A-Mathematical Physical and Engineering Sciences , 367, (1895), 1993-2009

Scaffold biomaterials for tissue engineering can be produced in many different ways depending on the applications and the materials used. Most research into new biomaterials is based on an experimental trial-and-error approach that limits the possibility of making many variations to a single material and studying its interaction with its surroundings. Instead, computer simulation applied to tissue engineering can offer a more exhaustive approach to test and screen out biomaterials. In this paper, a review of the current approach in biomaterials designed through computer-aided design (CAD) and through finite-element modelling is given. First we review the approach used in tissue engineering in the development of scaffolds and the interactions existing between biomaterials, cells and mechanical stimuli. Then, scaffold fabrication through CAD is presented and characterization of existing scaffolds through computed images is reviewed. Several case studies of finite-element studies in tissue engineering show the usefulness of computer simulations in determining the mechanical environment of cells when seeded into a scaffold and the proper design of the geometry and stiffness of the scaffold. This creates a need for more advanced studies that include aspects of mechanobiology in tissue engineering in order to be able to predict over time the growth and differentiation of tissues within scaffolds. Finally, current perspectives indicate that more efforts need to be put into the development of such advanced studies, with the removal of technical limitations such as computer power and the inclusion of more accurate biological and genetic processes into the developed algorithms.

JTD Keywords: Biomechanics, Tissue engineering, Biomaterials, Finite-element modelling


Engel, E., Michiardi, A., Navarro, M., Lacroix, D., Planell, J. A., (2008). Nanotechnology in regenerative medicine: the materials side Trends in Biotechnology , 26, (1), 39-47

Regenerative medicine is an emerging multidisciplinary field that aims to restore, maintain or enhance tissues and hence organ functions. Regeneration of tissues can be achieved by the combination of living cells, which will provide biological functionality, and materials, which act as scaffolds to support cell proliferation. Mammalian cells behave in vivo in response to the biological signals they receive from the surrounding environment, which is structured by nanometre-scaled components. Therefore, materials used in repairing the human body have to reproduce the correct signals that guide the cells towards a desirable behaviour. Nanotechnology is not only an excellent tool to produce material structures that mimic the biological ones but also holds the promise of providing efficient delivery systems. The application of nanotechnology to regenerative medicine is a wide issue and this short review will only focus on aspects of nanotechnology relevant to biomaterials science. Specifically, the fabrication of materials, such as nanoparticles and scaffolds for tissue engineering, and the nanopatterning of surfaces aimed at eliciting specific biological responses from the host tissue will be addressed.

JTD Keywords: Animals, Biocompatible Materials/ metabolism, Humans, Nanoparticles, Nanotechnology/ methods, Regenerative Medicine/ methods, Tissue Scaffolds


Engel, E., Del Valle, S., Aparicio, C., Altankov, G., Asin, L., Planell, J. A., Ginebra, M. P., (2008). Discerning the role of topography and ion exchange in cell response of bioactive tissue engineering scaffolds Tissue Engineering Part A , 14, (8), 1341-1351

Surface topography is known to have an influence on osteoblast activity. However, in the case of bioactive materials, topographical changes can affect also ion exchange properties. This makes the problem more complex, since it is often difficult to separate the strictly topographical effects from the effects of ionic fluctuations in the medium. The scope of this paper is to analyze the simultaneous effect of topography and topography-mediated ion exchange on the initial cellular behavior of osteoblastic-like cells cultured on bioactive tissue engineering substrates. Two apatitic substrates with identical chemical composition but different micro/nanostructural features were obtained by low-temperature setting of a calcium phosphate cement. MG63 osteoblastic-like cells were cultured either in direct contact with the substrates or with their extracts. A strong and permanent decrease of calcium concentration in the culture medium, dependent on substrate topography, was detected. A major effect of the substrate microstructure on cell proliferation was observed, explained in part by the topography-mediated ion exchange, but not specifically by the ionic Ca(2+) fluctuations. Cell differentiation was strongly enhanced when cells were cultured on the finer substrate. This effect was not explained by the chemical modification of the medium, but rather suggested a strictly topographical effect.

JTD Keywords: Alkaline Phosphatase/metabolism, Bone Cements/pharmacology, Calcium/metabolism, Calcium Phosphates/pharmacology, Cell Adhesion/drug effects, Cell Differentiation/drug effects, Cell Proliferation/drug effects, Cell Shape/drug effects, Cells, Cultured, Culture Media, Durapatite/pharmacology, Humans, Interferometry, Ion Exchange, Materials Testing, Osteoblasts/ cytology/drug effects/enzymology/ultrastructure, Phosphorus/metabolism, Powders, Tissue Engineering, Tissue Scaffolds


Navarro, M., Michiardi, A., Castano, O., Planell, J. A., (2008). Biomaterials in orthopaedics Journal of the Royal Society Interface , 5, (27), 1137-1158

At present, strong requirements in orthopaedics are still to be met, both in bone and joint substitution and in the repair and regeneration of bone defects. In this framework, tremendous advances in the biomaterials field have been made in the last 50 years where materials intended for biomedical purposes have evolved through three different generations, namely first generation (bioinert materials), second generation (bioactive and biodegradable materials) and third generation (materials designed to stimulate specific responses at the molecular level). In this review, the evolution of different metals, ceramics and polymers most commonly used in orthopaedic applications is discussed, as well as the different approaches used to fulfil the challenges faced by this medical field.

JTD Keywords: Biomaterials, Orthopaedics, Tissue engineering, Bioactive materials, Biodegradable materials, Bioinert materials


Sandino, C., Planell, J. A., Lacroix, D., (2008). A finite element study of mechanical stimuli in scaffolds for bone tissue engineering Journal of Biomechanics 41, (5), 1005-1014

Mechanical stimuli are one of the factors that affect cell proliferation and differentiation in the process of bone tissue regeneration. Knowledge on the specific deformation sensed by cells at a microscopic level when mechanical loads are applied is still missing in the development of biomaterials for bone tissue engineering. The objective of this study was to analyze the behavior of the mechanical stimuli within some calcium phosphate-based scaffolds in terms of stress and strain distributions in the solid material phase and fluid velocity, fluid pressure and fluid shear stress distributions in the pores filled of fluid, by means of micro computed tomographed (CT)-based finite element (FE) models. Two samples of porous materials, one of calcium phosphate-based cement and another of biodegradable glass, were used. Compressive loads equivalent to 0.5% of compression applied to the solid material phase and interstitial fluid flows with inlet velocities of 1, 10 and 100 mu m/s applied to the interconnected pores were simulated, changing also the inlet side and the viscosity of the medium. Similar strain distributions for both materials were found, with compressive and tensile strain maximal values of 1.6% and 0.6%, respectively. Mean values were consistent with the applied deformation. When 10 mu m/s of inlet fluid velocity and 1.45 Pa s viscosity, maximal values of fluid velocity were 12.76 mm/s for CaP cement and 14.87 mm/s for glass. Mean values were consistent with the inlet ones applied, and mean values of shear stress were around 5 x 10(-5) Pa. Variations on inlet fluid velocity and fluid viscosity produce proportional and independent changes in fluid velocity, fluid shear stress and fluid pressure. This study has shown how mechanical loads and fluid flow applied on the scaffolds cause different levels of mechanical stimuli within the samples according to the morphology of the materials.

JTD Keywords: Bone tissue engineering, Finite element analysis, Scaffolds, Mechanical stimuli


Charles-Harris, M., Koch, M. A., Navarro, M., Lacroix, D., Engel, E., Planell, J. A., (2008). A PLA/calcium phosphate degradable composite material for bone tissue engineering: an in vitro study Journal of Materials Science-Materials in Medicine , 19, (4), 1503-1513

Biodegradable polymers reinforced with an inorganic phase such as calcium phosphate glasses may be a promising approach to fulfil the challenging requirements presented by 3D porous scaffolds for tissue engineering. Scaffolds' success depends mainly on their biological behaviour. This work is aimed to the in vitro study of polylactic acid (PLA)/CaP glass 3D porous constructs for bone regeneration. The scaffolds were elaborated using two different techniques, namely solvent-casting and phase-separation. The effect of scaffolds' micro and macrostructure on the biological response of these scaffolds was assayed. Cell proliferation, differentiation and morphology within the scaffolds were studied. Furthermore, polymer/glass scaffolds were seeded under dynamic conditions in a custom-made perfusion bioreactor. Results indicate that the final architecture of the solvent-cast or phase separated scaffolds have a significant effect on cells' behaviour. Solvent-cast scaffolds seem to be the best candidates for bone tissue engineering. Besides, dynamic seeding yielded a higher seeding efficiency in comparison with the static method.

JTD Keywords: Biocompatible Materials/ chemistry, Bone and Bones/ metabolism, Calcium Phosphates/ chemistry, Cell Differentiation, Cell Proliferation, Humans, Lactic Acid/ chemistry, Microscopy, Confocal, Microscopy, Electron, Scanning, Osteoblasts/metabolism, Permeability, Polymers/ chemistry, Porosity, Solvents/chemistry, Tissue Engineering/ methods


Montufar, E. B., Gil, C., Traykova, T., Ginebra, M. P., Planell, J., (2008). Foamed beta-tricalcium phosphate scaffolds Bioceramics: Key Engineering Materials 20th International Symposium on Ceramics in Medicine (ed. Daculsi, G., Layrolle, P.), Trans Tech Publications Ltd (Nantes, France) 20, 323-326

The design and processing of 3D macroporous bioactive scaffolds is one of the milestones for the progress of bone tissue engineering and bone regeneration. Calcium phosphate based ceramics are among the most suitable materials, due to their similarity to the bone mineral. Specifically, beta-tricalcium phosphate (beta-TCP) is known to be a resorbable and bioactive material, with well established applications as bone regeneration material. The aim of this work is to explore a new OF route to obtain beta-TCP macroporous scaffolds starting from calcium phosphate cements. To this end foamed calcium phosphate cement.. composed of alpha tricalcium phosphate as starting powder was used as initial material. The set foamed structures, made of calcium deficient hydroxyapatite (CDHA) were sintered to obtain the final beta-TCP macroporous architecture. The interconnected macroporosity was maintained.. whereas the porosity in the nanometric range was strongly reduced by the sintering process. The sintering produced also an increase in the mechanical properties of the scaffold.

JTD Keywords: Calcium-phosphate ceramics, Cements, Scaffolds, Foams, Macroporous, Tissue engineering


Koch, M. A., Engel, E., Planell, J. A., Lacroix, D., (2008). Cell seeding and characterisation of PLA/glass composite scaffolds for bone tissue engineering Journal of Biomechanics 16th Congress, European Society of Biomechanics , Elsevier (Lucerne, Switzerland) 41, (Supplement 1), S162

In this study polymer-glass composite scaffolds were characterized by permeability and porosity, two important properties for the use in perfusion bioreactors. These scaffolds were seeded with osteoblast-like cells to assess the efficiency of the used bioreactor. The used PLA/glass composite scaffolds are adequate for the perfusion culture. The high porosity and pore interconnectivity allow an even cell distribution and incorporation of a high cell number. For optimisation of the perfusion bioreactor system, further research has to be dedicated to the cell seeding and culture.

JTD Keywords: Biomedical materials, Bioreactors, Bone, Cellular biophysics, Composite materials, Orthopaedics, Permeability, Polymers, Porosity, Porous materials, Tissue engineering