Publications

Porous zirconia scaffolds manufactured using polymer-infiltrated ceramic network (PICN) and additive manufacturing technologies are emerging as promising alternatives to traditional ceramic materials in dental restorations. However, incomplete osseointegration and bacterial infections still represent challenges for the long-term performance of this new composite material. To address this, the present study aims to investigate the effect of peptide biofunctionalization on the biological performance of infiltrated zirconia scaffold surfaces. The samples used in the work consisted of a 3D-printed zirconia scaffold infiltrated with a dimethacrylate copolymer. Surface biofunctionalization was achieved using a synthetic platform containing the cell-adhesive sequence RGD and the antibacteria LF1-11 peptide (RGD-LF). The attachment of the molecule was characterized through fluorescence confocal laser scanning microscopy and X-ray photoelectron spectroscopy. The biological performance of the samples was evaluated in terms of human mesenchymal stem cell adhesion and early attachment of S. aureus. The physicochemical characterization verified the successful anchoring of the biomolecule to the surface, leading to a peptide density of 288 pmol/cm2. 2 . The biological assays confirmed the potential of RGD-LF to improve cell adhesion and spreading. In this sense, the average cell area increased fourfold in the biofunctionalized surface. Regarding bacterial adhesion, it was demonstrated that RGD-LF significantly inhibited it, reducing early adhesion by half compared to the untreated surface. Overall, this study provides valuable insights into the biofunctionalization of polymer-infiltrated 3D scaffolds for the development of cell-instructive and antibacterial surfaces tailored for dental applications.

JTD Keywords: Antibacteria, Biocompatibility, Cell adhesion, Composite, Dental ceramics, Direct ink writing, Fabrication, Infiltrated zirconia scaffolds, Mechanical-properties, Peptide biofunctionalization, Picn material, Strength, Surface, Topograph, Wear behavior

Hydrogels with cell adhesive moieties stand out as promising materials to enhance tissue healing and regeneration. Nonetheless, bacterial infections of the implants represent an unmet major concern. In the present work, we developed an alginate hydrogel modified with a multifunctional peptide containing the RGD cell adhesive motif in combination with an antibacterial peptide derived from the 1-11 region of lactoferrin (LF). The RGD-LF branched peptide was successfully anchored to the alginate backbone by carbodiimide chemistry, as demonstrated by 1H NMR and fluorescence measurements. The functionalized hydrogel presented desirable physicochemical properties (porosity, swelling and rheological behavior) to develop biomaterials for tissue engineering. The viability of mesenchymal stem cells (MSCs) on the peptide-functionalized hydrogels was excellent, with values higher than 85 % at day 1, and higher than 95 % after 14 days in culture. Moreover, the biological characterization demonstrated the ability of the hydrogels to significantly enhance ALP activity of MSCs as well as to decrease bacterial colonization of both Gram-positive and Gram-negative models. Such results prove the potential of the functionalized hydrogels as novel biomaterials for tissue engineering, simultaneously displaying cell adhesive activity and the capacity to prevent bacterial contamination, a dual bioactivity commonly not found for these types of hydrogels. In this work we report on the functionalization of an alginate hydrogel with a tailor-made multifunctional peptide containing the cell adhesive RGD motif and the LF1-11 antibacterial peptide. Such novel multifunctional biomaterial ensures the viability of human mesenchymal stem cells, enhances ALP activity and decreases bacterial infections of both Gram-positive and Gram-negative models. image

JTD Keywords: Alginate hydrogel, Alginates, Anti-bacterial agents, Antimicrobial peptid, Antimicrobial peptide, Antimicrobial peptides, Arginyl-glycyl-aspartic acid, Biocompatible materials, Biofunctionalization, Bone, Cell adhesion, Cell survival, Composite hydrogels, Cross-linking, Hlf1-11 peptide, Human lactoferrin, Humans, Hydrogels, Immobilization, Mesenchymal stem cells, Multifunctional peptide, Oligopeptides, Peptides, Physical-properties, Scaffolds, Surfac, Tissue engineering

The 3D printing of titanium (Ti) offers countless possibilities for the development of personalized implants with suitable mechanical properties for different medical applications. However, the poor bioactivity of Ti is still a challenge that needs to be addressed to promote scaffold osseointegration. The aim of the present study was to functionalize Ti scaffolds with genetically modified elastin-like recombinamers (ELRs), synthetic polymeric proteins containing the elastin epitopes responsible for their mechanical properties and for promoting mesenchymal stem cell (MSC) recruitment, proliferation, and differentiation to ultimately increase scaffold osseointegration. To this end, ELRs containing specific cell-adhesive (RGD) and/or osteoinductive (SNA15) moieties were covalently attached to Ti scaffolds. Cell adhesion, proliferation, and colonization were enhanced on those scaffolds functionalized with RGD-ELR, while differentiation was promoted on those with SNA15-ELR. The combination of both RGD and SNA15 into the same ELR stimulated cell adhesion, proliferation, and differentiation, although at lower levels than those for every single moiety. These results suggest that biofunctionalization with SNA15-ELRs could modulate the cellular response to improve the osseointegration of Ti implants. Further investigation on the amount and distribution of RGD and SNA15 moieties in ELRs could improve cell adhesion, proliferation, and differentiation compared to the present study.

JTD Keywords: 3d printing, adhesion, biofunctionalization, elastin-like recombinamers, functionalization, hydroxyapatite, osseointegration, polymers, purification, technology, titanium, 3d printing, Surfaces, Titanium

Cell-material interactions are regulated by mimicking bone extracellular matrix on the surface of biomaterials. In this regard, reproducing the extracellular conditions that promote integrin and growth factor (GF) signaling is a major goal to trigger bone regeneration. Thus, the use of synthetic osteogenic domains derived from bone morphogenetic protein 2 (BMP-2) is gaining increasing attention, as this strategy is devoid of the clinical risks associated with this molecule. In this work, the wrist and knuckle epitopes of BMP-2 are screened to identify peptides with potential osteogenic properties. The most active sequences (the DWIVA motif and its cyclic version) are combined with the cell adhesive RGD peptide (linear and cyclic variants), to produce tailor-made biomimetic peptides presenting the bioactive cues in a chemically and geometrically defined manner. Such multifunctional peptides are next used to functionalize titanium surfaces. Biological characterization with mesenchymal stem cells demonstrates the ability of the biointerfaces to synergistically enhance cell adhesion and osteogenic differentiation. Furthermore, in vivo studies in rat calvarial defects prove the capacity of the biomimetic coatings to improve new bone formation and reduce fibrous tissue thickness. These results highlight the potential of mimicking integrin-GF signaling with synthetic peptides, without the need for exogenous GFs.© 2022 The Authors. Advanced Healthcare Materials published by Wiley-VCH GmbH.

JTD Keywords: adhesion formation, bmp-2, cell adhesions, in-vivo, integrin, mesenchymal stem-cells, morphogenetic protein-2, multifunctionality, osteoblastic differentiation, osteogenic differentiation, rgd-dwiva, rgd-peptides, titanium biofunctionalization, titanium surfaces, Animals, Biocompatible materials, Biomimetic peptides, Bone morphogenetic protein 2, Bone regeneration, Cell adhesions, Cell differentiation, Epitopes, Extracellular matrix, Integrins, Marrow stromal cells, Multifunctionality, Osteogenesis, Osteogenic differentiation, Peptides, Rats, Rgd-dwiva, Titanium, Titanium biofunctionalization

Endothelial coverage of an exposed cardiovascular stent surface leads to the occurrence of restenosis and late-stent thrombosis several months after implantation. To overcome this difficulty, modification of stent surfaces with topographical or biochemical features may be performed to increase endothelial cells’ (ECs) adhesion and/or migration. This work combines both strategies on cobalt-chromium (CoCr) alloy and studies the potential synergistic effect of linear patterned surfaces that are obtained by direct laser interference patterning (DLIP), coupled with the use of Arg-Gly-Asp (RGD) and Tyr-Ile-Gly-Ser-Arg (YIGSR) peptides. An extensive characterization of the modified surfaces was performed by using AFM, XPS, surface charge, electrochemical analysis and fluorescent methods. The biological response was studied in terms of EC adhesion, migration and proliferation assays. CoCr surfaces were successfully patterned with a periodicity of 10 µm and two different depths, D (≈79 and 762 nm). RGD and YIGSR were immobilized on the surfaces by CPTES silanization. Early EC adhesion was increased on the peptide-functionalized surfaces, especially for YIGSR compared to RGD. High-depth patterns generated 80% of ECs’ alignment within the topographical lines and enhanced EC migration. It is noteworthy that the combined use of the two strategies synergistically accelerated the ECs’ migration and proliferation, proving the potential of this strategy to enhance stent endothelialization.

JTD Keywords: adhesion, bare-metal, biofunctionalization, biomaterials, cell adhesive peptides, cobalt-chromium alloy, direct laser interference patterning (dlip), endothelial cell migration, functionalization, matrix, proliferation, selectivity, shear-stress, surfaces, Biofunctionalization, Cell adhesive peptides, Cobalt-chromium alloy, Direct laser interference patterning (dlip), Drug-eluting stents, Endothelial cell migration

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

STUDY QUESTION Is the attachment of biofunctionalized polysilicon barcodes to the outer surface of the zona pellucida an effective approach for the direct tagging and identification of cultured embryos? SUMMARY ANSWER The results achieved provide a proof of concept for a direct embryo tagging system using biofunctionalized polysilicon barcodes, which could help to minimize the risk of mismatching errors (mix-ups) in human assisted reproduction technologies. WHAT IS KNOWN ALREADY Even though the occurrence of mix-ups is rare, several cases have been reported in fertility clinics around the world. Measures to prevent the risk of mix-ups in human assisted reproduction technologies are therefore required. STUDY DESIGN, SIZE, DURATION Mouse embryos were tagged with 10 barcodes and the effectiveness of the tagging system was tested during fresh in vitro culture (n=140) and after embryo cryopreservation (n = 84). Finally, the full-term development of tagged embryos was evaluated (n =105). PARTICIPANTS/ MATERIALS, SETTING, METHODS Mouse pronuclear embryos were individually rolled over wheat germ agglutinin-biofunctionalized polysilicon barcodes to distribute them uniformly around the ZONA PELLUCIDA surface. Embryo viability and retention of barcodes were determined during 96 h of culture. The identification of tagged embryos was performed every 24 h in an inverted microscope and without embryo manipulation to simulate an automatic reading procedure. Full-term development of the tagged embryos was assessed after their transfer to pseudo-pregnant females. To test the validity of the embryo tagging system after a cryopreservation process, tagged embryos were frozen at the 2-cell stage using a slow freezing protocol, and followed in culture for 72 h after thawing. MAIN RESULTS AND THE ROLE OF CHANCE Neither the in vitro or in vivo development of tagged embryos was adversely affected. The tagging system also proved effective during an embryo cryopreservation process. Global identification rates higher than 96 and 92% in fresh and frozen-thawed tagged embryos, respectively, were obtained when simulating an automatic barcode reading system, although these rates could be increased to 100% by simply rotating the embryos during the reading process. LIMITATIONS, REASONS FOR CAUTION The direct embryo tagging developed here has exclusively been tested in mouse embryos. Its effectiveness in other species, such as the human, is currently being tested. WIDER IMPLICATIONS OF THE FINDINGS The direct embryo tagging system developed here, once tested in human embryos, could provide fertility clinics with a novel tool to reduce the risk of mix-ups in human assisted reproduction technologies. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by Spanish Ministry of Education and Science (TEC2011-29140-C03) and by the Generalitat de Catalunya (2009SGR-00282).

JTD Keywords: Assisted reproductive technologies (ART), Biofunctionalization, Embryo tagging, Mix-ups, Traceability