Institute for Bioengineering of Catalonia

PhD Discussions Sessions: Aida Baelo and Arnau Biosca
2018-02-23 10:00 - 23:00 (Europe/Madrid) | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
From the understanding to the treatment of biofilm wound infections
Aida Baelo, Bacterial infections: antimicrobial therapies group
Wounds represent a very common and serious health problem worldwide. The exposure of host tissue to the external environment allows the proliferation of a broad variety of pathogenic microorganisms, causing severe infections that are difficult to eradicate, such as diabetic foot ulcers, burn and surgery wounds. Bacteria infecting wounds arrange themselves in polymicrobial communities known as biofilms. The features displayed by the biofilm hinder and delay the healing processes, as such bacterial communities exhibit higher resistance to antibiotics and higher ability to evade the immune response. The understanding of the interaction between the components of this microbial community within the host is essential in order to develop new healing strategies that target bacteria growing in wounds. However, classic culture methods do not allow the simultaneous co-culture of different bacterial species, or the study in a more-realistic infection site environment, where several host factors are presented.
We use a novel in vitro culture approach, optimizing a method to assess bacterial viability in a wound biofilm model. This in vitro multispecies biofilm model resembles the natural conditions present in wounds and allows us to study a Pseudomonas aeruginosa and Staphylococcus aureus co-culture, which are the predominant bacteria found in wounds. We show that P. aeruginosa and S. aureus reach an equilibrium in the wound-like environment, with both microorganisms replicating under these conditions. As replication is a crucial step to initiate an infection, we have first focused on the study of the differential role of the different P. aeruginosa Ribonucleotide Reductase (RNR) enzymes in bacterial growth within the wound biofilm model, as RNR are essential enzymes in DNA replication. Also, we use a set of biofilm-degrading enzymes targeting the wound biofilm so as to improve the antibiotic delivery in the local area of the infection site.
New antimalarial strategies and drug delivery systems based on nanotechnology
Arnau Biosca, Nanomalaria joint group
Despite the undeniable importance of malaria elimination on the global research agenda, available front-line drugs are rapidly loosing efficacy. Thus, alternative therapeutic strategies working through radically new mechanism are urgently needed. Also, improving the delivery of old antimalarial compounds to decrease their side effects and avoid resistance appearance is a priority. In this work we study two new antimalarial strategies and two new delivery systems.
First, using a combinational approach that uses experimental data and bioinformatics, we are exploring the cytotoxicity of protein aggregation on Plasmodium falciparum parasites as a new antimalarial strategy. He have observed and purified highly insoluble protein aggregates form living parasites from which we have selected a list of potential protein candidates to be tested as antimalarial agents. Also, we are investigating the antimalarial properties of Domiphen bromide (DMB) a highly hydrophobic compound predicted to inhibit a key enzyme of the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway, absent in humans and found on the apicoplast organelle, a relic chloroplast of Plasmodium parasites.
Second, malaria eradication calls for strategies to reduce the transmission of the parasite from the human host to the mosquito vector. In this regard, and taking as a template the previously immunoliposome model developed in our group, we are now engineering a dual immunoliposome that targets gametocytes, the transmissible form of the parasite, and is loaded with two distinctive drugs. On the bilayer, the hydrophobic drug DMB is incorporated, and on the aqueous phase, the potent anti-gemetocidal compound, pyronoridine tetraphosphate, is actively encapsulated.
Finally, curcumin, a natural compound found in turmeric (Curcuma longa) presents promising antimalarial activity in vitro, but its low stability and intestinal absorption compromise their effectiveness in vivo. In this regard, we are working on the development of polymeric nanovectors for the improved abortion of curcumin in the intestinal mucosa.

PhD Discussions Sessions: Andreu Matamoros and Maider Badiola
2018-03-23 10:00 - 11:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Role of the Cellular Prion Protein in hippocampal neurotransmission, learning and memory
Andreu Matamoros, Molecular and cellular neurobiotechnology
MisfoldedCellular Prion protein (PrPC) was described as the causative agent of the transmissible spongiform encephalopathies (TSEs), independently of its origin (sporadic, iatrogenic or genetic). PrPC is present at the synaptic terminal, especially in the cerebral cortex including the hippocampus. It is involved in numerous cellular processes: cell proliferation and differentiation, copper homeostasis and cell signaling, among others. Recently has been demonstrated that most of these functions are based in misinterpretation of the mice models and need to be reevaluated. On the other hand, PrPC protein levels are decreased in TSEs. This opened a new insight in the study of TSEs: understanding the pathology not just as a gain of function due to the prion aggregation, but as a loss of function due to the reduction of PrPC.
Our goal is to elucidate the role of PrPC in hippocampal circuitry and its derived functions (i.e. learning and memory) using a new PrPC knockout mice (ZH3). Spontaneous firing and network formation are monitored with Calcium imaging in hippocampus primary cultures. Object Recognition Test and Skinner’s Test are performed to evaluate memory and learning in ZH3 mice. LTP and evocated potentials are also measured in CA3-CA1 connection in vivo. Glutamate neurotransmission is evaluated behaviourally and electrophysiologycally using kainate administration. Finally, mRNA from ZH3 mice hippocampus has been sequenced to identify differential gene expression compared to Wt mice.
Dissecting the role of PrPC in hippocampus neurotransmission will allow us to better understand alterations in the brain of TSEs patients.

Paving the way towards an in-vitro 3D mechanosensory-motor circuit on a chip
Maider Badiola, Nanobioengineering
Neuromuscular diseases (NMD) are neurological disorders affecting muscles and their control through nervous system. They often involve afferent and efferent pathways of the Peripheral Nervous System, and their effects might be reflected in the mechanosensory-motor circuit at different cellular levels (including sensory and motor neurons, glia and muscle dysfunctions), and in the connexion among them.
The aim of this research is to create an in-vitro model to mimic the 3D microenvironment of a neural circuit for locomotion to understand and find treatments for NMDs. To that end, organ-on-a-chip technologies are used for the integration of sensorial and motor neural components together with a functional muscular unit.
For that purpose, we first fabricated a compartmentalised microfluidic device in PDMS using soft lithography techniques. Then the afferent and efferent pathways of the Peripheral Nervous System were mimicked in 2D culturing primary neurons involved in the locomotion circuit (motoneurons and dorsal root ganglia) with Schwann cells in the microdevice.
But 2D cultures offer many limitations compared to 3D, and the assessment of the afferent pathway separately often means a complication. Optogenetics technique can be used in skeletal muscle to induce contraction, mimicking a natural innervation to some length and facilitating the study of the afferent pathway separately. Therefore, we propose a study model where primary spinal motor- or dorsal root ganglia sensory- neurons are cultured in 3D in different compartments together with optogenetically sensitive myocytes (a channelrhodopsin-2 positive cell line). This could make possible to evaluate the functionality of efferent and afferent pathways separately.
This study provides the basis for future steps towards NMD in-vitro study models.

PhD Discussions Sessions: Jesús Ordoño and Ernest Latorre
2018-06-01 10:00 - 11:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Lactate-based strategy for cardiac tissue engineering
Jesús Ordoño, Biomaterials for regenerative therapies
Lactate is a metabolite of glycolysis, commonly produced by cells consuming glucose. However, growing evidences suggest new roles for this molecule, as it has shown to act as a signalling molecule in many tissues. In this work, we explore the effects of lactate on cardiac cells for tissue engineering applications. Our results demonstrate that lactate enhance cardiomyocyte proliferation and modulates different cell cycle related proteins, supporting thus the idea that this molecule can be able to reprogram cardiomyocytes towards a more immature stage. Cardiac fibroblasts also show a dose-dependent response to lactate by modifying their secretome, hence promoting a suitable environment for cardiac regeneration. Ex vivo culture of mouse hearts revealed the ability of lactate to increase survival of cardiomyocytes as well as to prolong the beating capacity of the cardiac tissue.
With all these new evidences of the action of lactate, we cultured cardiac cells on a 3D scaffold based on collagen and elastin, allowing engraftment and beating of the cardiac tissue. The response of such system to external electrical stimulus was evaluated using a pulsatile electric field stimulation, showing a proliferative and more immature behaviour of the tissue in the presence of lactate. Cardiac cells also showed expression of specific lactate receptors and transporters, such as MCT1, MCT4 and GPR81. The correct development of sarcomeric structures was confirmed, as well as the coupling and presence of intercalated disks. In conclusion, lactate arises as a novel and feasible option to promote cardiac regeneration, and therefore lactate-releasing scaffolds are a suitable strategy for cardiac tissue regeneration.
Active superelasticity revealed by three-dimensional epithelial sheets of controlled size and shape
Ernest Latorre, Integrative cell and tissue dynamics
Fundamental processes in development and physiology are determined by the three-dimensional architecture of epithelial sheets. How these sheets deform and fold into complex structures has remained unclear, however, because their mechanical properties in three-dimensions have not been accessed experimentally. By combining measurements of epithelial tension, shape, and luminal pressure, here we show that epithelial cell sheets are active superelastic materials. We develop a new micropattering approach to produce massive arrays of epithelial domes with controlled basal shape and size. By measuring 3D deformations of the substrate and curvature of the dome we obtain a direct measurement of luminal pressure and epithelial tension. Observations over time-scales of hours allow us to map the epithelial tension-strain response, revealing a tensional plateau over several-fold areal strain reaching 300%. We show that these extreme nominal strains are accommodated by a highly heterogeneous stretching of individual cells, with barely deformed cells coexisting with others reaching 1000% areal strain, in seeming contradiction with the measured tensional uniformity. This phenomenology is reminiscent of superelasticity, a mechanical response generally attributed to microscopic material instabilities in metal alloys. We provide evidence that this instability is triggered in epithelial cells by limited availability of components of the actomyosin cortex. Finally, we implement 3D vertex model, which captures both the tension/strain relationship and strain heterogeneity. Our study unveils a new type of mechanical behavior -active superelasticity- that enables epithelial sheets to sustain extreme stretching under constant tension.

PhD Discussions Sessions: Helena Lozano and Martina Maier
2018-06-29 10:00 - 11:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Electrical and morphological characterization of bacterial polar flagella
Helena Lozano, Nanoscale bioelectrical characterization
The electric polarization of proteins in response to external electric fields plays an important role to understand the electrostatic interaction of proteins with charged biomolecules and ions [1]. Even if dielectric studies on individual proteins have not been reported, yet, there have been some studies on protein complexes involving a relatively small number of proteins, such as for instance, virus capsids and virus tails [2,3]. I am going to present the results of a dielectric study performed on a third protein complex system, the bacterial polar flagellum, which is composed of protein subunits called flagellin arranged in several intertwined chains [4]. In particular, we perform a comparative study of the dielectric properties of single flagella belonging to two different bacteria types, namely, Shewanella oneidensis MR-1 and Pseudomona aeruginosas PAO1. The dielectric properties (dielectric constant) have been obtained by combining Electrostatic Force Microscopy images [2] and 3D finite element numerical calculations. The values obtained are within the range of values obtained with macroscopic techniques [1].
References:
[1] Simonson, T., Rep. Prog. Phys. 66 (2003) 737–787.
[2] Fumagalli, L., Esteban-Ferrer, et al. Nature Materials 11 (2012) 743
[3] Cuervo, A., Dans, P. D., et al. PNAS 111 (2014) E3624.
[4] Lozano, H., Fábregas, R., Blanco-Cabra, N., Millán-Solsona, R. Torrents, E., Gomila, G. (in preparation).
Rehabilitation of cognitive deficits and depression after stroke
Martina Maier, SPECS
Cognitive deficits and depression are common consequences of stroke [1], [2]. Both have detrimental effects on quality of life and the activities of daily living [3]. In addition, they have been linked to poor functional outcome and more severe impairment [4], [5], [6] than observed in patients without cognitive deficit or depression. Post-stroke depression has been related to cognitive impairment [7], but the dynamics of that relationship are not well understood, as cognitive deficits and depression are typically studied and treated in isolation. The aim of our work is twofold. On one hand, we investigate in how depression modulates cognitive functioning after a stroke. On the other, we propose a new rehabilitation method that treats cognitive deficits and depression in conjunction. For this reason, we conducted a longitudinal randomized clinical trial with chronic stroke patients. All patients had a cognitive impairment as measured with the Montreal Cognitive Assessment. In addition, they expressed various degrees of depression expressed by varying scores on the Hamilton Depression Scale. Our results so far suggest that the presence of depression modulates attentional processing similarly to a cognitive load in a psychophysical task. Moreover, we see that depressive patients profited most of the conjunctive cognitive training and that the improvement was most evident in the attention domain. Further analysis will shed light on the underlying mechanisms of this improvement. We hope that this work will aid in not only find better rehabilitation methods, but also improve current diagnostic tools.
References:
[1] M. Lesniak, T. Bak, W. Czepiel, J. Seniów, and A. Czlonkowska, “Frequency and prognostic value of cognitive disorders in stroke patients,” Dement. Geriatr. Cogn. Disord., vol. 26, no. 4, pp. 356–363, 2008.
[2] M. L. Hackett and K. Pickles, “Part I: Frequency of depression after stroke: An updated systematic review and meta-analysis of observational studies,” Int. J. Stroke, vol. 9, pp. 1017–1025, 2014.
[3] L. Mercier, T. Audet, R. Hébert, A. Rochette, and M. F. Dubois, “Impact of motor, cognitive, and perceptual disorders on ability to perform activities of daily living after stroke.,” Stroke., vol. 32, no. 11, pp. 2602–2608, 2001.
[4] R. G. Robinson and R. E. Jorge, “Post-stroke depression: A review,”Am. J. Psychiatry, vol. 173, pp. 221–23, 2016.
[5] R. Gillen, H. Tennen, T. E. McKee, P. Gernert-Dott, and G. Affleck, “Depressive symptoms and history of depression predict rehabilitation efficiency in stroke patients,” Arch. Phys. Med. Rehabil., vol. 82, pp. 1645–1649, 2001.
[6] S. Paolucci, G. Antonucci, E. Gialloreti, M. Traballesi, S. Lubich, L. Pratesi, and L. Palombi, “Predicting Stroke Inpatient Rehabilitation Outcome: The Prominent Role of Neuropsychological Disorders,” Eur. Neurol., vol. 36, no. 6, pp. 385–390, 1996.
[7] M. L. Kauhanen, J. T. Korpelainen, P. Hiltunen, E. Brusin, H. Mononen, R. Maatta, et al., “Poststroke depression correlates with cognitive impairment and neurological deficits,” Stroke, vol. 30, pp. 1875–1880, 1999.

PhD Discussions Sessions: Anna Vila and Alexandre Gomila
2018-10-26 10:00 - 11:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Engineering of 3D Small Intestinal Mucosa Models
Anna Vila, Biomimetic Systems for Cell Engineering
Hydrogels have been used as a scaffold for engineering tissue-like structures due to their biocompatibility and properties similar to the extracellular matrix. There are two main types of hydrogels (i) natural hydrogels, such as gelatin, which are biodegradable and present cell adhesion motifs and (ii) synthetic hydrogels, such as poly(ethylene glycol) diacrylate (PEGDA), which are non-biodegradable and can withstand long-term cell cultures but do not have bioadhesion sequences [El-Sherbiny M.I. et al., 2013]. Both types of hydrogels present complementary benefits.
For this reason, we fabricated a hydrogel co-polymer composed of gelatin methacrylate (GelMA) co-polymerized with PEGDA [Hutson. B.C. et al., 2011]. We employed a single-step, moldless, UV-photolithography-based fabrication technique [Castaño A. et al., submitted] to fabricate scaffolds mimicking the 3D architecture of the small intestinal mucosa. Using this approach, we obtained hydrogel co-polymers with finger-like microstructures, with the roundness and dimensions found in the villi of the native tissue. Mechanical and physicochemical properties such as an approach of the Young’s modulus, degradation rate and swelling ratio of the hydrogels have been characterized. Adding PEGDA to the GelMA hydrogels have provided hydrogels with lower degradation and higher Young’s modulus, which ca be easily tuned by changing the composition of GelMA and PEGDA polymers. Furthermore, we have demonstrated that our scaffolds support the growth and differentiation of intestinal epithelial Caco-2 cells up to 21 days, obtaining a matured epithelial monolayer with effective tissue barrier properties.
Additionally, we increased the complexity of our model of the small intestinal mucosa by incorporating an additional cell compartment to mimic the stroma of the in vivo tissue. To do that, we embedded 3T3-NIH fibroblasts in our scaffolds during the photopolymerization procedure. Then, we cultured Caco-2 cells on top of the 3T3-NIH fibroblast-laden hydrogels up to 21 days. Our preliminary results showed that the co-culture of Caco-2 cells with 3T3-NIH fibroblasts favours the epithelial cell growth and improves their barrier function. Taking all together, we have generated an intestinal mucosa model that allows for the co-culture of different intestinal cell types distributed in compartments, mimicking the spatial-physiological features of the small intestinal mucosa. We believe our model better recapitulates cell-cell crosstalk and cell-matrix interactions found in vivo, being an improved alternative for the cell-base in vitro assays.

In vivo photomodulation of GABA and Glycine receptor channels
Alexandre Gomila, Nanoprobes and Nanoswitches
Neuronal networks are highly complex interactions, which determine even the finest behaviour. The major inhibitory pathways in the central nervous system (CNS) act through chloride ion flux, which are mostly driven by fast-acting ionotropic GABAA and Glycine receptors (GlyR). All of them share structural similarities and belong to pentameric ligand-gated ion channels of the Cys-loop family. Photoswitchable molecules have become a powerful tool in any applied field of bioscience, and are broadly used in biomedical research due to their capacity for enlightening biological aspects from their very basics, such as molecular level, up to entire neuronal networks.
Photopharmacology has proven to be advantageous for spatial and temporal control of biological processes without interfering the system natural dynamics and outcomes. Physiology can be tuned with photomimetic ligands and naturally occurring complex network responses can be segmented into light dependent activities discerning relevant data from a vast matrix of results. As these new tools are broadly used for biomedical purposes and most of them focused towards medical applications, an interpretative analytical platform is needed to screen and identify potential photoswitchable molecules.
Zebrafish (Danio rerio) larvae constitute an excellent animal model for studying and screening photoswitchable molecules in vivo. Zebrafish present a transparent body during larval stages, and therefore are capable of receiving specific and determined light applications. From the 19th hour post fertilisation they acquire behavioural traits, from spontaneous twisting movements to full swimming capacities, which are easily traceable and measurable. The use of up to 96 animals simultaneously allows a parallel high throughput data recovery system to analyse high complexity movements and behaviours, all of it with the use of photopharmacology. Here, we aimed at introducing an effective and reliable methodology for high throughput screening of photoswitchable compounds, including photopharmacological derivatives or peptidoswitches, for any in vivo possible target, from specific neuronal correlated diseases up to possible toxicological outcomes. We focused on the study of the main inhibitory neuronal pathways and their locomotion outcomes on a reliable and comparable animal model.
Hence, several photoswitchable compounds with a common benzodiazepine core and an azobenzene photoswitchable moiety were tested. A first light dependent activity ratio was applied in order to discern the most promising candidates. We identified the UR-DW290 molecule as a light dependent trigger of activity in larvae zebrafish. Larvae zebrafish treated with UR-DW290 maintained higher activity in terms of swimming distance (mm) during the relaxation period and UV-Blue light cycles in comparison to controls. Whole-cell recordings of GABAA and glycine mediated currents by URDW290 showed a potentiating of inhibition when irradiated with ultraviolet light in comparison to visible light illumination.
We propose the combination of high through-put screening and optopharmacology tools for the study and characterisation of zebrafish larvae behaviour focusing on their swimming activity. We identified a first photoswitchable molecule for glycine receptor modulation in vitro and in vivo, UR-DW290, which increases basal activity in zebrafish larvae. This increase is tuneable with UV and Blue light illumniation.

PhD Discussions Sessions: Martí Checa and Javier Rodríguez
2018-11-30 10:00 - 11:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Nanoscale Dielectric Imaging by 3D-Electrostatic Force Microscopy
Martí Checa, Nanoscale bioelectrical characterization
We present Electrostatic Force Volume Microscopy (EFVM) for nanoscale dielectric imaging. EFVM is a new 3D-SPM technique, based in the acquisition of electrostatic force approach curves at each point of a sample and its post-processing and quantification to obtain both Electrostatic Force Microscopy (EFM) images and dielectric constant maps. We show that with a single set of EFVM data one can obtain EFM images in all currently available EFM imaging modes (e.g. constant height, lift mode, constant electric force, etc.) and at any desired tip-sample distance or electric force set point. EFVM enables, in addition, obtaining EFM images under acquisition settings that cannot be implemented in any existing EFM instrument. Finally, EFVM allows obtaining maps of the dielectric constant of the sample with unparalleled accuracy and spatial resolution, irrespectively of the sample topography. We report applications of EFVM to thin oxide films, silver nanowires and single bacterial cells to show the broad applicability of the technique. EFVM is expected to have an important impact in the nanoscale dielectric mapping of topographically complex samples in Materials and Life Sciences.

Sudden cardiac death risk stratification of idiopathic cardiomyopathy patients by the application of cardiovascular coupling analysis
Javier Rodríguez, Biomedical signal processing and interpretation
Cardiovascular diseases are one of the most common cause of death. Early detection of patients at high risk of sudden cardiac death (SCD) is still an issue. The aim of this study was to analyze the cardio-vascular couplings based on heart rate variability (HRV) and blood pressure variability (BPV) analysis in order to introduce new indices that allow noninvasive risk stratification in idiopathic dilated cardiomyopathy patients (IDC).
High-resolution electrocardiogram (ECG) and continuous noninvasive blood pressure (BP) signals were recorded from 91 IDC patients and 49 healthy subjects (CON) for 30 minutes. During a follow-up period of 2 years, 14 patients either died or suffered life-threatening complications due to their cardiac condition. From the ECG and BP signals, the beat-to-beat interval, and systolic and diastolic blood pressure values were extracted. All this new information was analyzed, in univariate and bivariate ways, using the segmented Poincaré plot analysis, the high resolution joint symbolic dynamics and the normalized short time partial directed coherence methods. Indices with statistical significance between different SCD risk levels were selected. Support vector machine (SVM) models were built in order to classify these patients by their level of SCD risk. Patients at high risk of SCD (IDCHR) presented lowered HRV and increased BPV compared to both the low risk patients (IDCLR) and the control subjects, suggesting a depression in their vagal activity and a compensation from the sympathetic activity. The coupling strength from both, the systolic and diastolic blood pressure to the cardiac activity were stronger in high risk patients. Additionally, the cardio-systolic coupling analysis revealed that the systolic influence over the heart rate gets weaker as the risk increases. The SVM IDCLR vs IDCHR model achieved 98.9% accuracy with an area under the curve (AUC) of 0.96. When comparing IDC vs CON groups, 93.6% and 0.94 accuracy and AUC were obtained, respectively. In order to simulate the case were the original status of the subject is unknown, a cascade model was built fusing the aforementioned models, achieving 94.4% accuracy.
In conclusion, this study introduced a novel method of SCD risk stratification of IDC patients based on new indices from coupling analysis and non-linear HRV and BPV. We uncovered some of the complex interactions within the autonomic regulation in this type of patients.

PhD Discussions Sessions: Elena Lantero and Manuel López
2019-03-29 10:00 - 12:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Targeting Plasmodium falciparum with DNA aptamers
Elena Lantero, Nanomalaria
Malaria still remains as one of the main causes of mortality in many developing countries. Caused by parasitic infection of Plasmodium species, the World Health Organization has launched an ambitious plan to eradicate malaria, which will require population mass screening and treatment. In this context, the current diagnostic gold standard is light microscopy of peripheral blood smears, which is time and labor intensive, and needs prepared personnel and continuous training. Alternatively, antigen-based rapid tests have limited sensitivity and do not provide quantitative measure; and PCR-based molecular methods, although sensitive, demand for highly trained personnel and costly reactives. Besides, most current antimalarials have known resistances and new forms of treatment such as targeted delivery are interesting tools to prevent further spreading of such resistances.
Accordingly, malaria massive screening and treatment will require new rapid, sensitive, simple and economically affordable methods, able to detect even asymptomatic infected patients and low-density infections. Screening for new bioreceptors is required in order to increase the sensitivity of current antigen-based malaria rapid diagnosis or to develop new treatments. Antibody production often involves the use of laboratory animals and is time-consuming and costly, especially when the target is Plasmodium, whose variable antigen expression complicates the development of long-lived biomarkers. To circumvent these obstacles we have applied the Systematic Evolution of Ligands by EXponential enrichment (SELEX) method to the rapid identification of DNA aptamers against Plasmodium-infected red blood cells (pRBCs). This type of synthetic bioreceptor is expected to display higher dry-storage and lyophilisation stability than antibodies. Five 70 bp-long ssDNA sequences having a highly specific binding of pRBCs versus non-infected erythrocytes have been identified by using cell-SELEX with fixed pRBCs.
Photons to electrons: Single molecule and time resolved Photosynthetic complex electron transfer study
Manuel López , Nanoprobes and nanoswitches
We present our work on the electron transfer process of plant photosynthetic complex I (PSI). PSI is a membrane protein complex that captures sunlight energy and uses it to shuttle and energize electrons throughout thylakoid membrane, bringing them from a low energy state in one side of the membrane to a very energetic state in the other side.
We study the electron transfer process between photo-oxidized and photo-reduced peripherical cofactors of the protein and their respective protein redox partners performing electrochemical bulk and single molecule photocurrent measurements as protein sample is irradiated with LED and femtosecond pulsed lasers.

PhD Discussions Sessions: Andrés Marco and Davia Prischich
2019-04-26 10:00 - 12:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
An iCRISPR Platform for Human Functional Genetics in Human Pluripotent Stem Cells (hPSCs)
Andrés Marco, Pluripotency for organ regeneration
Self-renewal and pluripotency are the two major functional properties defining human pluripotent stem cells (hPSCs), allowing them to be cultured indefinitely in a dish and maintaining their capacity to differentiate to virtually any human cell lineage, tissue or organ. Hence, they represent an ideal model for studying cellular and multicellular behaviors in both physiological and pathological conditions.
Up until recently, the genome of hPSCs was difficult to manipulate, limiting considerably their use for functional genetics. To solve this problem, here we present the results of our ongoing effort to engineer an iCRISPR platform for highly efficient genome engineering in hPSCs. We have targeted the safe harbor AAVS1 locus using an inducible Cas9 editing vector (iC2), an inducible dCas9 activator (iCa) or an inducible dCas9 repressor (iCr). iCRISPR allows inducible gene knockout, gene upregulation and gene repression.
All together, these lines will greatly expand the repertoire of applications that can be addressed with hPSCs. Our final goal is to use the iCRISPR platform to dissect kidney development and disease in hPSC-derived kidney organoids.

Traffic Lights peptides to photocontrol clathrin-mediated endocytosis in yeasts
Davia Prischich, Nanoprobes and nanoswitches
Clathrin-mediated endocytosis (CME) is crucial to all eukaryotic cells. It is implicated in a variety of cellular processes that range from nutrient uptake, signal transduction and regulation of the membrane components including surface proteins. The functioning of this transient machinery requires a complex network of proteins that cannot be untangled only by means of genetic modification and immunological depletion. In this sense, photopharmacology provides a powerful aid by complementing the selectivity of drugs with the remote and reversible control offered by light.
Traffic Lights (TLs) peptides are cell-permeable, photoswitchable inhibitors specifically developed to target the main adaptor complex of the CME machinery. These peptides, named TL1 and TL2, have already proved capable of inhibiting CME in a light-regulated manner when tested in mammalian cells. Here we show that TL peptides retain their activity in yeast. After having confirmed photoregulation of CME events in this extremely versatile eukaryotic model system, we now aim to achieve in situ activation of the peptide so to directly address the role of endocytosis in cellular processes such as cytokinesis or cell migration.

PhD Discussions Sessions: Ariadna Marin and Ignasi Casanellas
2019-07-26 10:00 - 12:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Linking epithelial size, tension and pressure in curved epithelial monolayers
Ariadna Marin, Integrative Cell and Tissue Dynamics
Epithelia are thin cellular layers that act as mechanical and biochemical barriers. They are dynamic tissues that present strong intercellular junctions needed to maintain their integrity while growing and regenerating. During embryogenesis, they fold progressively and give rise to highly reproducible 3D shapes that guide the shape and positioning of organs.
The way pressure and tension depend on the size of 3D epithelial structures can help us understand how epithelia fold into determined shapes and are able to maintain them even under the continuous remodelling due to cell division. In this project we generate simple fluid-filled MDCK 3D monolayers to study the link between epithelial size, luminal pressure and intercellular tension.
Nanoscale surface adhesiveness continually modulates intercellular communication in cartilage development
Ignasi Casanellas, Nanobioengineering
Nanoscale inputs of the extracellular matrix (ECM) affect cell behavior, including differentiation. We have developed a method for the simple production of large-scale substrates functionalized with cell-adhesive moieties of arginine-glycine-aspartate (RGD) dendrimers, with uneven local densities at the nanoscale.
In the first stages of cartilage formation, mesenchymal stem cells gather together, forming condensates with an extensive gap junctional intercellular communication (GJIC) network. The establishment of this communication network is imperative for the development of healthy cartilage tissue. We have used nanopatterned substrates to locally control cell-substrate adherence during mesenchymal condensation, a prevalent morphogenetic transition, and promote stem cell differentiation towards chondrogenesis. We here demonstrate that local ligand density defines gap junctional protein Cx43 network architecture and GJ functionality.
By a condensate transplantation assay, we then reveal that differentiating stem cells are sensitive to evolving substrate inputs in a continuous feedback mode after condensation. The renewal of optimal ligand conditions led to a revamp of Cx43 expression.
This knowledge provides new insight into cell-matrix nanoscale interactions during morphogenesis. It is relevant for the design of nanopatterned platforms for cell-based regenerative therapies of mesenchymal tissues such as cartilage.

PhD Discussions Sessions: Ferran Velasco and Fabio Riefolo
2019-11-29 10:00 - 12:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Cellulose-based cryogels for long-term culture of pancreatic islets and skeletal muscle tissue
Ferran Velasco, Biosensors for Bioengineering
Islet encapsulation inside traditional hydrogels is one of the most common techniques to study insulin secretion for Diabetes Mellitus studies. However, it’s proved that cells encapsulated in a depth of more than 100 microns die due the lack of nutrient diffusion. As pancreatic islets are spherical aggregations of around 100 microns in diameter, this problem increases exponentially. To solve this problem, in this project we propose the use of new Carboxymethyl cellulose – gelatin biocomposite in combination with cryogelation technique to engineer a new in vitro model to mimic the insulin-mediated skeletal muscle glucose metabolism.
Carboxymethyl cellulose (CMC) is biocompatible, but not mammalian cell-degradable and shows extraordinary elasticity features. Gelatin is able to provide the 3D microenvironment for the proliferation of different cell types and cell-interactive biological activity, very desirable properties for muscle and pancreas tissue scaffold. Cryogelation technique consists in freezing a prepolymer solution at sub-zero temperatures, so water-ice crystals are formed while the material crosslinks. When it’s defrosted, these water-ice crystals lead to “empty” cavities that forms a macroporous and very interconnected scaffold that fits with our needs of morphology and nutrient diffusion.
We first optimize the protocol to achieve the desired morphology; for the pancreatic tissue we achieved a random porosity with high interconnected pores and for the skeletal muscle we fabricate it with an anisotropic structure. We characterize it by stiffness, pore distribution, SEM images and swelling to know its mechanical properties. Then we seed cells in the specific cryogel to characterize its biological behavior depending the cryogel approach used.
Our results are promising for seeding both cell types, as the morphology and pore distributions fits with our needs. These scaffolds show higher nutrient diffusion, good material properties and a better manipulation compared to traditional hydrogels for these tissues.
Photocontrol of Muscarinic Receptors and Applications In Vivo
Fabio Riefolo, Nanoprobes and Nanoswitches
Remote control of physiological functions with light offers the promise of unveiling their complex spatiotemporal dynamics in vivo, and enabling highly focalized therapeutic interventions with reduced systemic toxicity. Optogenetic methods have been implemented in the heart, but the need of genetic manipulation jeopardizes clinical applicability. We present a method to modulate cardiac function with light through a photoswitchable compound and without genetic manipulation. A new light-regulated drug, named PAI, was designed and synthesized to be active on M2 muscarinic acetylcholine receptor (mAChR). PAI can be reversibly photoisomerized between cis and trans conformations under UV and visible light and is able to photocontrol the activation M2 mAChRs in vitro.
We show that PAI has different light-dependent cardiac effects in a mammalian animal model. Finally, we demonstrate the reversible, real-time photocontrol of cardiac function in translucent wildtype tadpoles: PAI induced bradycardia and this effect could be reversibly switched using UV and visible illumination. PAI can also effectively activate M2 receptors using two-photon excitation with near-infrared light, which overcomes the scattering and low penetration of short-wavelength illumination. Such a new approach may enable enhanced spatial and temporal selectivity for cardiovascular drugs.

PhD Discussions Sessions: Maria Arista
2019-12-13 00:30 - 14:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Super-resolution microscopy for understanding the formation and inhibition of influenza virus structures
Maria Arista, Nanoscopy for Nanomedicine
Super-resolution microscopy is a mighty tool that has the ability to study fluorescence samples beyond the diffraction limit, achieving a spatial resolution around 20 nm. The study of viruses can greatly benefit from super-resolution imaging, mainly due to their small size, between 50 and 200 nm. Here we show that, thanks to this technique, we are able to visualize and study two relevant viral structures: filaments of influenza virus using stochastic optical construction microscopy (STORM) and virus-like particles formed from influenza using DNAPAINT (Points accumulation for imaging in nanoscale topography) .
Influenza A virus is highly pleomorphic, and virions can have either spherical or filamentous morphology. Influenza A virus strain A/Udorn/72 (H3N2) produces copious amounts of long and thin filaments on the surface of infected cells, led mainly by the matrix protein M1 and the membrane protein M2. These filaments are strongly related to the infectivity of influenza and cell-to-cell communication, however, due to the small size of these filaments (200 nm of width), they are hard to characterize in detail using immunofluorescence microscopy.
Here, we show with super-resolution microscopy that filament formation was inhibited by the treatment of cells with specific IgG2a and IgG1 antibodies but was not inhibited with the isotype control antibodies. Our results demonstrate that M2e-specific IgGs reduces the level influenza A virus replication in vitro and suggest that the inhibition of virus replication lead by M2especific antibodies is due to the fragmentation of filamentous virions and the loss of filament formation from the surfaces of infected cells.
Moreover, we study virus-like particles produced from influenza proteins transfected on mammalian cells. These structures mimic viruses but they lack viral genetic material, for this reason they are great models to study influenza particles without risks. Influenza expresses 3 different proteins on the surface of the particle and the distribution and homogeneity between particles is not well understood. To study this distribution, we are analyzing with DNA-PAINT the differential expression and distribution of these 3 proteins on the surface of the particles. Overall we show how super-resolution is suitable to study nanoscale viral structures and can provide new insights into anti-viral therapies.

PhD Discussions Sessions: Xarxa Quiroga and Adrián López
2020-02-28 10:00 - 12:00 | IBEC, floor 11, Tower I, Baldiri Reixac 4-8, 08028 Barcelona, Spain
Biochemical responses to cell membrane mechanical remodelling
Xarxa Quiroga, Cellular and molecular mechanobiology
In a range of physiological processes, from extravasation to endocytosis, cells are constantly submitted to morphological changes, which eventually entail plasma membrane reshaping and adaptation. This remodelling could be harnessed by cells to detect and respond to shape changes, enabling mechanosensing mechanisms. However, how this occurs is still largely unknown.
To increase our understanding on how such process can happen, we have engineered a cell-stretching system that allows us to induce controlled plasma membrane remodelling while monitoring the whole process with the help of a microscope.
By using this set up, we have found that cell de-stretch triggers the formation of transient membrane evaginations whose resorption is actively regulated by BAR protein recruitment and actin polymerisation. The described process may be the first part of a molecular cascade used by cells in response to stretch.

Development of Microphysiological Systems for the Evaluation of Regenerative Therapies
Adrián López, Biomaterials for Regenerative Therapies
The modelling of human organs has long been a task for scientist in order to lower the costs of therapeutic development and understand the pathological onset of human disease. Animal models remain the gold standard for drug discovery, despite their widely recognized limitations such as their marked differences with humans in terms of genetics and etiology or their high cost.
During the last decade, the advancements in tissue engineering and microfabrication gave rise to innovative models known as organs-on-a-chip or microphysiological systems, which aim to build functional miniaturized tissues in vitro that closely mimic the actual in vivo microenvironment. In this work, we will present two microphysiological platforms that we are developing with the goal of understanding and evaluating biomaterial-based regenerative therapies. The first model is aimed at replicating the bone healing microenvironment to evaluate the angiogenic potential of calcium-releasing scaffolds. The second model will be focused on the generation of an ischemic injury on a physiologically relevant cardiac tissue to test if lactate-releasing scaffolds are able to stimulate cardiac tissue regeneration.

PhD Discussions Sessions: Nimesh Ramesh and Sock Ching Low
2020-12-11 10:00 - 12:00
Microfluidic device for engineering 3D epithelial monolayers with controlled pressure
Nimesh Ramesh, Integrative Cell and Tissue Dynamics
The remarkable feature of the epithelial sheets is to form specialized 3D structures suited to their physiological roles, such as highly branched structures in the lungs, drastic shape changes during embryonic development, or self-organizing organoids. These tissues are distinctive not just in the forms cells assume, but also in function. To achieve this, tissues and the cells in them exhibit coordinated behavior across the spatial and temporal scale. In a sense, 3D epithelia resemble an active material that adapts and changes in response to its biophysical-chemical stimuli like gene expression, morphogen gradients, and lumen pressure. A rheological study of the epithelia would provide unique insight on two fronts. First, to understand the fundamental physical rules of the biology, and second for inspiration of new engineering tools and design principles.
Our study focuses on the tissue response to physical forces, specifically pressure, tension, and curvature. We have fabricated a microfluidic setup to subject epithelial tissues to lumen pressure at different spatial and temporal scales. The epithelial monolayer is grown on a porous surface with circular low adhesion zones. On applying controlled pressure, the monolayer delaminates into a spherical cap (dome). Laplace law for spherical shells allows us to compute tension in the 3D structure with applied pressure and the radius of the dome.
This microfluidic device helps us to characterize the 3D epithelial shape along with the mapping of physical forces. Here, we demonstrate that the device can subject MDCK epithelial cells to a range of lumen pressure at different rates. Drastic reduction in pressure results in tissue collapsing into wrinkles; showing buckling tendency of the tissue under compression. We think that our device enables studying geometrical and biophysical constraints of tissues and unravel emergent phenomena in tissues.

Saccade rate is associated with number of items in working memory
Sock Shing Low, Synthetic, Perceptive, Emotive and Cognitive Systems (SPECS)
Working memory has been shown to rely on theta oscillations for item representations, and the successful recall of items depends greatly on theta’s phase during both encoding and recall. At the same time, it has been observed that saccadic eye movements during visual exploration trigger theta phase-resets, raising the question of whether the neuronal substrates of mnemonic processing rely on motor-evoked responses. To quantify the relationship between saccadic eye movements and working memory load, we tested human participants performing an n-back Sternberg auditory task in combination with a colour-based catch detection task. We observed a task-specific interference in performance and an increase in saccade rate when both tasks were carried out simultaneously. Saccade rate also increased concurrently with working memory load in the Sternberg task’s pre-response stage, reflecting its hypothesised role in memory recall. Our results suggest an interplay between saccades and hippocampal theta during retrieval of items in working memory.
The PhD discussions session will be held ONLINE at the GoToMeeting platform

PhD Discussions Sessions: Gerard Rubí and Ignasi Ferrer
2021-01-29 10:00 - 12:00
Development of an in vitro three-dimensional colorectal tumor model for drug screening
Gerard Rubí, Biomaterials for Regenerative Therapies
The majority of morphogenetic and pathological processes are driven by cells responding to the surrounding matrix cues, including matrix composition, architecture, and mechanical properties. Despite the increased evidence of extracellular matrix (ECM) properties, in vitro substitutes still fail to effectively mimic the native microenvironment. In this study, we aim to develop and characterize cell-derived extracellular matrices (CDMs) obtained through a protein deposition from human mesenchymal stem cells cultured in sacrificial 3D scaffold templates of poly-lactic acid (PLA) microcarriers. Obtained decellularized CDMs closely mimic biochemical, physical, and mechanical properties of native tissues’ ECM. The produced novel CDMs, are currently tested as a 3D cell culture platform for disease modelling. This is achieved through CDMs repopulation with colorectal cancer cells and cancer associated fibroblasts (CAFs). The new 3D CDMs-cancer platform will provide an in vitro tumor model to study the cells-ECM interactions and potential therapeutic targets, to finally serve as a drug-screening platform for personalized medicine.
Novel m-Health and multimodal physiological biomarkers for non-invasive monitoring and home healthcare of Obstructive Sleep Apnea and COPD patients with comorbidities
Ignasi Ferrer, Biomedical Signal Processing and Interpretation
Obstructive sleep apnea (OSA) is a sleep disorder in which repetitive upper airway obstructive events occur during sleep. These events can induce hypoxia, which is a risk factor for multiple cardiovascular and cerebrovascular diseases. OSA is also known to be position-dependent in some patients, which is referred to as positional OSA (pOSA). The gold-standard technique for diagnosing OSA is nocturnal polysomnography (PSG), which consists in recording multiple physiological signals while the patient is asleep in a hospital sleep lab. However, PSG has some important limitations, such as the high cost of the diagnostic test; the diagnosis is usually performed with a one-night sleep assessment, which does not account for the variability of sleep performance in the patient; and the sleep quality varies from that at home, because the patient has to sleep in a different bed connected to a lot of electrodes and wires.
In this study we aim to study how smartphones could be used to diagnose and monitor sleep apnea at home. Since smartphones are worldwide available devices, with a lot of embedded sensors, they appear as a feasible mHealth tools that could help overcome these limitations.
The PhD discussions session will be held ONLINE at the GoToMeeting platform

PhD Discussions Sessions: Héctor López Carral
2021-03-26 10:00 - 12:00
Subjective ratings of emotive stimuli predict the impact of the COVID-19 quarantine on affective states
Héctor López Carral, SPECS lab, IBEC
The COVID-19 crisis resulted in a large proportion of the world’s population having to employ social distancing measures and self-quarantine. Given that limiting social interaction impacts mental health, we assessed the effects of quarantine on emotive perception as a proxy of affective states. To this end, we conducted an online experiment whereby 112 participants provided affective ratings for a set of normative images and reported on their well-being during COVID-19 self-isolation. We found that current valence ratings were significantly lower than the original ones from 2015. This negative shift correlated with key aspects of the personal situation during the confinement, including working and living status, and subjective well-being. These findings indicate that quarantine impacts mood negatively, resulting in a negatively biased perception of emotive stimuli. Moreover, our online assessment method shows its validity for large-scale population studies on the impact of COVID-19 related mitigation methods and well-being.
The PhD discussions session will be held ONLINE at the GoToMeeting platform

PhD Discussion Sessions: Teodora Andrian and Xavier Arqué
2021-04-30 10:00 - 12:00
Correlating super-resolution microscopy and transmission electron microscopy for nanoparticle characterisation
Teodora Andrian, Nanoscopy for nanomedicine group
The functionalization of nanoparticles with surface functional moieties is a key strategy to achieve bioactivity and cell targeting in nanomedicine. The interplay between size and ligand number and distribution is crucial for the formulation performance and needs to be properly characterized to understand nanoparticle structure-activity relations. However, the particle-to-particle heterogeneity poses a serious challenge due to the lack of methods able to measure both size and ligand number and distribution at the same time and at the single particle level. Here we address this issue by introducing a correlative method combining super-resolution microscopy (SRM) and transmission electron microscopy (TEM) imaging. Correlative light and electron microscopy (CLEM) techniques proved their potential in structural biology but to the best of our knowledge, they have not yet been explored for the structural characterization of nanoparticles. Here we apply our super-resCLEM method to characterize the relationship between size and ligand number, and ligand density in PLGA-PEG nanoparticles at the single particle and single-molecule level. We highlight how heterogeneity found in nanoparticle size can impact ligand distribution, and we discuss the implications on formulation performance. We show how a significant part of the nanoparticle population goes completely undetected in the single-technique analysis, demonstrating that the characterization of nanomaterials using a multiparametric correlative method outplays the information obtained compared to a one-method-at-a-time approach. Using SRM alone, we demonstrated how PEG architecture can influence ligand conjugation efficiency and accessibility. The applicability of our method spans beyond PLGA-PEG nanoparticles and holds great promise for the multiparametric analysis of several other parameters and nanomaterials.
Unraveling the fundamental aspects of enzyme-powered micromotors
Xavier Arqué, Smart Nano-Bio-Devices group
Enzyme-coated micro- and nanomotors self-propel by the biocatalytic conversion of substrates into products and show great promise as actively navigating agents in the fields of biomedicine and environmental applications. However, many of the fundamental aspects underlying enzyme-powered self-propulsion have yet to be fully understood and are crucial for their optimal implementation. Under this framework, this research is focused on elucidating and studying the intrinsic (catalytic turnover or structural flexibility) and extrinsic (bulk and local ionic media) enzymatic properties that lead to an improved active motion powered by bio-catalysis. This is enabled by exploring novel types of both i) enzymes that can act as active motion engines and ii) platforms with appealing properties to be used as chassis of enzymatic micro- and nanomotors. Overall, this work contributes to a better understanding of the mechanism of motion of enzymatic active motion, expands the current library of enzymatic engines and chassis materials available, and provides new insights into the feasibility of implementation of enzyme-powered micro- and nanomotors.
The session will be held online using the GoToMeeting Platform

PhD Discussions: Ignasi Granero
2021-07-02 10:00 - 12:00
Ignasi Granero, Celular and molecular mechanobiology group
In eukaryotic cells, the cell nucleus contains the DNA and directs the reaction to important signalling cascades, both biochemical and biomechanical. At the Cellular and Molecular Mechanobiology lab we study how forces and the physical characteristics of the cell environment affect cell behaviour and homeostasis. In my case, I am interested in how the cell nucleus senses physical changes and triggers a cell response to them.
For this purpose, we have engineered a battery of fluorescent proteins, which change their location depending on the physical state of the nucleus. Higher forces in the nucleus correspond with higher nuclear accumulation of these proteins. Indicating that these proteins react to mechanical/morphological stimuli. Then, we have chosen the variant that can be used as a mechano-sensor. Currently, we are testing the sensor in single- and multi-cell approaches to understand its behaviour in different systems and the ruling principles under its functioning.
This PhD Discussion will be held online using the GoToMeeting app

PhD Discussions: Yolanda Castillo and Marc Molina
2021-11-26 10:00 - 12:00
Assessment of trunk function in patients with spinal cord injury using electromyography and smartphone accelerometry
Yolanda Castillo, Biomedical Signal Processing and Interpretation
Spinal cord injury (SCI) causes motor and sensory impairment below the level of the injury, but also many other health problems. A common consequence of SCI is the lack of control over trunk muscles, leading to deficits in postural control and balance while sitting. Since trunk stability is essential to maintain upright posture and support functional movements, impaired trunk function constitutes a major cause of motor disability in SCI patients, limiting their independence and quality of life. However, trunk stability is rarely examined in studies of mobility after SCI, and one of the reasons is the lack of quantitative measures for assessing trunk function. Here we propose to record and analyze electromyographic (EMG) and smartphone accelerometric data to extract quantitative measures for the evaluation of trunk function. Our aims were: 1) to characterize muscle activity and movement patterns of trunk flexion during a reaching task in healthy subjects and patients with SCI, 2) to compare the impact of cervical and thoracic injuries in trunk function, and 3) to investigate the potentially destabilizing effects of a startling acoustic stimulus in this task. For these purposes, during a reaching movement requiring trunk flexion, we recorded the EMG activity of 8 trunk, neck, and shoulder muscles and smartphone accelerometer data from individuals with cervical SCI, thoracic SCI, and healthy control subjects. We analyzed these signals and extracted different features, including the response time until pressing a target button, EMG onset latencies and amplitudes, and trunk tilt, lateral deviation, and other movement features from accelerometry. The proposed outcome measures revealed deficits in postural control and compensatory strategies employed by SCI patients, including delayed responses and higher lateral deviations, which might have important consequences for rehabilitation. The combination of EMG and smartphone accelerometer data can help to develop more suitable methods for the assessment of trunk function in individuals with SCI, thus improving the follow-up and management of these patients.
Development of a model of “macro” substrates for the analysis of 3D chromatin structure and transcriptional profiling
Marc Molina, Cellular and Molecular Mechanobiology
Mechanically-induced changes in the genome are increasingly recognized as major drivers of cell and tissue function. However, current and past studies on this topic in vitro have often been limited by the sample size required for these genomic analyses. Here we describe the development of a polyacrylamide gel (pAAg) substrate with larger area than gels previously generated in labs worldwide. These substrates display the same tunable stiffness as their smaller counterparts and are particularly suitable to accommodate large numbers of cells. We tested the substrates to assess the effect that changes in rigidity have in the cell’s genome, both at the level of chromatin organization and transcriptional regulation. For this, two different rigidities were used (soft vs stiff) with three conditions which included: i) a control group, ii) a transiently expressing mutant of RanQ69L that prevents all nucleocytoplasmic transport and iii) a transiently expressing mutant of NES1-KASH that prevents force transmission to the nucleus. Our preliminary results suggest that cells seeded on pAAg substrates of different rigidities display differences in chromatin structure and gene expression, but more data will be necessary to further support these results. Ultimately, the aim of our project is to understand down the road how changes in rigidity affect: i) the 3D structure and interaction map of chromatin and ii) the activated or repressed transcriptional programs in soft and stiff substrates. Overall, this new model will help us to characterize how the genome is affected by rigidity both at the structural and transcriptional levels. More broadly, we expect the potential findings of this work to help the community detailing the effect of changes in tissue stiffness in the genome spatial organization.
This PhD Discussion will be hybrid. Yolanda Castillo will be doing her presentation online and Marc from the Baobab room, located in tower I floor 11. To follow the session online, find here the link, we will be using the GoToMeeting app. If you wish to attend in person, the free spots will be assigned on a first come first served basis, the capacity of the room is for 30 people.
