In Vitro Models for Studying Chronic Drug-Induced Liver Injury

[EN] Drug-induced liver injury (DILI) is a major clinical problem in terms of patient morbidity and mortality, cost to healthcare systems and failure of the development of new drugs. The need for consistent safety strategies capable of identifying a potential toxicity risk early in the drug discovery pipeline is key. Human DILI is poorly predicted in animals, probably due to the well-known interspecies differences in drug metabolism, pharmacokinetics, and toxicity targets. For this reason, distinct cellular models from primary human hepatocytes or hepatoma cell lines cultured as 2D monolayers to emerging 3D culture systems or the use of multi-cellular systems have been proposed for hepatotoxicity studies. In order to mimic long-term hepatotoxicity in vitro, cell models, which maintain hepatic phenotype for a suitably long period, should be used. On the other hand, repeated-dose administration is a more relevant scenario for therapeutics, providing information not only about toxicity, but also about cumulative effects and/or delayed responses. In this review, we evaluate the existing cell models for DILI prediction focusing on chronic hepatotoxicity, highlighting how better characterization and mechanistic studies could lead to advance DILI prediction.This work has been supported by the Institute of Health Carlos III (ISCIII, Plan Estatal de I+D+i 2013-2016) and co-financed by the European Regional Development Fund "A way to achieve Europe" (FEDER) through grant PI21/00223, by the Spanish Ministry of Science and Innovation Ministry-Spanish Research Agency through the Project PID2019-106000RB-C22/AEI/10.13039/501100011033, and by the Generalitat Valenciana (PROMETEO/2019/060).Donato, MT.; Gallego-Ferrer, G.; Tolosa, L. (2022). In Vitro Models for Studying Chronic Drug-Induced Liver Injury. International Journal of Molecular Sciences. 23(19):1-30. https://doi.org/10.3390/ijms231911428130231

In Vitro Modeling of Non-Solid Tumors: How Far Can Tissue Engineering Go?

[EN] In hematological malignancies, leukemias or myelomas, malignant cells present bone marrow (BM) homing, in which the niche contributes to tumor development and drug resistance. BM architecture, cellular and molecular composition and interactions define differential microenvironments that govern cell fate under physiological and pathological conditions and serve as a reference for the native biological landscape to be replicated in engineered platforms attempting to reproduce blood cancer behavior. This review summarizes the different models used to efficiently reproduce certain aspects of BM in vitro; however, they still lack the complexity of this tissue, which is relevant for fundamental aspects such as drug resistance development in multiple myeloma. Extracellular matrix composition, material topography, vascularization, cellular composition or stemness vs. differentiation balance are discussed as variables that could be rationally defined in tissue engineering approaches for achieving more relevant in vitro models. Fully humanized platforms closely resembling natural interactions still remain challenging and the question of to what extent accurate tissue complexity reproduction is essential to reliably predict drug responses is controversial. However, the contributions of these approaches to the fundamental knowledge of non-solid tumor biology, its regulation by niches, and the advance of personalized medicine are unquestionable.PROMETEO/2016/063 project is acknowledged. The CIBER-BBN initiative is funded by the VI National R&D&I Plan 2008-2011, Iniciativa Ingenio 2010, Consolider Program. CIBER actions are financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund. This work was also supported by the Spanish Ministry of Science, Innovation and Universities through Grant FPU17/05810 awarded to Sandra Clara-Trujillo.Clara-Trujillo, S.; Gallego Ferrer, G.; Gómez Ribelles, JL. (2020). In Vitro Modeling of Non-Solid Tumors: How Far Can Tissue Engineering Go?. International Journal of Molecular Sciences. 21(16):1-31. https://doi.org/10.3390/ijms21165747S1312116Langer, R., & Vacanti, J. P. (1993). Tissue Engineering. Science, 260(5110), 920-926. doi:10.1126/science.8493529Kelm, J. M., Lal-Nag, M., Sittampalam, G. S., & Ferrer, M. (2019). Translational in vitro research: integrating 3D drug discovery and development processes into the drug development pipeline. Drug Discovery Today, 24(1), 26-30. doi:10.1016/j.drudis.2018.07.007Pradhan, S., Hassani, I., Clary, J. M., & Lipke, E. A. (2016). Polymeric Biomaterials for In Vitro Cancer Tissue Engineering and Drug Testing Applications. 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PCL-Coated Multi-Substituted Calcium Phosphate Bone Scaffolds with Enhanced Properties

[EN] Ionic substitutions within the hydroxyapatite lattice are a widely used approach to mimic the chemical composition of the bone mineral. In this work, Sr-substituted and Mg- and Sr-co-substituted calcium phosphate (CaP) scaffolds, with various levels of strontium and magnesium substitution, were prepared using the hydrothermal method at 200 degrees C. Calcium carbonate skeletons of cuttlefish bone, ammonium dihydrogenphosphate (NH4H2PO4), strontium nitrate (Sr(NO3)(2)), and magnesium perchlorate (Mg(ClO4)(2)) were used as reagents. Materials were characterized by means of X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). Whole powder pattern decomposition refinements of XRD data indicated that increased magnesium content in the Mg- and Sr-co-substituted scaffolds was related to an increased proportion of the whitlockite (WH) phase in the biphasic hydroxyapatite (HAp)/WH scaffolds. In addition, refinements indicate that Sr2+ ions have replaced Ca2+ sites in the WH phase. Furthermore, PCL-coated Mg-substituted and Sr- and Mg-co-substituted scaffolds, with the HAp:WH wt. ratio of 90:10 were prepared by vacuum impregnation. Results of compression tests showed a positive impact of the WH phase and PCL coating on the mechanical properties of scaffolds. Human mesenchymal stem cells (hMSCs) were cultured on composite scaffolds in an osteogenic medium for 21 days. Immunohistochemical staining showed that Mg-Sr-CaP/PCL scaffold exhibited higher expression of collagen type I than the Mg-CaP/PCL scaffold, indicating the positive effect of Sr2+ ions on the differentiation of hMSCs, in concordance with histology results. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis confirmed an early stage of osteogenic differentiation.This research was funded by the Croatian Science Foundation (project IP-2014-09-3752) and the European Structural and Investments Funds (grant KK.01.1.1.07.0014.). The authors thank Inga Urli, Faculty of Science, University of Zagreb for providing Hek293 and hMSC cells. Compression experiments were carried out at the Centre for Biomaterials and Tissue Engineering (CBIT), Universitat Politecnica de Valencia, Valencia, Spain under the PID2019-106000RB-C21/AEI/10.13039/501100011033 project. The authors would like to thank Jorge Mas-Estelles for his generous assistance.Bauer, L.; Antunovic, M.; Gallego-Ferrer, G.; Ivankovic, M.; Ivankovic, H. (2021). PCL-Coated Multi-Substituted Calcium Phosphate Bone Scaffolds with Enhanced Properties. Materials. 14(16):1-19. https://doi.org/10.3390/ma14164403S119141

Synthesis and Characterization of Oxidized Polysaccharides for In Situ Forming Hydrogels

[EN] Polysaccharides are widely used as building blocks of scaffolds and hydrogels in tissue engineering, which may require their chemical modification to permit crosslinking. The goal of this study was to generate a library of oxidized alginate (oALG) and oxidized hyaluronic acid (oHA) that can be used for in situ gelling hydrogels by covalent reaction between aldehyde groups of the oxidized polysaccharides (oPS) and amino groups of carboxymethyl chitosan (CMC) through imine bond formation. Here, we studied the effect of sodium periodate concentration and reaction time on aldehyde content, molecular weight of derivatives and cytotoxicity of oPS towards 3T3-L1 fibroblasts. It was found that the molecular weights of all oPs decreased with oxidation and that the degree of oxidation was generally higher in oHA than in oALG. Studies showed that only oPs with an oxidation degree above 25% were cytotoxic. Initial studies were also done on the crosslinking of oPs with CMC showing with rheometry that rather soft gels were formed from higher oxidized oPs possessing a moderate cytotoxicity. The results of this study indicate the potential of oALG and oHA for use as in situ gelling hydrogels or inks in bioprinting for application in tissue engineering and controlled release.This work was supported by the Deutscher Akademischer Austauschdienst DAAD (grant No. 91605199 to MM) and Deutsche Forschungsgemeinschaft (grant Gr1290/11-1 to TG). The kind support by Spanish State Research Agency (AEI) through the PID2019-106000RB-C21/AEI/10.13039/501100011033 project (including the FEDER financial support) to GGF is acknowledged. We acknowledge the financial support within the funding programme "Open Access Publishing" by the German Research Foundation (DFG).We are very thankful to Andrea Liedmann for her guidance during the cell experiments and Alexandros Repanas for his help during the synthesis and characterization of oPs and data analyses. Furthermore, Marie-Luise Trutschel is acknowledged for her guidance during the rheological measurements.Muhammad, M.; Willems, C.; Rodríguez-Fernández, J.; Gallego Ferrer, G.; Groth, T. (2020). Synthesis and Characterization of Oxidized Polysaccharides for In Situ Forming Hydrogels. Biomolecules. 10(8):1-18. https://doi.org/10.3390/biom10081185S118108Ratner, B. D. (2019). Biomaterials: Been There, Done That, and Evolving into the Future. Annual Review of Biomedical Engineering, 21(1), 171-191. doi:10.1146/annurev-bioeng-062117-120940Morais, J. M., Papadimitrakopoulos, F., & Burgess, D. J. (2010). Biomaterials/Tissue Interactions: Possible Solutions to Overcome Foreign Body Response. The AAPS Journal, 12(2), 188-196. doi:10.1208/s12248-010-9175-3Domingues, R. M. A., Silva, M., Gershovich, P., Betta, S., Babo, P., Caridade, S. G., … Gomes, M. E. (2015). Development of Injectable Hyaluronic Acid/Cellulose Nanocrystals Bionanocomposite Hydrogels for Tissue Engineering Applications. Bioconjugate Chemistry, 26(8), 1571-1581. doi:10.1021/acs.bioconjchem.5b00209Pop-Georgievski, O., Zimmermann, R., Kotelnikov, I., Proks, V., Romeis, D., Kučka, J., … Werner, C. (2018). Impact of Bioactive Peptide Motifs on Molecular Structure, Charging, and Nonfouling Properties of Poly(ethylene oxide) Brushes. Langmuir, 34(21), 6010-6020. doi:10.1021/acs.langmuir.8b00441Wen, Q., Mithieux, S. M., & Weiss, A. S. (2020). Elastin Biomaterials in Dermal Repair. Trends in Biotechnology, 38(3), 280-291. doi:10.1016/j.tibtech.2019.08.005Trujillo, S., Gonzalez-Garcia, C., Rico, P., Reid, A., Windmill, J., Dalby, M. J., & Salmeron-Sanchez, M. (2020). Engineered 3D hydrogels with full-length fibronectin that sequester and present growth factors. Biomaterials, 252, 120104. doi:10.1016/j.biomaterials.2020.120104Xu, M., Pradhan, S., Agostinacchio, F., Pal, R. K., Greco, G., Mazzolai, B., … Yadavalli, V. K. (2019). Easy, Scalable, Robust, Micropatterned Silk Fibroin Cell Substrates. Advanced Materials Interfaces, 6(8), 1801822. doi:10.1002/admi.201801822Köwitsch, A., Zhou, G., & Groth, T. (2017). Medical application of glycosaminoglycans: a review. Journal of Tissue Engineering and Regenerative Medicine, 12(1), e23-e41. doi:10.1002/term.2398Yang, Y., Lu, Y., Zeng, K., Heinze, T., Groth, T., & Zhang, K. (2020). Recent Progress on Cellulose‐Based Ionic Compounds for Biomaterials. Advanced Materials, 33(28), 2000717. doi:10.1002/adma.202000717Yu, Y., Shen, M., Song, Q., & Xie, J. (2018). Biological activities and pharmaceutical applications of polysaccharide from natural resources: A review. Carbohydrate Polymers, 183, 91-101. doi:10.1016/j.carbpol.2017.12.009Grasdalen, H. (1983). High-field, 1H-n.m.r. spectroscopy of alginate: sequential structure and linkage conformations. Carbohydrate Research, 118, 255-260. doi:10.1016/0008-6215(83)88053-7Criado-Gonzalez, M., Fernandez-Gutierrez, M., San Roman, J., Mijangos, C., & Hernández, R. (2019). Local and controlled release of tamoxifen from multi (layer-by-layer) alginate/chitosan complex systems. Carbohydrate Polymers, 206, 428-434. doi:10.1016/j.carbpol.2018.11.007Kirdponpattara, S., Khamkeaw, A., Sanchavanakit, N., Pavasant, P., & Phisalaphong, M. (2015). Structural modification and characterization of bacterial cellulose–alginate composite scaffolds for tissue engineering. Carbohydrate Polymers, 132, 146-155. doi:10.1016/j.carbpol.2015.06.059Price, R. D., Berry, M. G., & Navsaria, H. A. (2007). Hyaluronic acid: the scientific and clinical evidence. Journal of Plastic, Reconstructive & Aesthetic Surgery, 60(10), 1110-1119. doi:10.1016/j.bjps.2007.03.005Kristiansen, K. A., Potthast, A., & Christensen, B. E. (2010). Periodate oxidation of polysaccharides for modification of chemical and physical properties. Carbohydrate Research, 345(10), 1264-1271. doi:10.1016/j.carres.2010.02.011Millan, C., Cavalli, E., Groth, T., Maniura-Weber, K., & Zenobi-Wong, M. (2015). Engineered Microtissues Formed by Schiff Base Crosslinking Restore the Chondrogenic Potential of Aged Mesenchymal Stem Cells. Advanced Healthcare Materials, 4(9), 1348-1358. doi:10.1002/adhm.201500102Reyes, J. M. G., Herretes, S., Pirouzmanesh, A., Wang, D.-A., Elisseeff, J. H., Jun, A., … Behrens, A. (2005). A Modified Chondroitin Sulfate Aldehyde Adhesive for Sealing Corneal Incisions. Investigative Opthalmology & Visual Science, 46(4), 1247. doi:10.1167/iovs.04-1192Peppas, N. A., Hilt, J. Z., Khademhosseini, A., & Langer, R. (2006). Hydrogels in Biology and Medicine: From Molecular Principles to Bionanotechnology. Advanced Materials, 18(11), 1345-1360. doi:10.1002/adma.200501612Van Tomme, S. R., Storm, G., & Hennink, W. E. (2008). In situ gelling hydrogels for pharmaceutical and biomedical applications. International Journal of Pharmaceutics, 355(1-2), 1-18. doi:10.1016/j.ijpharm.2008.01.057Mota, C., Camarero-Espinosa, S., Baker, M. B., Wieringa, P., & Moroni, L. (2020). Bioprinting: From Tissue and Organ Development to in Vitro Models. Chemical Reviews, 120(19), 10547-10607. doi:10.1021/acs.chemrev.9b00789Matyash, M., Despang, F., Ikonomidou, C., & Gelinsky, M. (2014). Swelling and Mechanical Properties of Alginate Hydrogels with Respect to Promotion of Neural Growth. Tissue Engineering Part C: Methods, 20(5), 401-411. doi:10.1089/ten.tec.2013.0252Berger, J., Reist, M., Mayer, J. M., Felt, O., Peppas, N. A., & Gurny, R. (2004). Structure and interactions in covalently and ionically crosslinked chitosan hydrogels for biomedical applications. European Journal of Pharmaceutics and Biopharmaceutics, 57(1), 19-34. doi:10.1016/s0939-6411(03)00161-9Segura, T., Anderson, B. C., Chung, P. H., Webber, R. E., Shull, K. R., & Shea, L. D. (2005). Crosslinked hyaluronic acid hydrogels: a strategy to functionalize and pattern. Biomaterials, 26(4), 359-371. doi:10.1016/j.biomaterials.2004.02.067De la Riva, B., Nowak, C., Sánchez, E., Hernández, A., Schulz-Siegmund, M., Pec, M. K., … Évora, C. (2009). VEGF-controlled release within a bone defect from alginate/chitosan/PLA-H scaffolds. European Journal of Pharmaceutics and Biopharmaceutics, 73(1), 50-58. doi:10.1016/j.ejpb.2009.04.014Yang, Y., Köwitsch, A., Ma, N., Mäder, K., Pashkuleva, I., Reis, R. L., & Groth, T. (2015). Functionality of surface-coupled oxidised glycosaminoglycans towards fibroblast adhesion. Journal of Bioactive and Compatible Polymers, 31(2), 191-207. doi:10.1177/0883911515599999Köwitsch, A., Yang, Y., Ma, N., Kuntsche, J., Mäder, K., & Groth, T. (2011). Bioactivity of immobilized hyaluronic acid derivatives regarding protein adsorption and cell adhesion. Biotechnology and Applied Biochemistry, 58(5), 376-389. doi:10.1002/bab.41Korzhikov, V., Roeker, S., Vlakh, E., Kasper, C., & Tennikova, T. (2008). Synthesis of Multifunctional Polyvinylsaccharide Containing Controllable Amounts of Biospecific Ligands. Bioconjugate Chemistry, 19(3), 617-625. doi:10.1021/bc700383wZhao, M., Li, L., Zhou, C., Heyroth, F., Fuhrmann, B., Maeder, K., & Groth, T. (2014). Improved Stability and Cell Response by Intrinsic Cross-Linking of Multilayers from Collagen I and Oxidized Glycosaminoglycans. Biomacromolecules, 15(11), 4272-4280. doi:10.1021/bm501286fTang, Q.-Q., Otto, T. C., & Lane, M. D. (2004). Commitment of C3H10T1/2 pluripotent stem cells to the adipocyte lineage. Proceedings of the National Academy of Sciences, 101(26), 9607-9611. doi:10.1073/pnas.0403100101Alarake, N. Z., Frohberg, P., Groth, T., & Pietzsch, M. (2017). Mechanical Properties and Biocompatibility of in Situ Enzymatically Cross-Linked Gelatin Hydrogels. The International Journal of Artificial Organs, 40(4), 159-168. doi:10.5301/ijao.5000553Morra, M. (2005). Engineering of Biomaterials Surfaces by Hyaluronan. Biomacromolecules, 6(3), 1205-1223. doi:10.1021/bm049346iZhang, R., Xue, M., Yang, J., & Tan, T. (2011). A novel injectable and in situ crosslinked hydrogel based on hyaluronic acid and α,β-polyaspartylhydrazide. Journal of Applied Polymer Science, 125(2), 1116-1126. doi:10.1002/app.34828Jejurikar, A., Seow, X. T., Lawrie, G., Martin, D., Jayakrishnan, A., & Grøndahl, L. (2012). Degradable alginate hydrogels crosslinked by the macromolecular crosslinker alginate dialdehyde. Journal of Materials Chemistry, 22(19), 9751. doi:10.1039/c2jm30564jEmami, Z., Ehsani, M., Zandi, M., & Foudazi, R. (2018). Controlling alginate oxidation conditions for making alginate-gelatin hydrogels. Carbohydrate Polymers, 198, 509-517. doi:10.1016/j.carbpol.2018.06.080Yegappan, R., Selvaprithiviraj, V., Mohandas, A., & Jayakumar, R. (2019). Nano polydopamine crosslinked thiol-functionalized hyaluronic acid hydrogel for angiogenic drug delivery. Colloids and Surfaces B: Biointerfaces, 177, 41-49. doi:10.1016/j.colsurfb.2019.01.035Bouhadir, K. H., Lee, K. Y., Alsberg, E., Damm, K. L., Anderson, K. W., & Mooney, D. J. (2001). Degradation of Partially Oxidized Alginate and Its Potential Application for Tissue Engineering. Biotechnology Progress, 17(5), 945-950. doi:10.1021/bp010070pSträtz, J., Liedmann, A., Heinze, T., Fischer, S., & Groth, T. (2019). Effect of Sulfation Route and Subsequent Oxidation on Derivatization Degree and Biocompatibility of Cellulose Sulfates. Macromolecular Bioscience, 20(2), 1900403. doi:10.1002/mabi.201900403Elahipanah, S., O’Brien, P. J., Rogozhnikov, D., & Yousaf, M. N. (2017). General Dialdehyde Click Chemistry for Amine Bioconjugation. Bioconjugate Chemistry, 28(5), 1422-1433. doi:10.1021/acs.bioconjchem.7b00106Huang, G., & Huang, H. (2018). Application of hyaluronic acid as carriers in drug delivery. Drug Delivery, 25(1), 766-772. doi:10.1080/10717544.2018.1450910Qhattal, H. S. S., & Liu, X. (2011). Characterization of CD44-Mediated Cancer Cell Uptake and Intracellular Distribution of Hyaluronan-Grafted Liposomes. Molecular Pharmaceutics, 8(4), 1233-1246. doi:10.1021/mp2000428Andersen, T., Auk-Emblem, P., & Dornish, M. (2015). 3D Cell Culture in Alginate Hydrogels. Microarrays, 4(2), 133-161. doi:10.3390/microarrays4020133Poveda-Reyes, S., Moulisova, V., Sanmartín-Masiá, E., Quintanilla-Sierra, L., Salmerón-Sánchez, M., & Ferrer, G. G. (2016). Gelatin-Hyaluronic Acid Hydrogels with Tuned Stiffness to Counterbalance Cellular Forces and Promote Cell Differentiation. Macromolecular Bioscience, 16(9), 1311-1324. doi:10.1002/mabi.201500469Poveda-Reyes, S., Rodrigo-Navarro, A., Gamboa-Martínez, T. C., Rodíguez-Cabello, J. C., Quintanilla-Sierra, L., Edlund, U., & Ferrer, G. G. (2015). Injectable composites of loose microfibers and gelatin with improved interfacial interaction for soft tissue engineering. Polymer, 74, 224-234. doi:10.1016/j.polymer.2015.08.01

Bone-Mimicking Injectable Gelatine/Hydroxyapatite Hydrogels

[EN] Bioactive synthetic hydrogels have emerged as promising materials because they can provide molecularly tailored biofunctions and adjustable mechanical properties. To mimic the mineralogical and organic components of the natural bone, hydroxyapatite and a tyramine conjugate of gelatine were combined in this study. The effect of various amounts of in situ synthesized hydroxyapatite in gelatine-tyramine on the morphology and physical properties of injectable hydrogels was investigated. Mineralogical identification confirmed successful precipitation of in situ formed hydrox yapatite. Better distribution of hydroxyapatite crystal agglomerates within modified gelatine was found at 5 % of hydroxyapatite, which could be responsible for increased storage modulus with respect to pure gelatine hydrogel. Prepared composite hydrogels are non-toxic and support the proliferation of Hek293 cells.The authors are grateful for the financial support of the Spanish Ministry of Economy and Competitiveness through the MAT2016-76039-C4-1-R project (including Feder funds) and the Croatian Science Foundation under the project IP-2014-09-3752.Rogina, A.; Sandrk, N.; Teruel Biosca, L.; Antunovic, M.; Ivankovic, M.; Gallego Ferrer, G. (2019). Bone-Mimicking Injectable Gelatine/Hydroxyapatite Hydrogels. Chemical and Biochemical Engineering Quarterly Journal. 33(3):325-335. https://doi.org/10.15255/CABEQ.2019.1663S32533533

Novel microgel culture system as semi-solid three-dimensional in vitro model for the study of multiple myeloma proliferation and drug resistance

[EN] Multiple myeloma (MM) is a hematological malignancy in which the patient's drug resistance is one of the main clinical problems. As 2D cultures do not recapitulate the cellular microenvironment, which has a key role in drug resistance, there is an urgent need for better biomimetic models. Here, a novel 3D platform is used to model MM. The semi-solid culture consists of a dynamic suspension of microspheres and MM cells, termed as microgel. Microspheres are synthesized with acrylic polymers of different sizes, compositions, and functionalities (fibronectin or hyaluronic acid). Optimal conditions for the platform in terms of agitation speed and microsphere size have been determined. With these parameters the system allows good proliferation of the MM cell lines RPMI8226, U226, and MM1.S. Interestingly, when used for drug resistance studies, culture of the three MM cell lines in microgels showed close agreement in revealing the role of acrylic acid in resistance to anti-MM drugs such as dexamethasone and bortezomib. This work presents a unique platform for the in vitro modeling of non-solid tumors since it allows keeping non-adherent cells in suspension conditions but in a 3D context that can be easily tuned with different functionalizations.This work was funded by the Spanish State Research Agency (AEI) through the PID2019-106099RB-C41/AEI/10.13039/501100011033 Project. CIBER-BBN is an initiative funded by the VI National R&D&I Plan 2008-2011, Iniciativa Ingenio 2010, Consolider Program. CIBER Actions were financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund. This work was also supported by the Spanish Ministry of Science, Innovation and Universities through Grant NO FPU17/05810 awarded to Sandra Clara-Trujillo. The Microscopy Service of the UPV (UniversitatPolitecnica de València) is gratefully acknowledged for helping with FESEM characterization.Clara-Trujillo, S.; Tolosa, L.; Cordón, L.; Sempere, A.; Gallego Ferrer, G.; Gómez Ribelles, JL. (2022). Novel microgel culture system as semi-solid three-dimensional in vitro model for the study of multiple myeloma proliferation and drug resistance. Biomaterials Advances. 135:1-13. https://doi.org/10.1016/j.bioadv.2022.21274911313

Biomimetic microspheres for 3D mesenchymal stem cell culture and characterization

[EN] Stem cells reside in niches, specialized microenvironments that sustain and regulate their fate. Extracellular matrix (ECM), paracrine factors or other cells are key niche regulating elements. As the conventional 2D cell culture lacks these elements, it can alter the properties of naive stem cells. In this work we designed a novel biomimetic microenvironment for cell culture, consisting of magnetic microspheres, prepared with acrylates and acrylic acid copolymers and functionalized with fibronectin or hyaluronic acid as ECM coatings. To characterize cell proliferation and adhesion, porcine mesenchymal stem cells (MSCs) were grown with the different microspheres. The results showed that the 3D environments presented similar proliferation to the 2D culture and that fibronectin allows cell adhesion, while hyaluronic acid hinders it. In the 3D environments, cells reorganize the microspheres to grow in aggregates, highlighting the advantages of microspheres as 3D environments and allowing the cells to adapt the environment to their requirements.PROMETEO/2016/063 project is acknowledged. This work was partially financed with FEDER funds (CIBERONC (CB16/12/00284)). The CIBER-BBN initiative is funded by the VI National R&D&I Plan 2008-2011, Iniciativa Ingenio 2010, Consolider Program. CIBER actions are financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund. This work was also supported by the Spanish Ministry of Science, Innovation and Universities trough Sandra Clara-Trujillo FPU17/05810 grant.Clara-Trujillo, S.; Marin-Paya, JC.; Cordón, L.; Sempere, A.; Gallego Ferrer, G.; Gómez Ribelles, JL. (2019). Biomimetic microspheres for 3D mesenchymal stem cell culture and characterization. Colloids and Surfaces B Biointerfaces. 177:68-76. https://doi.org/10.1016/j.colsurfb.2019.01.050S687617

Glass transition and water dynamics in hyaluronic acid hydrogels

Glass transition and water dynamics in hydrated hyaluronic acid (HA) hydrogels crosslinked by divinyl sulfone (DVS) were studied by differential scanning calorimetry (DSC), dielectric relaxation spectroscopy (DRS) and water sorption-desorption (ESI) measurements. A critical water fraction of about h (w) = 0.17 (g of water per g of hydrated HA) for a change in the hydration properties of the material was estimated. Water crystallization was recorded by DSC during cooling and heating for water fraction values h (w) a parts per thousand yenaEuro parts per thousand 0.31. The glass transition of the hydrated system was recorded in the water fraction region 0.06 a parts per thousand currency signaEuro parts per thousand h (w) a parts per thousand currency signaEuro parts per thousand 0.59. The T (g) was found to decrease with increasing hydration level, starting from T (g) = -48 A degrees C down to about T (g) = -80 A degrees C and then to stabilize there, for the hydration levels where water crystallization occurs, suggesting that the origin of the glass transition is the combined motion of uncrystallized water molecules attached to primary hydration sites and segments of the HA chains. DRS studies revealed two relaxation peaks, associated with the main secondary relaxation process of uncrystallized water molecules (UCW) triggering the mobility of polar groups and the segmental mobility of HA chains (alpha relaxation). The alpha relaxation was in good agreement with the results by DSC. A qualitative change in the dynamics of the alpha relaxation was found for h (w) = 0.23 and was attributed to a reorganization of water in the material due to structural changes. Finally, the dielectric strength of the relaxation of UCW was found to decrease in the water fraction region of the structural changes, i.e. for h (w) similar to 0.23.Panagopoulou, A.; Vázquez Molina, J.; Kyritsis, A.; Monleón Pradas, M.; Vallés Lluch, A.; Gallego Ferrer, G.; Pissis, P. (2013). Glass transition and water dynamics in hyaluronic acid hydrogels. Food Biophysics. 8(3):192-202. doi:10.1007/s11483-013-9295-2S19220283T.C. Laurent, Ciba Foundation Symposium, vol. 143 (John Wiley and Sons, New York, 1989), pp. 1–298J. Necas, L. Bartosikov, P. Brauner, J. Kolar, Vet. Med. 53(8), 397–411 (2008)M.K. Cowman, M. Li, E.A. Balazs, Biophys. J. 75, 2030–2037 (1998)M.K. Cowman, S. Matsuoka, Carbohydr. Res. 340, 791–809 (2005)C.E. Schanté, G. Zuber, C. Herlin, T.F. Vendamme, Carbohydr. Polym. 85, 469–489 (2011)E.J. Oh, K. Park, K.S. Kim, J. Kim, J.-A. Yang, J.-H. Kong, M.Y. Lee, A.S. Hoffman, S.K. Hahn, J. Control. Release 141, 2–12 (2010)A.S. Hoffman, Adv. Drug Deliv. 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Capacitively Coupled Electrical Stimulation of Rat Chondroepiphysis Explants: A Histomorphometric Analysis

[EN] The growth plate is a cartilaginous layer present from the gestation period until the end of puberty where it ossifies joining diaphysis and epiphysis. During this period several endocrine, autocrine, and paracrine processes within the growth plate are carried out by chondrocytes; therefore, a disruption in cellular functions may lead to pathologies affecting bone development. It is known that electric fields impact the growth plate; however, parameters such as stimulation time and electric field intensity are not well documented. Accordingly, this study presents a histomorphometrical framework to assess the effect of electric fields on chondroepiphysis ex-plants. Bones were stimulated with 3.5 and 7 mV/cm, and for each electric field two exposure times were tested for 30 days (30 min and 1 h). Results evidenced that electric fields increased the hypertrophic zones compared with controls. In addition, a stimulation of 3.5 mV/cm applied for 1 h preserved the columnar cell density and its orientation. Moreover, a pre-hypertrophy differentiation in the center of the chondroepiphysis was observed when explants were stimulated during 1 h with both electric fields. These findings allow the understanding of the effect of electrical stimulation over growth plate organization and how the stimulation modifies chondrocytes morphophysiology.This research was supported by COLCIENCIAS Administrative Department of Science, Technology and Innovation. The authors gratefully thank the research support from the Biotechnology Institute of the Universidad Nacional de Colombia, for providing the lab space at the Biomimetics Laboratory and the reactants to perform the experimental approach of this study. Research reported in this publication was supported by COLCIENCIAS Administrative Department of Science, Technology and Innovation (Announcement 712-2015 Grant No 50457).Vaca-González, JJ.; Escobar, J.; Guevara, J.; Hata, YA.; Gallego Ferrer, G.; Garzón-Alvarado, DA. (2019). Capacitively Coupled Electrical Stimulation of Rat Chondroepiphysis Explants: A Histomorphometric Analysis. Bioelectrochemistry. 126:1-11. https://doi.org/10.1016/j.bioelechem.2018.11.004S11112