Examinando por Autor "Marchal, Juan Antonio"
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Ítem A soft 3D polyacrylate hydrogel recapitulates the cartilage niche and allows growth-factor free tissue engineering of human articular cartilage(Elsevier, 2019) Jiménez-González, Gema; Venkateswaran, Seshasailam; López-Ruiz, Elena; Perán, Macarena; Pernagallo, Salvatore; Díaz-Manchón, Juan José; Canadas, Raphaël; Antich, Cristina; Oliveira, Joaquim Miguel; Callanan, Anthony; Wallace, Robert; Reis, Rui L.; Montañez, Elvira; Carrillo-Delgado, Esmeralda; Bradley, Mark; Marchal, Juan AntonioCartilage degeneration or damage treatment is still a challenge, but, tissue engineering strategies, which combine cell therapy strategies, which combine cell therapy and scaffolds, and have emerged as a promising new approach. In this regard, polyurethanes and polyacrylates polymers have been shown to have clinical potential to treat osteochondral injuries. Here, we have used polymer microarrays technology to screen 380 different polyurethanes and polyacrylates polymers. The top polymers with potential to maintain chondrocyte viability were selected, with scale-up studies performed to evaluate their ability to support chondrocyte proliferation during long-term culture, while maintaining their characteristic phenotype. Among the selected polymers, poly (methylmethacrylate-co methacrylic acid), showed the highest level of chondrogenic potential and was used to create a 3D hydrogel. Ultrastructural morphology, microstructure and mechanical testing of this novel hydrogel revealed robust characteristics to support chondrocyte growth. Furthermore, in vitro and in vivo biological assays demonstrated that chondrocytes cultured on the hydrogel had the capacity to produce extracellular matrix similar to hyaline cartilage, as shown by increased expression of collagen type II, aggrecan and Sox9, and the reduced expression of the fibrotic marker’s collagen type I. In conclusion, hydrogels generated from poly (methylmethacrylate-co-methacrylic acid) created the appropriate niche for chondrocyte growth and phenotype maintenance and might be an optimal candidate for cartilage tissue-engineering applications.Ítem Activin A/BMP2 chimera AB235 drives efficient redifferentiation of long term cultured autologous chondrocytes(Nature Research, 2015) Jiménez-González, Gema; López-Ruiz, Elena; Kwiatkowski, Witek; Montañez, Elvira; Arrebola, Francisco; Carrillo-Delgado, Esmeralda; Gray, Peter; Izpisua, Juan Carlos; Choe, Senyon; Perán, Macarena; Marchal, Juan AntonioAutologous chondrocyte implantation (ACI) depends on the quality and quantity of implanted cells and is hindered by the fact that chondrocytes cultured for long periods of time undergo dedifferentiation. Here we have developed a reproducible and efficient chondrogenic protocol to redifferentiate chondrocytes isolated from osteoarthritis (OA) patients. We used morphological, histological and immunological analysis together with a RT-PCR detection of collagen I and collagen II gene expression to show that chondrocytes isolated from articular cartilage biopsies of patients and subjected to long-term culture undergo dedifferentiation and that these cells can be redifferentiated following treatment with the chimeric Activin A/BMP2 ligand AB235. Examination of AB235-treated cell pellets in both in vitro and in vivo experiments revealed that redifferentiated chondrocytes synthesized a cartilage-specific extracellular matrix (ECM), primarily consisting of vertically-orientated collagen fibres and cartilage-specific proteoglycans. AB235 treated cell pellets also integrated into the surrounding subcutaneous tissue following transplantation in mice as demonstrated by their dramatic increase in size while non-treated control pellets disintegrated upon transplantation. Thus, our findings describe an effective protocol for the promotion of redifferentiation of autologous chondrocytes obtained from OA patients and the formation of a cartilage-like ECM that can integrate into the surrounding tissue in vivo.Ítem Cellular extracts from post-mortem human cardiac tissue direct cardiomyogenic differentiation of human adipose tissue-derived stem cells(Elsevier, 2013) Perán, Macarena; López-Ruiz, Elena; González-Herrera, Lucas; Bustamante, Milán; Valenzuela, Aurora; Marchal, Juan AntonioBackground aims. Human adipose tissue-derived stem cells (hASCs) can be easily (and inexpensively) expanded in culture, and their high plasticity allows their conversion to different cell types. We study the potential capacity of postmortem cardiac tissue to direct cardiac differentiation of hASCs in vitro. Methods. Cardiac tissue collected from autopsies was used to obtain cell extracts and conditioned medium, and both approaches were tested for cardiac induction. Results. Gene expression analyses proved that post-mortem human cardiac tissue maintains genetic integrity. hASCs exposed to the cell extracts or conditioned medium for 2 weeks achieved the appearance of myotube-like structures and were positive for cardiac markers such as sarcomeric a-actinin, cardiac troponin I and T and desmin as proved by immunofluorescence. In addition, differentiated cells showed increased expression of cardiomyocyte-related genes analyzed by reverse transcriptase polymerase chain reaction (GATA-4, myocyte-enhancer factor-2c, a-cardiac actin and cardiac troponin I). Conclusions. For the first time, post-mortem human cardiac tissue was used to induce hASC differentiation into myocardial-like cells. The methodology described here would serve as a useful model to obtain cardiomyocyte-like cells in vitro.Ítem Cellular extracts from post-mortem human cardiac tissue direct cardiomyogenic differentiation of human adipose tissue-derived stem cells(Elsevier, 2010) Perán, Macarena; Marchal, Juan Antonio; López-Ruiz, Elena; Jiménez-Navarro, Manuel; Boulaiz, Houria; Rodríguez-Serrano, Fernado; Carrillo-Delgado, Esmeralda; Sánchez-Espín, Gema; de-Teresa, Eduardo; Tosh, David; Aránega, AntoniaBackground aims. The goal was to induce the transdifferentiation (or conversion) of human adipose-derived stem cells to cardiomyocytes using an intracellular extract obtained from adult human heart tissue. Methods. Human adult stem cells from lipoaspirates were transiently permeabilized, exposed to human atrial extracts and allowed to recover in culture. Results. After 21 days, the cells acquired a cardiomyocyte phenotype, as demonstrated by morphologic changes (appearance of binucleate, striated cells and branching fibers), immunofl uorescence detection of cardiac-specifi c markers (connexin-43, sarcomeric a-actinin, cardiac troponin I and T, and desmin) and the presence of cardiomyocyte-related genes analyzed by reverse transcription – polymerase chain reaction (cardiac myosin light chain 1, a -cardiac actin, cardiac troponin T and cardiac b -myosin). Conclusions. We have demonstrated for the fi rst time that adult cardiomyocytes obtained from human donors retain the capacity to induce cardiomyocyte differentiation of mesenchymal stromal cells. The use of autologous extracts for reprogramming adult stem cells may have potential therapeutic implications for treating heart disease.Ítem Chondrocytes extract from patients with osteoarthritis induces chondrogenesis in infrapatellar fat pad-derived stem cells(Elsevier, 2013) López-Ruiz, Elena; Perán, Macarena; Cobo-Molinos, Jesús; Jiménez-González, Gema; Picón, Manuel; Bustamante, Milán; Arrebola, Francisco; Hernández-Lamas, María Carmen; Delgado-Martínez, Alberto Damián; Montañez, Elvira; Marchal, Juan AntonioObjective: Infrapatellar fat pad of patients with osteoarthritis (OA) contains multipotent and highly clonogenic adipose-derived stem cells that can be isolated by low invasive methods. Moreover, nuclear and cytoplasmic cellular extracts have been showed to be effective in induction of cell differentiation and reprogramming. The aim of this study was to induce chondrogenic differentiation of autologous mesenchymal stem cells (MSCs) obtained from infrapatellar fat pad (IFPSCs) of patients with OA using cellular extracts-based transdifferentiation method. Design: IFPSCs and chondrocytes were isolated and characterized by flow cytometry. IFPSCs were permeabilized with Streptolysin O and then exposed to a cell extract obtained from chondrocytes. Then, IFPSCs were cultured for 2 weeks and chondrogenesis was evaluated by morphologic and ultrastructural observations, immunologic detection, gene expression analysis and growth on 3-D poly (DL-lactic-coglycolic acid) (PLGA) scaffolds. Results: After isolation, both chondrocytes and IFPSCs displayed similar expression of MSCs surface makers. Collagen II was highly expressed in chondrocytes and showed a basal expression in IFPSCs. Cells exposed to chondrocyte extracts acquired a characteristic morphological and ultrastructural chondrocyte phenotype that was confirmed by the increased proteoglycan formation and enhanced collagen II immunostaining. Moreover, chondrocyte extracts induced an increase in mRNA expression of chondrogenic genes such as Sox9, L-Sox5, Sox6 and Col2a1. Interestingly, chondrocytes, IFPSCs and transdifferentiated IFPSCs were able to grow, expand and produce extracellular matrix (ECM) on 3D PLGA scaffolds. Conclusions:We demonstrate for the first time that extracts obtained from chondrocytes of osteoarthritic knees promote chondrogenic differentiation of autologous IFPSCs. Moreover, combination of transdifferentiated IFPSCs with biodegradable PLGA 3D scaffolds can serve as an efficient system for themaintenance and maturation of cartilage tissue. These findings suggest its usefulness to repair articular surface in OA.Ítem Evaluation of Glycerylphytate Crosslinked Semi- and Interpenetrated Polymer Membranes of Hyaluronic Acid and Chitosan for Tissue Engineering(MDPI, 2020) Mora Boza, Ana; López Ruiz, Elena; López Donaire, María Luisa; Jiménez, Gema; Rosa Aguilar, María; Marchal, Juan Antonio; Perdiz, Jose Luis; Vázquez Lasa, Blanca; San Román, Julio; Gálvez Martín, PatriciaIn the present study, semi- and interpenetrated polymer network (IPN) systems based on hyaluronic acid (HA) and chitosan using ionic crosslinking of chitosan with a bioactive crosslinker, glycerylphytate (G1Phy), and UV irradiation of methacrylate were developed, characterized and evaluated as potential supports for tissue engineering. Semi- and IPN systems showed significant di erences between them regarding composition, morphology, and mechanical properties after physicochemical characterization. Dual crosslinking process of IPN systems enhanced HA retention and mechanical properties, providing also flatter and denser surfaces in comparison to semi-IPN membranes. The biological performance was evaluated on primary human mesenchymal stem cells (hMSCs) and the systems revealed no cytotoxic e ect. The excellent biocompatibility of the systems was demonstrated by large spreading areas of hMSCs on hydrogel membrane surfaces. Cell proliferation increased over time for all the systems, being significantly enhanced in the semi-IPN, which suggested that these polymeric membranes could be proposed as an e ective promoter system of tissue repair. In this sense, the developed crosslinked biomimetic and biodegradable membranes can provide a stable and amenable environment for hMSCs support and growth with potential applications in the biomedical field.Ítem Functionalized nanostructures with application in regenerative medicine(MPDI, 2012) Perán, Macarena; García-Chaves, María Ángel; López-Ruiz, Elena; Bustamante, Milán; Jiménez-González, Gema; Madeddu, Roberto; Marchal, Juan AntonioIn the last decade, both regenerative medicine and nanotechnology have been broadly developed leading important advances in biomedical research as well as in clinical practice. The manipulation on the molecular level and the use of several functionalized nanoscaled materials has application in various fields of regenerative medicine including tissue engineering, cell therapy, diagnosis and drug and gene delivery. The themes covered in this review include nanoparticle systems for tracking transplanted stem cells, self-assembling peptides, nanoparticles for gene delivery into stem cells and biomimetic scaffolds useful for 2D and 3D tissue cell cultures, transplantation and clinical application.Ítem How Can Nanotechnology Help to Repair the Body? Advances in Cardiac, Skin, Bone, Cartilage and Nerve Tissue Regeneration(MPDI, 2013) Perán, Macarena; García, María Ángel; López Ruiz, Elena; Jiménez, Gema; Marchal, Juan AntonioNanotechnologists have become involved in regenerative medicine via creation of biomaterials and nanostructures with potential clinical implications. Their aim is to develop systems that can mimic, reinforce or even create in vivo tissue repair strategies. In fact, in the last decade, important advances in the field of tissue engineering, cell therapy and cell delivery have already been achieved. In this review, we will delve into the latest research advances and discuss whether cell and/or tissue repair devices are a possibility. Focusing on the application of nanotechnology in tissue engineering research, this review highlights recent advances in the application of nano-engineered scaffolds designed to replace or restore the followed tissues: (i) skin; (ii) cartilage; (iii) bone; (iv) nerve; and(v) cardiac.Ítem Human cardiac tissue induces transdifferentiation of adult stem cells towards cardiomyocytes(Elsevier, 2010) Perán, Macarena; Marchal, Juan Antonio; López-Ruiz, Elena; Jiménez-Navarro, Manuel; Boulaiz, Houria; Rodríguez-Serrano, Fernando; Carrillo-Delgado, Esmeralda; Sánchez-Espín, Gema; de-Teresa, Eduardo; Tosh, David; Aránega, AntoniaBackground aims. The goal was to induce the transdifferentiation (or conversion) of human adipose-derived stem cells to cardiomyocytes using an intracellular extract obtained from adult human heart tissue. Methods. Human adult stem cells from lipoaspirates were transiently permeabilized, exposed to human atrial extracts and allowed to recover in culture. Results. After 21 days, the cells acquired a cardiomyocyte phenotype, as demonstrated by morphologic changes (appearance of binucleate, striated cells and branching fi bers), immunofl uorescence detection of cardiac-specifi c markers (connexin-43, sarcomeric a-actinin, cardiac troponin I and T, and desmin) and the presence of cardiomyocyte-related genes analyzed by reverse transcription – polymerase chain reaction (cardiac myosin light chain 1, a cardiac actin, cardiac troponin T and cardiac b-myosin). Conclusions. We have demonstrated for the fi rst time that adult cardiomyocytes obtained from human donors retain the capacity to induce cardiomyocyte differentiation of mesenchymal stromal cells. The use of autologous extracts for reprogramming adult stem cells may have potential therapeutic implications for treating heart disease.Ítem Papel de las células madre tumorales en la respuesta a nuevos fármacos selectivos frente a cáncer de colon y mama(Jaén : Universidad de Jaén, 2014) Ramírez-Rivera, Alberto; Marchal, Juan Antonio; Perán, Macarena; Boulaiz, Houria; Universidad de Jaén. Departamento de Ciencias de la Salud[ES] El cáncer es una de las enfermedades más importantes a nivel mundial. Las células madre cancerígenas (CSC) son clave en el proceso de tumorigénesis y en la aparición de quimio y radioresistencia, facilitando la aparición de recurrencia y metástasis. Actualmente hay un gran interés en desarrollar terapias selectivas frente a las células tumorales y las CSC. Nuestro objetivo ha sido seleccionar, de un conjunto de fármacos de nueva síntesis, el compuesto con más actividad antitumoral. Bozepinib ha mostrado una potente y selectiva actividad antiproliferativa frente a células tumorales y CSC. Además, actúa sobre vías metabólicas de la tumorigenesis y CSCs, poniendo de relieve su actividad anti cses. In vivo, Bozepinib no induce toxicidad aguda ni crónica a altas dosis y posee una potente actividad antitumoral. Estos resultados indican la eficacia de Bozepinib como agente farmacológico frente a CSCs pudiendo mejorar la respuesta de los pacientes con este tipo de patología.Ítem Study and evaluation of the efficacy of pancreatic proenzymes against the tumor microenvironment and chemoresistance in pancreatic cancer(2024-12-13) Toledo, María Belén; Perán, Macarena; Marchal, Juan Antonio; Giovanetti, Elisa; Universidad de Jaén. Departamento de Ciencias de la SaludEl cáncer es la segunda causa principal de muerte en los países desarrollados y se caracteriza por el crecimiento celular descontrolado y la diseminación a otros órganos. El adenocarcinoma ductal pancreático (PDAC) es uno de los tipos de cáncer más agresivos y letales, con una supervivencia a cinco años de menos del 12%. Su desarrollo está impulsado por mutaciones genéticas en genes clave y por un microambiente tumoral (TME) que favorece la quimiorresistencia. Este tipo de cáncer enfrenta grandes desafíos terapéuticos debido a su heterogeneidad y a la resistencia a tratamientos convencionales actuales como la quimioterapia. Esta tesis se ha centrado en el papel de las (pro)enzimas pancreáticas, que muestran promesas en la lucha contra las células quimioresistentes, en elementos del TME y en la inhibición de la progresión tumoral y la metástasis. Los enfoques terapéuticos utilizando estas (pro)enzimas combinadas con otros tratamientos podrían mejorar los resultados para los pacientes con PDAC y abordar la quimiorresistencia de manera efectiva. Cancer is the second leading cause of death in developed countries and is characterized by uncontrolled cellular growth and dissemination to other organs. Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal types of cancer, with a five-year survival rate of less than 12%. Its development is driven by genetic mutations in key genes and by a tumor microenvironment (TME) that promotes chemoresistance. This type of cancer faces significant therapeutic challenges due to its heterogeneity and resistance to current conventional treatments such as chemotherapy. This thesis has focused on the role of (pro)enzymes, which show promise in combating chemoresistant cells, elements of the TME, and in inhibiting tumor progression and metastasis. Therapeutic approaches utilizing these (pro)enzymes in combination with other treatments could improve outcomes for patients with PDAC and effectively address chemoresistance.Ítem Ultrastructural and molecular analyzes of insulin-producing cells induced from human hepatoma cells(Elsevier, 2011) Perán, Macarena; Sánchez-Ferrero, Aitor; Tosh, David; Marchal, Juan Antonio; López-Ruiz, Elena; Álvarez-Aránega, Pablo; Boulaiz, Houria; Rodríguez-Serrano, Fernando; Aránega, AntoniaBackground aims. Diabetes type I is an autoimmune disease characterized by the destruction of pancreatic insulin-producing (beta-) cells and resulting in external insulin dependence for life. Islet transplantation represents a potential treatment for diabetes but there is currently a shortage of suitable organs donors. To augment the supply of donors, different strategies are required to provide a potential source of beta-cells. These sources include embryonic and adult stem cells as well as differentiated cell types. The main goal of this study was to induce the transdifferentiation (or conversion of one type cell to another) of human hepatoma cells (HepG2 cells) to insulin-expressing cells based on the exposure of HepG2 cells to an extract of rat insulinoma cells (RIN). Methods. HepG2 cells were fi rst transiently permeabilized with Streptolysin O and then exposed to a cell extract obtained from RIN cells. Following transient exposure to the RIN extract, the HepG2 cells were cultured for 3 weeks. Results. Acquisition of the insulin-producing cell phenotype was determined on the basis of (i) morphologic and (ii) ultrastructural observations, (iii) immunologic detection and (iv) reverse transcription (RT)-polymerase chain reaction (PCR) analysis. Conclusions. This study supports the use of cell extract as a feasible method for achieve transdifferentiation of hepatic cells to insulin-producing cells.Ítem Validation of the 1,4-butanediol thermoplastic polyurethane as a novel material for 3D bioprinting applications(John Wiley & Sons, 2020-11) Chocarro-Wrona, Carlos; de-Vicente, Juan; Antich, Cristina; Jiménez-González, Gema; Martínez-Moreno, Daniel; Carillo-Delgado, Esmeralda; Montañez, Elvira; Gálvez-Martín, Patricia; Perán, Macarena; López-Ruiz, Elena; Marchal, Juan AntonioTissue engineering (TE) seeks to fabricate implants that mimic the mechanical strength, structure, and composition of native tissues. Cartilage TE requires the development of functional personalized implants with cartilage-like mechanical properties capable of sustaining high load-bearing environments to integrate into the surrounding tissue of the cartilage defect. In this study, we evaluated the novel 1,4-butanediol thermoplastic polyurethane elastomer (b-TPUe) derivative filament as a 3D bioprinting material with application in cartilage TE. The mechanical behavior of b-TPUe in terms of friction and elasticity were examined and compared with human articular cartilage, PCL, and PLA. Moreover, infrapatellar fat pad-derived human mesenchymal stem cells (MSCs) were bioprinted together with scaffolds. in vitro cytotoxicity, proliferative potential, cell viability, and chondrogenic differentiation were analyzed by Alamar blue assay, SEM, confocal microscopy, and RT-qPCR. Moreover, in vivo biocompatibility and host integration were analyzed. b-TPUe demonstrated a much closer compression and shear behavior to native cartilage than PCL and PLA, as well as closer tribological properties to cartilage. Moreover, b-TPUe bioprinted scaffolds were able to maintain proper proliferative potential, cell viability, and supported MSCs chondrogenesis. Finally, in vivo studies revealed no toxic effects 21 days after scaffolds implantation, extracellular matrix deposition and integration within the surrounding tissue. This is the first study that validates the biocompatibility of b-TPUe for 3D bioprinting. Our findings indicate that this biomaterial can be exploited for the automated biofabrication of artificial tissues with tailorable mechanical properties including the great potential for cartilage TE applications. © 2020 The Authors. Bioengineering & Translational Medicine published by Wiley Periodicals LLC on behalf of American Institute of Chemical Engineers.