Histology and histopathology Vol.30,nº12 (2015)
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- PublicationOpen AccessChondrocyte differentiation for auricular cartilage reconstruction using a chitosan based hydrogel(Universidad de Murcia. Departamento de Biología Celular e Histología, 2015) García-López, J.; Garciadiego-Cázares, D.; Melgarejo-Ramírez, Y.; Sánchez-Sánchez, R.; Solís-Arrieta, L.; García-Carvaja, Z.; Sánchez-Betancourt, J.I.; Ibarra, C.; Luna-Bárcena, G.; Velasquillo, C.Tissue engineering with the use of biodegradable and biocompatible scaffolds is an interesting option for ear repair. Chitosan-Polyvinyl alcohol-Epichlorohydrine hydrogel (CS-PVA-ECH) is biocompatible and displays appropriate mechanical properties to be used as a scaffold. The present work, studies the potential of CS-PVA-ECH scaffolds seeded with chondrocytes to develop elastic cartilage engineered-neotissues. Chondrocytes isolated from rabbit and swine elastic cartilage were independently cultured onto CS-PVA-ECH scaffolds for 20 days to form the appropriate constructs. Then, in vitro cell viability and morphology were evaluated by calcein AM and EthD-1 assays and Scanning Electron Microscopy (SEM) respectively, and the constructs were implanted in nu/nu mice for four months, in order to evaluate the neotissue formation. Histological analysis of the formed neotissues was performed by Safranin O, Toluidine blue (GAG’s), Verhoeff-Van Gieson (elastic fibers), Masson’s trichrome (collagen) and Von Kossa (Calcium salts) stains and SEM. Results indicate appropriate cell viability, seeded with rabbit or swine chondrocyte constructs; nevertheless, upon implantation the constructs developed neotissues with different characteristics depending on the animal species from which the seeded chondrocytes came from. Neotissues developed from swine chondrocytes were similar to auricular cartilage, while neotissues from rabbit chondrocytes were similar to hyaline cartilage and eventually they differentiate to bone. This result suggests that neotissue characteristics may be influenced by the animal species source of the chondrocytes isolated.
- PublicationOpen AccessTranscription factors GATA4 and TBX5 promote cardiomyogenic differentiation of rat bone marrow mesenchymal stromal cells(Universidad de Murcia. Departamento de Biología Celular e Histología, 2015) Chen, Wei; Zhang, Lei; Shao, Su-Xia; Wang, Hai-Ping; Cui, Shi-Jie; Zhang, Ya-Nan; Kong, Xiang-Zeng; Yin, Qing; Zhang, Jin-Ping. Bone marrow mesenchymal stromal cells (BMSCs) have potential applications in cell and gene therapies for cardiac disease. The cardiac-specific transcription factors GATA-binding protein 4 (GATA4) and T-Box protein 5 (TBX5) are considered to be pivotal in cardiogenesis. The aim of this study was to investigate the effects of GATA4 and TBX5 on cardiomyogenic differentiation of BMSCs. The BMSCs were initially isolated and identified. Vectors harboring cardiac transcription factor genes GATA4 and TBX5 or empty vectors were transferred into BMSCs. Cardiomyogenic cells differentiated from BMSCs were identified by expression of cardiac-specific markers including cardiac troponin T, connexin 43, β-myosin heavy chain, and myosin light chain-2 using immunocytochemical staining, western blotting, and quantitative real-time PCR. The ultrastructures of the differentiated cells were examined by transmission electron microscopy, which were similar to those of fetal cardiomyocytes. The differentiated cells exhibited L-type calcium current activities reflective of the electrophysiological characteristics of cardiomyocytes. These findings indicate that exogenous expression of cardiac-specific transcription factors GATA4 and TBX5 enhance cardiomyogenic differentiation of BMSCs
- PublicationOpen AccessDevelopment and preclinical evaluation of a new galactomannan-based dressing with antioxidant properties for wound healing(Universidad de Murcia. Departamento de Biología Celular e Histología, 2015) Castro, Begoña; Palomares, Teodoro; Azcoitia, Iker; Bastida, Felix; del Olmo, Maite; Soldevilla, Javier J.; Alonso-Varona, AnaWe describe a novel wound dressing (HR006) with two components: a lyophilized matrix of the galactomannan from locust bean gum (LBG) and an antioxidant hydration solution (AHsol ) containing curcumin and N-acetyl-L-cysteine (NAC). Physicostructural analyses of the LBG matrix revealed homogeneous interconnected pores with high absorbing capacity showing excellent properties for moist wound care (MWC). In an in vitro oxidative stress fibroblast injury model, the AHsol showed relevant protective effects reducing intracellular reactive oxygen species (ROS) production, rescuing cell viability, and regulating expression of inflammation-related genes (COX-2, TNFα, IL-1α, IL-1β). The new dressing showed good biocompatibility profile as demonstrated by cytotoxicity, hemocompatibility, and skin irritation tests. Moreover, in an in vivo skin wound model in pigs, this dressing enhanced the production of healthy and organized granulation tissue and re-epithelization. In summary, HR006 exhibits significant antioxidant activity, good biocompatibility, and excellent repair capabilities improving tissue remodeling and the healing of wounds.
- PublicationOpen AccessChrysin attenuates cardiomyocyte apoptosis and loss of intermediate filaments in a mouse model of mitoxantrone cardiotoxicity(Universidad de Murcia. Departamento de Biología Celular e Histología, 2015) Anghe, N.; Cotoraci, C.; Ivan, A.; Suciu, M.; Herman, H.; Balta, C.; Nicolescu, L.; Olariu, T.; Galajda, Z.; Ardelean, A.; Hermenean, A.y. Chrysin (CHR) is a natural flavonoid and is present in high concentration in honey, propolis and many plant extracts. The aim of the present study was to evaluate the effects of CHR to reduce cardiomyocyte apoptosis and loss of intermediate filaments in a mouse model of mitoxantrone cardiotoxicity. Morphology of the cardiomyocytes was determined by optic and transmission electron microscopy and biochemistry methods. The expression of Bcl-2, Bax and Caspase-3 were assessed by immunofluorecence. Tunel assay was used to assess apoptosis in cardiomyocytes. In addition, the distribution of desmin protein was evaluated using immunohistochemistry. Our results show that MTX treatment significantly increased serum levels of creatine kinase isoenzyme (CK-MB), indicator of cardiac injury and withdrawn under CHR protection. Expression levels of Bcl-2 decreased, while those of Bax and caspase-3 increased following MTX treatment. 50 mg/kg of daily CHR intake reduced Bax and caspase-3 immunopositivity and restored Bcl-2 levels to a value comparable to the control. TUNEL (+) cardiomyocyte nuclei of MTX group showed typical signs of apoptosis which almost completely disappeared in response to 50 mg/kg CHR treatment. In parallel, an irregular distribution and a weak expression of desmin is associated with MTX induced cardiotoxic effects which was also restored by CHR treatment. In conclusion chrysin inhibits MTX-triggered cardiomyocyte apoptosis via multiple pathways, including decrease of the Bax/Bcl-2 ratio and caspase-3 expression along with preservation of the desmin disarray.
- PublicationOpen AccessOligodendroglial markers in the cuprizone model of CNS de- and remyelination(Universidad de Murcia. Departamento de Biología Celular e Histología, 2015) Salinas Tejedor, Laura; Gudi, Viktoria; Kucman, Valeria; Pul, Refik; Gingele, Stefan; Sühs, Kurt-Wolfram; Stangel, Martin; Skripuletz, Thomasy. Oligodendrocytes are the myelinating cells of the central nervous system. Since many studies of demyelinating diseases focus their research on this cell type, there is growing interest for obtaining reliable markers that can specifically recognize oligodendroglia. Established markers are the myelin-associated neurite outgrowth inhibitor (NogoA), the transcription factor Olig2, and the antibody CC-1, the latter being directed against the protein adenomatous polyposis coli (APC). Unfortunately, it has been discussed whether APC and Olig2 could recognize astrocytes under pathological conditions as well. Hence, we performed immunohistochemical studies using the oligodendroglial markers NogoA, APC, and Olig2 in a murine model of cuprizone induced demyelination. We have found that APC colocalizes with NogoA and does not co-localize with the astrocytic marker GFAP. Olig2 shows co-localization with APC but there is also a small population of Olig2/GFAP double positive cells. Some Olig2/GFAP double positive cells are found in the corpus callosum in a narrow time window in which oligodendrocyte precursor cells proliferate in this model. In other brain regions including the cerebral cortex and hippocampus and in all regions in untreated control mice double positive Olig2/GFAP cells do not occur. In conclusion, our results underline that APC and NogoA are reliable markers for detection of mature oligodendrocytes. Olig2 is a suitable marker to stain cells of oligodendroglial origin but could be combined with GFAP to exclude the GFAP positive population of cells from the quantification of oligodendroglia.
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