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Information × Registration Number 0213U002059, 0107U003342 , R & D reports Title The transforming potential of native and modifiated stem cells and it realization in vitro and in vivo. popup.stage_title Head Kordium Vitaly Arnoldovich, Доктор біологічних наук Registration Date 14-01-2013 Organization Institute of Molecular Biology and Genetics popup.description2 Report 192., 120 fig., 1 tabl.bibliografii. 278 items.Object of study - mesenhymal stromal (stem) cells (MSC) from human umbilical cord Wharton's jelly and recombinant genes of several cytokines. Methods: molecular cloning, expression of the recombinant proteins, chromatography, sequencing, RT-PCR, Western-blot analysis, immunoprecipitation, transfection, methods of cell biology (eukaryotic cells cultivation, cyto-and histochemical studies, light and confocal microscopy, FACS-analysis). The past several years witnessed a rapid development of the field of stem cell research concerning mesenhymal (stem) cells, which are considered to be one of the most promising sources of autologous material for cell therapy and tissue engineering. Their potential for the differentiation into the different types of cells depending on the cytokine profile, immunomodulatory activity, and the potential for the reparation and restoration of functional tissues have been extensively tested to treat a variety of pathologies. The comparative analysis of different methods for isolation of MSC from human umbilical cord Wharton's jelly, depending on the quality of starting material, was performed as a part of this study on a large number of umbilical cords. The influence of different cultivation parameters (source of growth factors, composition of gas mixture, utilization developed of 3D matrices), and of the number of passages on the characteristics of obtained MSC (morphology, expression of surface markers) has been evaluated. The possible recommendations for the isolation and for future design of in vivo experiments have been developed on the basis of the research conducted. The artificial matrices from collagen I and II type have been developed. The matrices were characterized by scanning electron microscopy, and the data obtained from in vitro experiments revealed that they were able to support the proliferation and differentiation of MSC particularly into chondrocytes. Within the scope of this research we have studied the influence of intrarticular administration of isolated MSC on the course of the experimental osteoarthritis induced in rats. It has been demonstrated that the administration of MSC improved several indicators of osteoarthritis. Since the cytokines determine the fate and role of MCS in the body, as a preliminary stage of study of diverse potential of MCSs in vivo, the genes of human IL-10, SDF-1? and VEGF121 were cloned. The producers of these cytokines had been developed, the recombinant proteins were purified and their biological activity was confirmed. The recombinant genes of IL-10, LIF, and FGF-2 were used for the development of the vectors, which provide the expression of the genes in eukaryotic cells. Several transgenic cell sublines, that carry the genes, were obtained by transfection. The Western-blot data analysis revealed that LIF and FGF-2 were secreted by cell sublines carrying corresponding transgene. The cells of the developed genetically modified CHO-K1 cell line were encapsulated into alginate microbeads. It had been demonstrated that during the cultivation of encapsulated cells the products of the expression of recombinant genes were secreted into in culture medium. The method of the non-viral gene delivery to MSC, mediated by glycosylated PEI, had been optimized. The transfection experiments conducted with optimized protocol revealed the percentage of successfully transfected MSCs to be 4%. Thus, as a result of the conducted studies, the next phase of research aimed at realization of the reparative and restoring potential of MSC in vivo had been prepared. Product Description popup.authors В.І. Андрієнко Д.М. Іродов Л.І.Лихачева Л.Т. Рачкова М.В.Драгулян М.В.Ковальчук Н.С. Шувалова О.В. Окунєв О.Г. Дерябіна О.К.Топорова П.В. Моргунов С.І. Черних С.П.Шпильова С.Ю. Римар Т.М. Луценко Т.П. Гулько Ю.С Ніколаєв Я.О. Похоленко popup.nrat_date 2020-04-02 Close