Cultivation and cryopreservation of rat stem cells and their interaction with lyophilised acellular matrix

Abstract

With the rapid development of regenerative medicine in the 21st century, the study of the therapeutic potential of stem cells in both preclinical research and clinical trials has become particularly relevant. Preclinical studies on animals allow for a detailed understanding of the mechanisms of action of allogzeneic cell preparations, exploring their regenerative activity, pharmacodynamics, and potential side effects. The purpose of the study was to select optimal conditions for obtaining, cultivating, and cryopreserving mesenchymal stem cells from rats and analyse their interaction with the lyophilised acellular matrix. The enzymatic method was applied to obtain primary cell cultures from the umbilical cord, dermis, and muscles of Rattus norvegicus fetuses. Cell cultures were cultivated in vitro, and cell line proliferation rates were analysed using an inverted microscope. In addition, cryopreservation was performed to store cellular materials. The interaction of mesenchymal stem cells with an acellular matrix and cryopreservation of the obtained cells was at the 4 and 5th passages. It was shown that the optimal nutrient medium for cultivating the obtained lines of mesenchymal stem cells from the umbilical cord and dermis of rat fetuses is DMEM/F12 Advanced. It was established that the method of thawing the cell suspension by 10-fold dilution of dimethyl sulfoxide is more effective than the alternative method of immediate removal of cryoprotectant by centrifugation. The lyophilised acellular dermal matrix was found to have a cytotoxic effect on all cultured rat cells, while the pericardial matrix showed a positive effect on the growth of the investigated cell lines. Thus, the optimal nutrient medium and conditions for freezing/thawing of rat stem cells were selected, and the effect of lyophilised acellular matrix, planned for therapeutic use, on the obtained cell lines was determined

Keywords

• in vitro culture
• cryopreservation
• umbilical cord
• muscles
• dermis
• lyophilised matrix
• pericardium