Harvesting of bone marrow mesenchymal stem cells from live rats and the in vitro differentiation of bone marrow mesenchymal stem cells into neuron-like cells

Abstract

In the bone marrow, there are certain populations of stem cell sources with the capacity to differentiate into several different types of cells. Ideally, cell transplants would be readily obtainable, easy to expand and bank, and capable of surviving for sufficient periods of time. Bone marrow mesenchymal stem cells (BM-MSCs) possess all of these characteristics. One of the most important benefits in using BM-MSCs is the possibility of autologous therapy. Recently, numerous studies have evaluated strategies that attempt to promote axonal regeneration in central nervous system (CNS) injuries. Among these strategies, cell transplantation is considered to be the most effective way. The differentiation of stem cells into different neural lineages (such as astrocytes and neural like cells) before transplantation has a critical role in achieving the best results in studies of CNS injury. In this study, BM-MSCs were isolated from bone marrow aspirates taken from the femurs of 103 live rats. The detection of BM-MSCs was performed with RT-PCR analysis, and they were then induced to differentiate into neuron-like cells in serum-withdrawal medium over a two week period using a multistep protocol. In addition to the morphological evaluation of differentiated cells, the process of neural differentiation was proven by immunocytochemical techniques using primary antibodies to Neuron Specific Enolase (NSE) to assess cell differentiation. PT-PCR analysis was performed for the evaluation of neural specific genes, which included NSE, MAP2, nestin, and ?-tubulin. Morphological evaluations detect neuron like cells with longitudinal processes. Using RT- PCR and immunocytochemistry assays, neuron specific genes and proteins following treatment of cells in serum-withdrawal induction medium was expressed. This study showed a simple and practical method for differentiating MSCs into neuron like cells, and feasibility of aspirate bone marrow from a live rat for autologous grafting.