Adenovirus-mediated cardiotrophin-1 gene for modifying neural stem cells in rats
Aim: To provide material foundation for the transplantation of neural stem cells (NSCs) modified by cardiotrophin-1 (CT-1) gene in the treatment of central nervous system injury with the CT-1 gene modifying NSCs in rats. Methods: The experiment was performed at the Military Key Opened Laboratory, Institute of Battle Surgery, Third Military Medical University of Chinese PLA from June 2002 to June 2003. The embryos NSCs were isolated and cultured from embryo hippocampus and ependyma region in Wistar rats in order to establish the purified recombined adenovirus (rAd)-CT1 and rAd-enhanced green fluorescent protein (EGFP) carrier transfection into NSCs. The condition of growth and living was observed directly under the differential microscope. The NSCs and the special nestin, neurofilament, glial fibriliary acidic protein (GFAP) exogenous Brdu of the differentiation cells of NSCs after inducing and the mixed status of the EGFP were detected with immunocytochemical method and immunofluorescence technique, and the expression of the CT-1 gene in the NSCs was observed. Results: 1 The NSCs separated from embryo of rats could do many times of passage in vitro. The cell proliferation marker, Brdu mixture verified that the NSCs had the ability of splitting and proliferation. 2 The NSCs marker-nestin, neuron marker-neurofilament and the glial cell marker-GFAP were detected, which indicated that the NSCs had the multipotential differentiation. 3 After the transfection of rAd-CT1 and rAd-EGFP into NSCs. the CT1 and the expression of reported gene EGFP in the NSCs continuously increased significantly detected with immunohistochemical method and fluorescent tracer technique. Conclusion: 1 NSCs are isolated and cultured in the experiment successfully, and can perform passage proliferation in vitro in many times and can trans-differentiation into neurons and glial cells. 2 After the transfection of rAd-CT1 and rAd-EGFP into NSCs, the exogenous CT-1 gene and EGFP can express stablely in NSCs.