Abstract
Objective To study the effect of the recombinant Lentivirus containing calcitonin gene related peptide (CGRP) gene on cells biological activity and differentiation of rat bone marrow stem cells(MSCs). Methods Rat MSCs were isolated and cultured by granulocytes adherent. MSCs were transfected with Lenti-EGFP-CGRP(MSCsCGRP+ /+ group), While MSCs were transfected with Lenti-EGFP as control group. Cell transfection rate was detected by flow cytometry, protein secretion in the above-mentioned MSCsCGRP+ /+ supernatant was detected using ELISA method. Cells surface markers weare detected by flow cytometry and immunohistochemistry. Trypan blue was used to examin the survive rate, β-galactosidase staining was used to examin aging of MSCs transfection, and MTT was used to examine cell vitality. Results At first day after transfecting with Lenti-EGFP-CGRP, fluorescence was not observed by fluorescence microscope, but a small amount of CGRP protein was detected by ELISA in MSCsCGRP+ /+ group, at 3 days and 4 days after transfecting with MSCs, strong fluorescence was observed by fluorescence microscope (the cell transfection rates were 77.87% and 79.58%). The CGRP expression was significantly higher in MSCsCGRP+ /+ group than in control group 〔(19.53±0.50) pg/ml vs. (3.12±0.00) pg/ml, t=48.964, P 0.05). At 14 days after transfection with Lenti-EGFP-CGRP, aging of cell were decreased in MSCsCGRP+ /+ group as compared with control group (t=2.446, P<0.05). Conclusions After MSCs are transfected with Lenti-EGFP-CGRP, biological characteristics of MSCs has no significant effects, there is still proliferation and differentiation activity. Cell secretion of CGRP can promote the endothelial cell differentiation, and inhibit the differentiation to smooth muscle cells. The CGRP modification of MSCs may play a role in the regulation of angiogenesis. Key words: Mesenchymal stem cells; Calcitonin gene-related peptide; Transfection; Cytological techniques
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