Abstract:

BACKGROUND: Gene transfection can elevate the survival of bone marrow mesenchymal stem cells (BMSCs) in transplanted region, but there are a few reports addressing proliferation, apoptosis and secretion of adrenomedullin (ADM) gene-transfected BMSCs under hypoxic condition.
OBJECTIVE: To observe effects of ADM gene transfection on BMSCs proliferation, apoptosis and secretion of vascular endothelial growth factor under hypoxic and serum-free culture conditions.
METHODS: Rat BMSCs were isolated and amplified in vitro by differential adhesion method. The infection efficiency of recombinant adenovirus Ad-ADM in BMSCs was tested by x-gal staining. BMSCs were divided into non-transfection group, empty vector group and Ad-ADM transfection group. BMSCs cultured under hypoxia and serum-free conditions for 0, 3, 6, 9, 12, 16, 20, 24 hours respectively. Cell proliferation was detected by cell counting kit-8 assay. ADM and vascular endothelial growth factor in culture medium were detected by enzyme-linked immunosorbent assay. Apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick labeling.
RESULTS AND CONCLUSION: Infection rate of BMSCs increased with multiplicity of infection (MOI) in a dose-dependant manner. When MOI was 150, the infection rate was 95.4%. After BMSCs cultured in hypoxia and serum-free medium, cell growth inhibition and apoptosis increased, but the apoptotic rate in Ad-ADM transfection group was significantly lower than other 2 groups after cultured for 0, 3, 6, 9, 12, 16 hours (P < 0.05). At 9 and 12 hours, BMSCs secreted ADM and vascular endothelial growth factor reached a peak and were significantly higher in the Ad-ADM transfection group than other two groups (P < 0.05). Results have indicated that under hypoxia and serum-free culture conditions (< 20 hours), ADM gene transfection enhances the anti-apoptotic ability of BMSCs, which may be associated with the increased expression of ADM and vascular endothelial growth factor.