The Effect of HOXA10 Gene on Proliferation and Differentiation of Bovine Skeletal Muscle Satellite Cells
ZHANG Meng, ZHANG Wentian, YANG Li, LIU Lili, LÜ Ruixue, FU Xuepeng, ZHANG Weiwei*
This study explored the effects of HOXA10 gene on the proliferation and differentiation of BSMSCs (bovine skeletal muscle satellite cells). qRT-PCR (real-time quantitative PCR) was used to detect the ex pression of HOXA10 and PCNA genes in BSMSCs proliferating for 1, 2 and 3 days (P1, P2 and P3). HOXA10 gene overexpression plasmids, interference plasmids, and control plasmids were transfected into BSMSCs. qRT-PCR and Western blot were used to detect overexpression and interference effects. CCK-8, EdU, FCM (Flow CytoM etry), immunofluorescence, qRT-PCR and Western blot were used to detect cell viability, cell proliferation, cell cycle, cell differentiation, mRNA and protein expression of genes that mark cell proliferation and differentiation. The results showed that the mRNA of HOXA10 was significantly increased during the proliferation of BSMSCs (P<0.01, P<0.001). Compared with the control group, the overexpression and interference effects were signifi cant after 24 hours of transfection (P<0.01, P<0.001); the cell viability and the proportion of EdU-positive cells in BSMSCs were increased (P<0.001) after overexpression of HOXA10 gene, meanwhile the percentage of cells in G1 phase was significantly decreased (P<0.01); the percentage of cells in S phase was significantly increased (P<0.05); and the expression levels of proliferation marker genes PCNA, CCND1 mRNA and protein were sig nificantly increased (P<0.05, P<0.01). The formation of myotubes was significantly reduced, and the mRNA and protein expressions of the differentiation marker gene MyoG were significantly reduced (P<0.01, P<0.001). Compared with the control group, after silencing of the HOXA10 gene, the cell viability decreased, the propor tion of EdU-positive cells decreased (P<0.05); the percentage of cells in G1 phase increased (P<0.001); the per centage of cells in S phase decreased significantly (P<0.01). The levles of PCNA, CCND1 mRNA and protein were decreased significantly after silencing of the HOXA10 gene (P<0.05, P<0.001). The formation of myotubes was significantly increased, and the mRNA and protein expressions of the differentiation marker gene MyoG were significantly increased (P<0.001). In conclusion, HOXA10 gene promotes the proliferation and inhibits dif ferentiation of BSMSCs.
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