| Objective:To investigate the effect and mechanism of negative co-stimulatory molecule B7-H4 on the proliferation and survival of mouse mesenchymal stem cell line C3H10 T1/2(hereinafter referred to as C3H10).Methods:C3H10-shRNA-B7H4-1,C3H10-shRNA-B7H4-2,and C3H10-shRNA-B7H4-NC cell lines transfected with no load virus were constructed using RNA interference technology.The C3H10-shRNA-B7H4-1 group with significantly down-regulated B7-H4 expression was selected as the experimental group,the C3H10-NC group as the control group,and the C3H10 group as the blank control group for follow-up experiments.Cell proliferation in each group was detected by CCK-8 method,cell counting method and Westepn blot.Cell cycle and apoptosis were detected by FCM.The expressions of Cyclin DI and cyclin-2,BAX,caspase-3 and caspase-9 in each group were detected by Western blot.Results:FACS,Western blot and immunofluorescence staining showed that B7-H4 was highly expressed in C3H10,and its expression rate was significantly reduced after B7-H4 targeted shRNA interference with C3H10(P<0.05).(2)Compared with C3H10-NC,the proliferation capacity of C3H10-shRNA-B7H4-1 was significantly decreased(P<0.05).(3)In the C3H10-shRNA-B7H4-1 group,the G0/G1 phase was significantly increased(P<0.05),and the apoptosis was significantly increased in the early phase(P<0.05),while there was no significant difference in the late phase(P=0.717).(4)In the C3-H10-shRNA-B7H4-1 group,Cyclin D1 was down-regulated,BAX,caspase-3 and caspase-9 were up-regulated,and bcl-2 was down-regulated.Conclusion:Negative co-stimulatory molecule b7-h4 is highly expressed in mouse mesenchymal stem cell C3H10 T1/2,and is closely related to the proliferation and survival of the cells.The mechanism may be related to the involvement of B7-H4 in cell cycle regulation and the prevention of apoptosis. |
