| Objective To observe the effect of Beclin1 wild type and mutant plasmid on the expression of autophagy and apoptosis in osteoblasts. Methods Beclin1 mutant plasmid transfected osteoblast (MUT) and wild-type plasmid transfected osteoblast (WT) were constructed from rat skull osteoblasts isolated and cultured in primary culture. Cells were treated with autophagy activator rapamycin (Rapa), autophagy inhibitor BAF-A1 (Baf) and Beclin1/Bcl2 binding inhibitor mifepristone (Mif), respectively. Cells were divided into Beclin1 WT group and Beclin1 WT+mif group, Beclin1 WT+ Baf-A1 group, Beclin1 Mut group, and Beclin1 Mut+Rapa group. The Beclin1-BCL2 complex binding level was detected with immunoprecipitation. The autophagy and apoptosis of osteoblasts were detected with MDC, lysosome, flow detection, mitochondrial membrane potential, Hoechest, immunofluorescence, and WB, respectively. Results Immunoprecipitation detection: compared to that in NC group, Beclin1-Bcl2 complex level in MUT group significantly (p<0.05). MDC staining showed that compared to that in blank group, lysosome staining in WT cells increased significantly. Apoptosis: compared to those in NC group and WT group, the apoptosis rate in WT+Mif group increased significantly. The apoptosis rate in MUT cells was lower than that in WT cells, and the results were statistically different. JC-1 detection showed that compared to that in NC group, the mitochondrial transmembrane potential of WT cells changed abnormally, while the expression of transmembrane potential of MUT cell samples was active. Conclusions (1) Beclin1 overexpression inhibits the formation of Beclin1-BCl2 complex. (2) Beclin1 is an important gene involved in autophagy. Moderate expression of Beclin1 is conducive to maintaining the metabolic balance of cell population. Beclin1 overexpression has no significant effect on autophagy. (3) Beclin1 overexpression inhibits osteoblast apoptosis. |