Effects of Bushen Jianpi Huoxue Recipe on cell proliferation and alkaline phosphatase activity in Beclin-1 overexpressing and silencing MC-3T3-E1 cells

doi:10.12307/2021.164

Abstract

Abstract: BACKGROUND: Osteoblasts are one of the important cells involved in the fine regulation of the dynamic balance of bone metabolism, and their dysfunction is extremely essential for the pathogenesis of osteoporosis. Studies have found that knockout of Beclin-1, an autophagy gene, leads to a decrease in osteoblast differentiation and mineralization, suggesting that Beclin-1 has an important regulatory role in bone metabolism.
OBJECTIVE: To explore the effects of Bushen Jianpi Huoxue Recipe on cell proliferation and alkaline phosphatase activity in Beclin-1 overexpressing and silencing MC-3T3-E1 cells
METHODS: Recombinant lentiviral vectors overexpression and silencing Beclin-1 were constructed, and drug-containing serum of Bushen Jianpi Huoxue Recipe was prepared. MC-3T3-E1 cells were divided into six groups: blank serum + blank virus group, blank serum + Beclin-1 overexpression group, blank serum + Beclin-1 silence group, drug serum + blank virus group, drug serum + Beclin-1 overexpression group, AND drug serum + Beclin-1 silence group. Interventions in each group were accordingly carried out. Cell proliferation was detected using cell counting kit-8, cell cycle was determined using flow cytometry, and alkaline phosphatase activity was measured using an alkaline phosphatase kit.
RESULTS AND CONCLUSION: Compared with the blank serum + blank virus group, the cell proliferation and alkaline phosphatase activity were significantly increased in the drug serum + blank virus group, blank serum + Beclin-1 overexpression group, and drug serum + Beclin-1 overexpression group, and significantly decreased in the blank serum + Beclin-1 silence group and drug serum + Beclin-1 silence group. The most obvious changes were found in the drug serum + Beclin-1 overexpression group and blank serum + Beclin-1 silence group. From the perspective of cell cycle in each group, the proportion of cells in G2/M phase was relatively higher in the blank serum + Beclin-1 overexpression group and drug serum + Beclin-1 overexpression group; the proportion of cells in G0/G1 phase was relatively higher in the blank serum + blank virus group, blank serum + Beclin-1 silence group and drug serum + Beclin-1 silence group; the proportion of cells in S phase was relatively higher in the blank serum + Beclin-1 overexpression group and drug serum + blank virus group. To conclude, overexpression of Beclin-1 and serum containing Bushen Jianpi Huoxue Recipe, especially their combination, can improve the proliferation and alkaline phosphatase activity of MC-3T3-E1 cells. Whereas, silence of Beclin-1 can reduce the proliferation and alkaline phosphatase activity of MC-3T3-E1 cells, and serum containing Bushen Jianpi Huoxue Recipe can inhibit the adverse effects of Beclin-1 silence to some extent.

Key words: MC-3T3-E1 cells, lentivirus, transfection, Beclin-1, autophagy, traditional Chinese medicine, alkaline phosphatase

CLC Number:

Zheng Weipeng, Hu Weijian, Zhao Guoyuan, Wei Hewei, Liu Zhijun, Wan Lei, Chen Sheng, Liao Zhihao. Effects of Bushen Jianpi Huoxue Recipe on cell proliferation and alkaline phosphatase activity in Beclin-1 overexpressing and silencing MC-3T3-E1 cells[J]. Chinese Journal of Tissue Engineering Research, 2021, 25(29): 4650-4655.

Figures/Tables 6

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