Zuogui Pills down-regulating autophagy to inhibit osteoclast differentiation in the treatment of senile osteoporosis
Objective:To explore the effects of Zuogui Pills(ZGW)on senile osteoporosis(SOP)by down-regulating autophagy and inhibiting bone marrow-derived macrophage(BMMs)osteoclast differentiation in mice.Methods:The mouse model of senile osteoporosis was established and 20 mice were randomly divided into two groups:aging group(SOP group)and Zuogui Pills treatment group(ZGW group).After 2 months of treatment with Zuogui Pills,bone volume fraction(BV/TV)was measured by Micro-CT,the number of osteoclasts on the trabecular surface was detected by TRAP staining,and the expression of P-BECLIN1 in bone tissue was detected by immunohistochemistry(IHC).Mouse BMMs was extracted,and different concentrations of Zuogui Pills were used to interfere with the osteoclast(OCs)differentiation process of BMMs.The number of OCs was observed by TRAP staining,the mRNA expression of autophagy marker genes Atg5,Atg12,Lc3 and p62 was detected by RT-qPCR,and the protein expression levels of NFATC1,CTSK,LC3,P62 and P-BECLIN1 in cells were detected by Western Blot.Results:In vivo experiment,BV/TV in ZGW group was significantly higher than that in SOP group(P＜0.01),the number of OCs on trabecular surface in ZGW group was significantly less than that in SOP group(P＜0.01),and the expression of P-BECLIN1 in bone marrow cavity of ZGW group was significantly lower than that of SOP group(P＜0.01).In vitro experiment,compared with the control group,10,100,1 000 μg/mL Zuogui Pills could inhibit the OCs differentiation of BMMs induced by RANKL(P＜0.01).During the inhibition of OCs differentiation,the mRNA expression of autophagy genes Atg5,Atg12 and LC3 was decreased(P＜0.05,P＜0.01),while the mRNA expression of P62 was increased(P＜0.01).Western Blot suggested that Zuogui Pills could inhibit the protein expression of NFATC1,CTSK,LC3 and P-BECLIN1 and up-regulate the protein expression of P62.Conclusion:Zuogui Pills can down-regulate autophagy and inhibit OCs formation,which can treat senile osteoporosis.Zuogui Pills is a potential drug for the treatment of SOP and provides a new idea for clinical treatment of SOP.
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