Metformin alleviates dexamethasone-induced apoptosis by regulating autophagy via AMPK/mTOR/p70S6K in osteoblasts

Abstract

Glucocorticoid (GC)-induced osteoporosis (GIOP) is the most common type of secondary osteoporosis. Osteoblast apoptosis induced by GCs is now considered as a crucial factor for GIOP. Many clinical, in vivo, and in vitro studies have shown that metformin has a beneficial effect on bone metabolism and bone formation. To investigate whether metformin could be used to treat GIOP, we explored the influence of metformin on dexamethasone (Dex)-induced apoptosis of osteoblasts and its underlying mechanisms. In this study, the CCK8 assay was used to determine the optimal metformin concentration and processing time. The expression levels of target proteins were examined by Western blot and immunofluorescence; the expression levels of target genes were tested by quantitative PCR. Apoptotic cells were detected using flow cytometry. Characteristics of autophagy were observed by transmission electron microscopy. An autophagy inhibitor was administered to investigate whether autophagy decreases apoptosis. Sh-AMPK transfection and an mTOR activator were used to investigate the role of AMPK/mTOR signaling in metformin-induced autophagy. The results showed that metformin alleviated Dex-induced apoptosis of osteoblasts accompanied by increased autophagy. Treatment with the autophagy inhibitor 3-methyladenine (3-MA) attenuated the effect of metformin on apoptosis, autophagy, and the AMPK/mTOR/p70S6K signaling pathway. The anti-apoptotic effect of metformin on osteoblasts is associated with the promotion of autophagy. Furthermore, sh-AMPK transfection and the mTOR activator MHY1485 impaired metformin-mediated inhibition of osteoblast apoptosis and promotion of autophagy. The AMPK/mTOR/p70S6K signaling pathway plays a role in metformin-mediated apoptosis suppression and autophagy promotion. In conclusion, metformin can alleviate Dex-induced osteoblast apoptosis by inducing autophagy via the AMPK/mTOR/p70S6K pathway. This study highlights the potential value of metformin in the treatment of GIOP.