A-485 alleviates postmenopausal osteoporosis by activating GLUD1 deacetylation through the SENP1-Sirt3 signal pathway.

OBJECTIVE: Postmenopausal osteoporosis (OP) is a bone disease caused by estrogen deficiency. A-485 is a selective inhibitor of p300/CBP histone acetyltransferase (HAT) with potential regulatory effects on bone remodeling. This study aims to investigate the effects of A-485 on postmenopausal OP and its underlying mechanisms. METHODS: For animal experiments, 61 female Wistar rats were used to establish an OP model through ovariectomy (OVX). The rats were administered with A-485 (100 mg/kg/day) via intraperitoneal injection for six weeks. Bone mineral density (BMD) was measured using dual-energy X-ray absorptiometry (DXA). Histopathological changes were observed using HE and Masson's trichrome staining. ELISA was used to measure bone resorption markers (CTX-1, DPD) and the bone formation marker (P1NP) in rats. Osteoblast differentiation markers (Runx2, OCN), SENP1, Sirt3 expression levels, and GLUD1 acetylation were assessed via Western blot (WB) and RT-qPCR. In vitro, MC3T3-E1 osteogenic progenitor cells were cultured in osteogenic differentiation medium supplemented with ascorbic acid, β-glycerophosphate, dexamethasone, and fulvestrant. CCK-8 was performed to evaluate cell proliferation. Flow cytometry was selected to measure apoptosis and mitochondrial membrane potential. WB and RT-qPCR were employed to analyze ERα, ERβ, Runx2, Sirt3, and GLUD1 acetylation. Additionally, Alizarin red staining was applied to monitor osteoblast mineralization. ATP levels were detected using a commercial kit, and ROS levels were measured by MitoSOX Red. RESULTS: In vivo, ovariectomized rats exhibited lower BMD, impaired bone trabeculae, increased CTX-1 and DPD, and altered expression of Runx2 and OCN, all of which were reversed by A-485 treatment. In vitro, A-485 activated GLUD1 deacetylation, enhanced osteogenic differentiation, and improved mitochondrial function. Regarding the mechanism, A-485 activated the SENP1-Sirt3 signal pathway, with SENP1 knockdown negating the effects of A-485. In vivo, A-485 reduced GLUD1 acetylation and promoted improvement of OP, which were reversed by SENP1 knockdown. CONCLUSION: A-485 ameliorates postmenopausal OP by activating GLUD1 deacetylation via the SENP1-Sirt3 signal pathway, thus improving mitochondrial function, and promoting osteogenic differentiation and mineralization.