Metformin attenuates osteoclast-mediated abnormal subchondral bone remodeling and alleviates osteoarthritis via AMPK/NF-κB/ERK signaling pathway

Abstract

This study explored the mechanism by which metformin (Met) inhibits osteoclast activation and determined its effects on osteoarthritis (OA) mice. Bone marrow-derived macrophages were isolated. Osteoclastogenesis was detected using tartrate-resistant acid phosphatase (TRAP) staining. Cell proliferation was evaluated using CCK-8, F-actin rings were detected by immunofluorescence staining, and bone resorption was detected using bone slices. Nuclear factor kappa-B (NF-κB) and nuclear factor of activated T-cell cytoplasmic 1 (NFATc1) were detected using luciferase assays, and the adenosine monophosphate-activated protein kinase (AMPK), NF-κB, and mitogen-activated protein kinase (MAPK) signaling pathways were detected using western blotting. Finally, expression of genes involved in osteoclastogenesis was measured using quantitative polymerase chain reaction. A knee OA mouse model was established by destabilization of the medial meniscus (DMM). Male C57BL/6J mice were assigned to sham-operated, DMM+vehicle, and DMM+Met groups. Met (100 mg/kg/d) or vehicle was administered from the first day postoperative until sacrifice. At 4- and 8-week post OA induction, micro-computed tomography was performed to analyze microstructural changes in the subchondral bone, hematoxylin and eosin staining and Safranin-O/Fast Green staining were performed to evaluate the degenerated cartilage, TRAP-stained osteoclasts were enumerated, and receptor activator of nuclear factor κB ligand (RANKL), AMPK, and NF-κB were detected using immunohistochemistry. BMM proliferation was not affected by Met treatment below 2 mM. Met inhibited osteoclast formation and bone resorption in a dose-dependent manner in vitro. Met suppressed RANKL-induced activation of p-AMPK, NF-κB, phosphorylated extracellular regulated protein kinases (p-ERK) and up-regulation of genes involved in osteoclastogenesis. Met reversed decreases in BV/TV, Tb.Th, Tb.N, and CD, and an increase in Tb.Sp at 4 weeks postoperatively. The number of osteoclasts and OARSI score were decreased by Met without effect on body weight or blood glucose levels. Met inhibited RANKL, p-AMPK, and NF-κB expression in early OA. The mechanism by which Met inhibits osteoclast activation may be associated with AMPK/NF-κB/ERK signaling pathway, indicating a novel strategy for OA treatment.

Metformin alleviates osteoarthritis in mice by inhibiting chondrocyte ferroptosis and improving subchondral osteosclerosis and angiogenesis

“…In order to establish a mouse OA model, the right knee of mice was selected for DMM surgery [ 36 ]. Briefly, after mice were anesthetized with an intraperitoneal injection of 1% sodium pentobarbital (60 mg/kg) [ 37 ], the meniscotibial ligament of the right knee was transected to free the anterior crus of the meniscus, and sham surgery was performed without transecting the meniscotibial ligament. Mice were injected intra-articularly with Erastin (15 mg/kg) [ 38 ], and the sham and DMM groups were injected with an equal volume of saline twice a week.…”

Autophagy attenuates osteoarthritis in mice by inhibiting chondrocyte pyroptosis and improving subchondral bone remodeling

“…Li et al [60] found that OPG levels were decreased, and RANKL levels were increased in subchondral bone of early OA in vivo. According to our previous study, the RANK/RANKL/OPG axis played a key role in the process of osteoclast activation in the subchondral bone of early OA [61,62].…”

How zoledronic acid improves osteoporosis by acting on osteoclasts

“…A study has shown that AMP-activated protein kinase (AMPK) plays a negative regulatory role in osteoclast differentiation and function ( Tong et al, 2020 ). Multiple drugs can affect osteoclast differentiation and bone resorption through AMPK-mediated signaling pathways, such as AMPK/NF-κB/ERK, AMPK/mTOR/p70S6K, and AMPK/Nfr2 signaling pathways ( Li Z et al, 2018 ; Tong et al, 2018 ; Peng et al, 2019 ; Zhao et al, 2020 ; Guo et al, 2021 ). Unlike various other drugs whose effects on osteoclast differentiation and bone resorption through AMPK pathway have been widely studied, studies on how ZA affects osteoclasts through the AMPK pathway are relatively recent.…”