Metformin regulates chondrocyte senescence and proliferation through microRNA-34a/SIRT1 pathway in osteoarthritis.

Osteoarthritis (OA) is the most common degenerative disease in joints among elderly patients. Senescence is deeply involved in the pathogenesis of osteoarthritis. Metformin is widely used as the first-line drug for Type 2 diabetes mellitus (T2DM), and has great potential for the treatment of other aging-related disorders, including OA. However, the role of metformin in OA is not fully elucidated. Therefore, our aim here was to investigate the effects of metformin on human chondrocytes. After metformin treatment, expression level of microRNA-34a and SIRT1 in chondrocyte were detected with quantitative real-time PCR and immunofluorescence staining. Then, microRNA-34a mimic and small interfering RNA (siRNA) against SIRT1 (siRNA-SIRT1) were transfected into chondrocyte. Senescence-associated β-galactosidase (SA-β-gal) staining was performed to assess chondrocyte senescence. Chondrocyte viability was illustrated with MTT and colony formation assays. Western blot was conducted to detect the expression of P16, IL-6, matrix metalloproteinase-13 (MMP-13), Collagen type II (COL2A1) and Aggrecan (ACAN). We found that metformin treatment (1 mM) inhibited microRNA-34a while promoted SIRT1 expression in OA chondrocytes. Both miR-34a mimics and siRNA against SIRT1 inhibited SIRT1 expression in chondrocytes. SA-β-gal staining assay confirmed that metformin reduced SA-β-gal-positive rate of chondrocytes, while transfection with miR-34a mimics or siRNA-SIRT1 reversed it. MTT assay and colony formation assay showed that metformin accelerated chondrocyte proliferation, while miR-34a mimics or siRNA-SIRT1 weakened this effect. Furthermore, results from western blot demonstrated that metformin suppressed expression of senescence-associated protein P16, proinflammatory cytokine IL-6 and catabolic gene MMP-13 while elevated expression of anabolic proteins such as Collagen type II and Aggrecan, which could be attenuated by transfection with miR-34a mimics. Overall, our data suggest that metformin regulates chondrocyte senescence and proliferation through microRNA-34a/SIRT1 pathway, indicating it could be a novel strategy for OA treatment.

Yan S ，Dong W ，Li Z ，Wei J ，Han T ，Wang J ，Lin F ... -  《Journal of Orthopaedic Surgery and Research》

MicroRNA-34a affects chondrocyte apoptosis and proliferation by targeting the SIRT1/p53 signaling pathway during the pathogenesis of osteoarthritis.

Yan S ，Wang M ，Zhao J ，Zhang H ，Zhou C ，Jin L ，Zhang Y ，Qiu X ，Ma B ，Fan Q ... -  《-》

MiR-29a and MiR-140 Protect Chondrocytes against the Anti-Proliferation and Cell Matrix Signaling Changes by IL-1β.

Li X ，Zhen Z ，Tang G ，Zheng C ，Yang G ... -  《-》

Downregulation of microRNA-448 inhibits IL-1β-induced cartilage degradation in human chondrocytes via upregulation of matrilin-3.

Yang H ，Wu D ，Li H ，Chen N ，Shang Y ... -  《-》

Circ_0022383 alleviates IL-1β-induced apoptosis, inflammation and extracellular matrix degeneration in osteoarthritis cell model by miR-3619-5p/SIRT1 axis.

Circular RNAs (circRNAs) have been identified to play roles in cartilage homeostasis and chondrocyte development, and be associated with osteoarthritis (OA) pathophysiology. Here, we aimed to investigate the role and mechanism of circ_0022383 on OA progression. Chondrocytes in functional groups were treated with interleukin (IL)-1β. The levels of genes and proteins were assayed by qRT-PCR and western blotting. Cell proliferation and apoptosis were evaluated by cell counting kit-8 assay, 5-Ethynyl-2'-deoxyuridine (Edu) assay, and flow cytometry, respectively. The inflammation and extracellular matrix (ECM) degeneration were determined by assessing the activity of IL-6, tumor necrosis factor (TNF-α), Aggrecan (ACAN), collagen type II α 1 chain (COL2A1) and ADAMTS5 proteins. The binding between miR-3619-5p and circ_0022383 or silent information regulator 1 (SIRT1) was confirmed by dual-luciferase reporter, RIP and RNA pull-down assays. Circ_0022383 expression was lower in the cartilages of OA patients and IL-1β-induced primary chondrocytes. Functionally, ectopic overexpression of circ_0022383 alleviated IL-1β-induced proliferation arrest, apoptosis, the release of IL-6 and TNF-α, as well as the decrease of ACAN and COL2A1 and the increase of ADAMTS5 level in chondrocytes. Mechanistically, circ_0022383 acted as a sponge for miR-3619-5p, which was verified to target SIRT1. Rescue experiments showed that miR-3619-5p up-regulation reversed the protective effects of circ_0022383 on IL-1β-stimulated chondrocytes. Additionally, miR-3619-5p inhibition abolished IL-1β-induced apoptosis, inflammation and ECM degeneration in chondrocytes, which were counteracted by SIRT1 silencing. Circ_0022383 protected chondrocytes from IL-1β-induced apoptosis, inflammation and ECM degeneration by miR-3619-5p/SIRT1 axis, inspiring future therapy development for OA prevention.

Qian L ，Yu B ，Chen T ，Chen K ，Ma Z ，Wang Y ，Sun B ... -  《-》