SIRT3-mediated DRP1 acetylation changes are associated with the restoration of mitochondrial function and alleviation of anterior cruciate ligament injury-induced osteoarthritis.

Mitochondrial dysfunction and endoplasmic reticulum stress (ERS) play critical roles in the development of osteoarthritis (OA). Sirtuin 3 (SIRT3), a mitochondrial deacetylase, has regulatory effects on OA, but whether it can influence OA progression by modulating the crosstalk between mitochondrial dysfunction and ERS remains unclear. Therefore, the aim of this study was to investigate the effects of SIRT3 on mitochondrial dysfunction and ERS in OA. Anterior cruciate ligament transection was performed to establish an OA rat model, and C28/I2 chondrocytes were treated with IL-1β to establish an in vitro OA cell model for experimental investigation. Hematoxylin and eosin (H&E) staining, Safranin O/Fast Green staining, MitoTracker Red staining, and JC-1 staining were used to assess OA progression in rats and C28/I2 cell damage. RT-qPCR and Western blotting were used to measure the expression levels of target genes and proteins. Coimmunoprecipitation was utilized to examine protein interactions and acetylation levels. The result shows that SIRT3 expression was downregulated in OA rats and IL-1β-induced C28/I2 cells. SIRT3 overexpression significantly alleviated pathological damage to rat articular cartilage tissues, reduced chondrocyte loss, improved matrix metabolism in articular cartilage tissues, and ultimately mitigated OA progression in rats. Moreover, SIRT3 overexpression inhibited IL-1β-induced ERS in C28/I2 cells, reduced the activation of the unfolded protein response, and improved mitochondrial function. Additionally, the mitochondrial Ca2+ transport inhibitor Ru360 suppressed mitochondrial Ca2+ overload caused by disrupted endoplasmic reticulum calcium homeostasis and improved both mitochondrial function and ERS. DRP1 was upregulated in OA rats and IL-1β-stimulated cells, and changes in DRP1 acetylation signals together with decreased DRP1 protein abundance were observed following SIRT3 manipulation. Overall, SIRT3 manipulation was associated with acetylation signals of DRP1 and reduced DRP1 protein abundance, indicating that DRP1 is a potential mediator involved in the SIRT3‑linked restoration of mitochondrial function, attenuation of ERS, and alleviation of OA progression.