Diabetes-accelerated experimental osteoarthritis is prevented by MTOR inhibition

Abstract

Purpose: Type 2 Diabetes (T2D) is an important risk factor for Osteoarthritis (OA). Autophagy, an essential homeostatic mechanism in articular cartilage, is defective in T2D and OA, while mTOR inhibition has been shown to be protective in experimental OA. However, how T2D may influence OA disease progression and the molecular mechanisms associated is still unknown. The objective of this study was to determine how diabetes affects cartilage integrity and whether pharmacological activation of autophagy has disease-modifying efficacy in experimental-induced OA in diabetic mice. Methods: To determine if diabetes influences cartilage damage, experimental OA was performed in the right knee of 9 weeks-old C57Bl/6 male mice (Lean group, N = 8) and of 9 weeks-old B6.BKS (D)-Leprdb male mice (db/db group, N = 16) by transection of the medial meniscotibial ligament and the medial collateral ligament. Left knee was employed as a control knee. Db/db mice were randomized according to body weight and blood glucose levels in two groups of N = 8. Rapamycin (2 mg/kg weight/day) or vehicle (dimethyl sulphoxide, DMSO) were administered intraperitoneally 3 times a week for 10 weeks. Body weight and fasting blood glucose levels were monitorized weekly. Insulin levels in plasma were determined by ELISA at the end of the study. Histopathological changes in articular cartilage and synovium were evaluated by using a semiquantitative scoring system and a synovitis grading system, respectively. Western Blotting and Inmunohistochemical staining were employed to analyze the effect of diabetes and Rapamycin treatment on cartilage integrity, OA biomarkers and mTOR signaling pathway. Results: Diabetic mice have increased body weight, fasting blood glucose levels and insulin levels compared to lean mice. Importantly, Rapamycin treated mice have significantly reduced body weight and blood glucose and insulin compared to control db/db mice. The results indicated that cartilage damage was significantly increased in db/db mice compared to lean group after surgical induction of OA, while no differences are observed in the control knee. To evaluate whether mTOR inhibition protects against surgical-induced OA in db/db mice, Rapamycin was administered. The data showed a significant reduction in OA severity in the treated group. This reduction in cartilage damage was accompanied with a significant reduction in synovium inflammation and in the expression of OA biomarkers, such as Metalloproteinase-13 (MMP-13) and Aggrecanase-5 (ADAMTS-5). Furthermore, cartilage cellularity was maintained after Rapamycin treatment, suggesting that mTOR inhibition protects chondrocytes against death. Conclusions: Our findings indicate, for the first time, that diabetic mice exhibit accelerated- cartilage damage after experimental OA. Importantly, mTOR inhibition by Rapamycin reduces body weight, corrects metabolic parameters, and reduces cartilage damage and synovium inflammation. Furthermore, Rapamycin reduces the expression of OA biomarkers and maintains chondrocyte homeostasis. These results demonstrate that pharmacological inhibition of mTOR by Rapamycin is an effective therapy for diabetes-accelerated Osteoarthritis.