L. Wei (Providence, US)
Warren Alpert Medical School of Brown University Department of OrthopedicsPresenter Of 1 Presentation
12.2.8 - MicroRNA-195 contributes development of osteoarthritis via targeting Smad3
Abstract
Purpose
This study investigated the biological function of microRNA-195(miR-195) and the molecular mechanism in osteoarthritis(OA).
Methods and Materials
The knee cartilage tissues obtained from traumatic amputees (n=10) and OA patients (n=10). Real-time PCR and Western blot were used to detect the expression of miR-195 and Smad3 from cartilage. Human chondrocytes were transfected with the miR-195 mimic, the miR-195 inhibitor, or the control, respectively. The expression level of Smad3 , type II collagen (Col II) were detected by Real-time PCR 24h later or by Western blot 48h later. We constructed a Smad3 3'-UTR-luciferase reporter and examined the effect of miR-195 on Smad3 3'-UTR luciferase activity by dual luciferase reporter gene assay in 293T cells. To further understand whether Smad3 mediates the downstream effects of miR-195, human chondrocytes were transfected with the miR-195 inhibitor with and without siRNA-Smad3 (siSmad3). The study was approved by the Ethics Committee of The Second Hospital of ShanXi Medical University.
Results
miR-195 levels were significantly upregulated in OA tissue (P<0.05) while Smad3 mRNA/protein expression was substantially decreased in OA tissue (P<0.05). Consistently, upregulated miR-195 strongly inhibited Col II expression, while downregulated miR-195 significantly increased Col II expression (P<0.05). Furthermore, it has been demonstrated that Smad3 is a novel target of miR-195 using dual luciferase reporter assay. Most importantly, miR-195 mediated the changes of Col II were substantially attenuated by siSmad3 treatment(P<0.05).
Conclusion
Our study identifies that miR-195 is up-regulated in OA cartilage, while the content of Smad3 is decreased in OA cartilage. Smad3 is a novel target of miR-195. We further demonstrated that Smad3 is involved in the progress of miR-195 mediated OA cartilage degeneration.