Role of microRNA-27B-3P in the Regulation of Key Extracellular Matrix Markers in the Synovium during Osteoarthritis

Synovial fibrosis, characterized by excessive accumulation of extra-cellular matrix (ECM), is a key pathology contributing to joint destruction associated with Osteoarthritis (OA). microRNAs (miRNAs) are small non-coding RNAs that play key roles in various pathophysiological processes. We have previously shown that miR-27b-3p is abundantly expressed in the OA synovium and elevated in the synovial fluid of patients with late-stage knee OA. However, the exact role of miR-27b-3p in the regulation of ECM in the OA synovium is largely unknown. Hence, my PhD thesis project aimed to determine the role of miR-27b-3p in the regulation of ECM components in the synovium during OA. I first show that miR-27b-3p is prominently expressed in the human OA synovial lining and increased in late-stage knee OA compared to early-stage radiographic knee OA. I further show that miR-27b-3p is highly expressed in the synovial lining and fibrocartilage of a surgery-induced mouse model of knee OA. To decipher the role of miR-27b-3p in the regulation of synovial ECM, I transfected primary human OA fibroblast like synoviocytes (FLS, major synovial cells) with miR-27b-3p-mimic and subjected to ECM-specific qPCR array. Profiling the expression of 84 ECM genes in OA-FLS showed that miR-27b-3p-mimic significantly increases the expression of key ECM genes including pro-collagen-Type-I (COL1A1), A-disintegrin-and-metalloproteinase-with-thrombospondin-motifs-8 (ADAMTS8), Thrombospondin-1 (THBS1), Fibronectin-1 (FN1), Collagen-type-V-alpha-1 (COL5A1) and Collagen-type-XIV-alpha-1 (COL14A1). MiR-27b-3p-mimic also markedly increased COL1A1 protein expression as well as migration of OA-FLS. Integrative computational analysis of identified ECM genes predicted a closed relationship between miR-27b-3p signaling and ADAMTS8. To functionally evaluate this, I silenced ADAMTS8 by siRNA in miR-27b-3p-mimic transfected OA-FLS. Results showed that ADAMTS8 silencing attenuated the increased expression of COL14A1, COL5A1 and FN1 without any significant effect on COL1A1 and THSB1 expression in OA-FLS, suggesting that regulation of COL14A1, COL5A1 and FN1 expression by miR-27b-3p may in part be mediated by ADAMTS8. Finally, I performed RNA sequencing on miR-27b-3p-mimic transfected OA-FLS to profile the transcriptome of miR-27b-3p in FLS during OA. Overall, this study provides a crucial evidence on the role of miR-27b-3p in the regulation of key ECM markers in the OA synovium.Ph.D.2021-06-22 00:00:0