Regulation of lncRNA-H19/miR-140-5p in cartilage matrix degradation and calcification in osteoarthritis.

Abstract

To study the effect of long non-coding RNA H19/microRNA (miR)-140-5p regulatory axis on the degradation and calcification of osteoarthritis (OA) cartilage matrix. Twenty cases of clinical OA cartilage tissue and healthy cartilage tissue were collected. Construction of small interfering RNA vector (si-H19) and negative control (si-Control), miR-140-5p mimic and negative control mimic-NC, miR-140-5p inhibitor, and negative control inhibitor-NC transfected human chondrocytes HC-A cells. Chondrocytes were categorized into classes of si-H19, si-Control, mimic, mimic-NC, inhibitor, NC inhibitor, inhibitor + si-H19, NC inhibitor + si-H19. The expression levels of H19, miR-140-5p, alkaline phosphatase (ALP), osteocalcin (OCN), and bone sialoprotein (BSP) in cartilage tissue and chondrocytes were detected using real-time fluorescent quantitative PCR (RT-qPCR). In each group of chondrocytes, Western blotting was used to detect the levels of matrix metalloproteinase (MMP)- 1, MMP-13, and type II α1 collagen fibers (COL2A1). CCK-8 was used to detect the cell proliferation rate, and flow cytometry was used to detect the cell apoptosis rate. H19 was up-regulated in OA cartilage tissue (P<0.05); Silencing H19 not only prevented chondrocyte apoptosis but also facilitated chondrocyte proliferation (both P<0.05), down-regulated MMP1 and MMP-13 expression levels (both P<0.05), and up-regulated COL2A1 expression rates (P<0.05). Also, silenced H19 inhibited the mRNA level of ALP, OCN, and BSP and ALP activity of chondrocytes (all P<0.05). There was a negative correlation between miR-140-5p and H19 in OA cartilage tissue and chondrocytes (r=0.98, P<0.001). Silenced miR-140-5p significantly reversed the inhibitory effects of silenced H19 on matrix degradation and calcification markers in chondrocytes (all P<0.05). The present study showed that the H19/ miR-140-5p regulatory axis could affect the degradation and calcification of the cartilage matrix in OA cells.