LncRNA TM1-3P Regulates Proliferation, Apoptosis and Inflammation of Fibroblasts in Osteoarthritis through miR-144-3p/ONECUT2 Axis

Objective This study explores LncRNA TM1-3P effects on the proliferation, apoptosis, and inflammatory response of fibroblasts in osteoarthritis (OA) and its underlying mechanism. Methods Bioinformatics was performed to analyze OA disease-related genes, miRNA profiles, and function. The targeted regulation of LncRNA TM1-3P and miR-144-3p, ONECUT2 and miR-144-3p were analyzed by dual luciferase reporter gene assay, RNA Binding Protein Immunoprecipitation (RIP), and RNA pull down. Histopathological morphology of the knee joint was observed by hematoxylin-eosin (HE) and Annona Red O/Fast Green. The expressions of mRNAs and proteins were detected by RT-qPCR, Western blot, and immunohistochemistry. Unpaired T test was used between groups, and the one-way analysis of variance of repeated measurement data was applied for multi-group comparison, following Tukey's post-test. Results ONECUT2 and Smurf2 genes were significantly elevated in the osteoarthritis group compared with the normal group (P < 0.001, P < 0.001). Expressions of ONECUT2 and LncRNA TM1-3P were increased, and expression of miR-144-3p was decreased in interleukin (IL)-1 beta-induced human fibroblast synovial cells (hFSCs) (mRNA: 1.06 +/- 0.24 vs. 3.29 +/- 0.73, proteins: 0.22 +/- 0.03 vs. 0.46 +/- 0.22, 1.23 +/- 0.22 vs. 3.76 +/- 0.73, 1.06 +/- 0.25 vs. 0.37 +/- 0.13, P < 0.01, P < 0.001, P < 0.01, P < 0.05). Overexpression of miR-144-3p down-regulated the ONECUT2 expression, reduced cell proliferation, promoted apoptosis in hFSCs induced by IL-1 beta (mRNA: 0.89 +/- 0.14 vs. 0.15 +/- 0.01, P < 0.05; proteins: 0.46 +/- 0.01 vs. 0.23 +/- 0.01, P < 0.001; CCK8: 1.88 +/- 0.07 vs. 1.65 +/- 0.07; P < 0.05; EDU: 55.82 +/- 1.44 vs 40.57 +/- 2.24, P < 0.05; apoptosis: 10.57 +/- 0.79 vs 16.36 +/- 0.35, P < 0.0001). Overexpression of LncRNA TM1-3P up-regulated the expression of ONECUT2, promoted cell proliferation, and inhibited apoptosis (mRNA: 0.9 +/- 0.09 vs 1.94 +/- 0.12, P < 0.05; proteins: 0.61 +/- 0.05 vs 0.76 +/- 0.03, P > 0.05; CCK8: 2.07 +/- 0.05 vs 2.47 +/- 0.06; P < 0.01; EDU: 52.67 +/- 1.17 vs 60.06 +/- 3.24, P < 0.05; apoptosis: 10.57 +/- 0.79 vs 16.36 +/- 0.35, P < 0.001), which were reversed by the overexpression of miR-144-3p treatment (mRNA: 1.82 +/- 0.07 vs 0.31 +/- 0.07, P < 0.0001; proteins: 0.74 +/- 0.02 vs 0.35 +/- 0.01, P < 0.01; CCK8: 2.41 +/- 0.01 vs 1.67 +/- 0.02; P < 0.0001; EDU: 66.85 +/- 2.86 vs 44.68 +/- 1.97, P < 0.0001; apoptosis: 7.19 +/- 0.19 vs 13.36 +/- 0.53, P < 0.0001). Silencing LncRNA TM1-3P attenuated the injury of knee joint tissue, down-regulated the expression of ONECUT2, Smurf2, IL-1 beta, IL-6, TNF-alpha, and improved the expression of Rap1 in rats (0.71 +/- 0.04 vs 0.48 +/- 0.02, 0.68 +/- 0.06 vs 0.36 +/- 0.02, 0.74 +/- 0.03 vs 0.49 +/- 0.04, 0.78 +/- 0.01 vs 0.54 +/- 0.03, 0.68 +/- 0.02 vs 0.4 +/- 0.04, 0.24 +/- 0.01 vs 0.4 +/- 0.03, P < 0.05, P < 0.05, P < 0.05, P < 0.01, P < 0.01, P < 0.05). Conclusion LncRNA TM1-3P improved inflammation and damage of knee joints in OA rats through miR-144-3p/ONECUT2 axis, providing a new theoretical basis for gene therapy of OA.