LINE-1 methylation levels in synovial fibroblasts of knee osteoarthritis patients

Abstract

Background: Alterations of LINE-1 methylation can cause genomic instability and contribute to dysfunction of the innate immune system, leading to inflammatory disorders like osteoarthritis (OA). However, little work has been completed to determine the change of LINE-1 methylation in synovial fibroblasts (SFs) of patients with OA of the knee.

Objectives: The aim of this study was to examine LINE-1 methylation levels, expression of inflammatory cytokine genes and relative telomere length (RTL) in SFs induced by tumor necrosis factor- (TNF-), hydrogen peroxide (H2O2) and tocopheryl acetate (TA).

Methods: SFs isolated from 4 knee OA patients were treated with 10 ng/mL TNF-, 100 M H₂O₂ and 50 M tocopheryl acetate. LINE-1 methylation levels were assessed by quantitative combined bisulfite restriction analysis. Relative mRNA expression and RTL were measured by quantitative real-time polymerase chain reaction.

Results: LINE-1 methylation levels were significantly decreased in SFs after treatment with TNF- for 24 hours compared with 7 days ( = 0.03). In addition, the relative mRNA expression of IL-1ß, IL-6, MMP-3 and VEGF expression was remarkably higher in SFs treated with TNF- for 3 and 7 days. However, no significant differences in RTL were noted in SFs treated with and without TNF-. In SFs cultured with 10 ng/mL TNF-, 100 M H₂O₂ and pre-treatment with 50 M tocopheryl acetate, LINE-1 methylation levels were found to be reduced, whereas IL-1ß, IL-6, MMP-3 and VEGF mRNA expression were markedly elevated in TNF- stimulated SFs for 24 hours.

Conclusion: These findings suggest that LINE-1 hypomethylation might be implicated in inflammatory response, resulting in up-regulated mRNA expression IL-1ß, IL-6, MMP-3 and VEGF in patients with OA of the knee.