Effects of Tim-3 silencing on the viability of fibroblast-like synoviocytes and lipopolysaccharide-induced inflammatory reactions.

Abstract

The objective of the present study was to investigate the effects of Tim-3 silencing on cell viability and lipopolysaccharide (LPS)-induced inflammatory reactions in fibroblast-like synoviocytes (FLS). T-cell immunoglobulin mucin domain molecule (Tim)-3 expression in FLS obtained from patients with rheumatoid arthritis (RA) and normal controls were detected by western blot analysis and reverse transcription-polymerase chain reaction (RT-PCR). Small interfering (si)RNA was transfected using Lipofectamine® 2000 to decrease Tim-3 expression. Following transfection, FLS were stimulated by LPS. An MTT assay, RT-PCR and western blot analysis were performed to measure cell viability, Toll-like receptor 4 (TLR4) signaling pathway-related protein expression and inflammatory cytokine release, respectively. The results of the present study indicated that Tim-3 expression was increased in FLS from patients with RA compared with FLS from healthy controls. Transfection of Tim-3 siRNA significantly decreased Tim-3 expression in FLS from patients with RA. Notably, Tim-3 silencing decreased FLS cell viability. Following stimulation with LPS, cell viability and the expression of TLR4, myeloid differentiation protein gene 88 (MyD88) and nuclear factor-κB (NF-κB) p65 were enhanced in FLS. By contrast, Tim-3 silencing attenuated LPS-induced cell proliferation and the expression of TLR4, MyD88 and NF-κB p65. In addition, LPS significantly increased levels of cytokines in the supernatant, including tumor necrosis factor-α, interferon-γ and interleukin-6 (P<0.01). By contrast, Tim-3 silencing significantly decreased LPS-induced cytokine release (P<0.01). However, Tim-3 silencing did not affect TLR4, MyD88 and NF-κB p65 expression and the release of cytokines in cells that did not undergo treatment with LPS. Therefore, the results of the present study indicate that Tim-3 silencing decreases the viability of FLS in RA and attenuates the LPS-induced inflammatory reaction.