TIR-domain-containing adapter-inducing interferon-β contributes to TLR3/TLR4 triggered apoptosis and inflammation in nucleus pulposus cells

Abstract

The senescence and degeneration of the intervertebral disc are closely related to the reduction of nucleus pulposus (NP) cells caused by apoptosis. TIR-domain-containing adapter-inducing interferon-β (TRIF) is an adapter for Toll-like receptors 3/4 (TLR3/4), which involves in cell apoptosis. The aim of this study is to detect the role of TRIF in the apoptotic progress of NP cells. The expression of collagen II, aggrecan, TLR3/4, and TRIF were analyzed in different degrees of degenerated human NP samples from patients. NP cells were isolated from mild degenerated tissues and cultured with IL-1β to accelerate the degradation, and treated with TLR3/4 protein. siRNA was used to silence TRIF gene expression, and TRLF-plasmid was used to upregulate TRLF gene expression. We used flow cytometry assay to analyze cell apoptosis. The expression of collagen II, aggrecan, TLF3/4, TRIF, caspase-8/3, MMP-13, TNF-α was determined by immunofluorescence, Western blot, or RT-PCR. That the expression of collagen II and aggrecan markedly decreased, but TLF3/4, TRIF, caspase-8/3, MMP-3, TNF-α, and IL-1β were increased in severely degenerated disc tissues. IL-1β treatment induced NP cell degeneration and TLF3/4, TRIF, caspase-8/3, MMP-3, TNF-α overexpression. TLF3/4 protein treatment promoted NP cell degeneration and apoptosis by upregulation of TRIF, caspase-8/3, MMP-3, and TNF-α. Furthermore, TRIF silencing reversed the negative effect of TLF3/4 overexpression, and TRIF overexpression played the same role in NP cell apoptosis. Based on these results, we believe that TRIF is activated in a degenerated intervertebral disc. TLF3/4 promotes NP cell apoptosis and inflammation through the TRLF adaptor. TRLF expression is positively related to the apoptosis and inflammation in NP cells. These results suggest a therapeutic potential of the TRIF in the treatment of disc degeneration.