Resistin promotes CCL4 expression through toll-like receptor-4 and activation of the p38-MAPK and NF-κB signaling pathways: implications for intervertebral disc degeneration.

OBJECTIVE

This study was to investigate whether resistin induces the expression of chemokine ligand 4 (CCL4) during Intervertebral disc degeneration (IVDD) and whether toll-like receptor-4 (TLR-4) and the nuclear factor-κB (NF-κB) signaling pathway are involved in this process.

METHODS

The expression pattern of resistin and CCL4 in different degenerated human nucleus pulposus (NP) tissues were measured by quantitative reverse transcription-polymerase chain reaction (qPCR); Effect of resistin on the migration of macrophages was measured by cell migration assay. Resistin-induced CCL4 expression were analyzed by qPCR, Enzyme-linked immunosorbant assay (ELISA) and cell immunofluorescence. Involvement of TLR-4, p38-mitogen-activated protein kinase (p38-MAPK), and NF-κB signaling pathways were studied by small interfering RNA (siRNA) or Lenti-virus mediated knockdown, co-immunoprecipitation, and chromatin immunoprecipitation (ChIP) assay.

RESULTS

Expression of resistin and CCL4 was elevated in degenerated NP tissue. Resistin promoted macrophage migration through CCL4 and its receptor. Expression of CCL4 was significantly increased by resistin treatment. The pharmacological inhibition or siRNA knockdown of TLR-4 blocked the resistin-induced CCL4 expression. Co-immunoprecipitation data confirmed the binding of resistin to TLR4. Pharmacological inhibition of the NF-κB and p38-MAPK signaling pathways attenuated the resistin-induced CCL4 expression. A ChIP assay and lentivirus mediated knockdown showed that resistin regulate CCL4 expression through p65.

CONCLUSION

This study shows that resistin binds to TLR4 and increase the expression of CCL4 through p38-MAPK and NF-κB signaling pathways in NP cells, and this expression causes infiltration of macrophages. This study might provide a feasible therapeutic target for controlling the inflammatory response associated with IVDD.