Induction of matrix metalloproteinase-1 gene transcription by tumour necrosis factor alpha via the p50/p50 homodimer of nuclear factor-kappa B in activated human hepatic stellate cells

Abstract

Background/aims: Liver injury results in the activation of hepatic stellate cells (HSCs), which in turn produce matrix metalloproteinase (MMP) in response to pro-inflammatory cytokines for tissue remodelling. This study explored the transcriptional induction of the MMP-1 gene by tumour necrosis factor-alpha (TNF-alpha) in HSCs.

Methods: The LI90 human HSC line was used in the present study. Gelatin zymography, enzyme-linked immunosorbent assay, Northern blotting and gene promoter-reporter assays were used to analyse the induction of MMP-1 protein, mRNA expression and gene transcription respectively. Deletional or site-directed mutations were introduced into the promoter region and transiently transfected into LI90 cells to determine the cis-acting elements necessary for TNF-alpha inducibility. Gel shift mobility assays were used to determine the transcriptional factors involved in the TNF-alpha responsiveness.

Results: TNF-alpha upregulated MMP-1 protein and mRNA expression in a dose-dependent manner. A time-course experiment revealed a rapid induction of MMP-1 mRNA expression after TNF-alpha treatment. Mutation in a putative nuclear factor (NF)-kappaB-binding site at -2541 bp almost completely abolished the TNF-alpha response to MMP-1 gene-promoter activity, suggesting transcriptional regulation of MMP-1 expression by TNF-alpha via this site. Electrophoretic mobility shift assay and supershift assays indicated that this transcriptional regulation was regulated via the p50/p50 homodimer of NF-kappaB.

Conclusions: MMP-1 gene expression might be induced by TNF-alpha via the p50/p50 homodimer of NF-kappaB in activated human HSCs.