NF-κB inhibits expression of the α1(I) collagen gene

Fibrosis results from an increase in the synthesis and deposition of type I collagen, Fibrosis is frequently associated with inflammation, which is accompanied by increased levels of tumor necrosis factor-alpha (TNF alpha) and activation of the transcription factor NF-kappa B. However, several agents known to activate NF-kappa B, such as phorbol la-myristate 13-acetate (PMA) and TNF alpha, result in decreased expression of type I collagen. Therefore, me directly examined the effects of NF-kappa B on alpha 1(I) collagen gene expression in two collagen-producing cells, NIH 3T3 fibroblasts and hepatic stellate cells (HSCs). Transient transfections of NM. 3T3 cells or HSCs using NF-kappa B p50, p65, and c-Rel expression plasmids with collagen reporter gene plasmids demonstrated a strong inhibitory effect on transcription of the collagen gene promoter, Dose-response curves showed that p65 was a stronger inhibitor of collagen gene expression than was NF-kappa B p50 or c-Rel (maximum inhibition 90%), Transient transfections with reporter gene plasmids containing one or two Spl binding sites demonstrated similar inhibitory effects of NF-kappa B p65 on the activity of these reporter genes, suggesting that the inhibitory effects of NF-kappa B p65 are mediated through the critical Sp1 binding sites in the alpha 1(I) collagen gene promoter. Cotransfection experiments using either a super-repressor I[ke]B or Spl partially blocked the inhibitory effects of p65 on collagen reporter gene activity. Coimmunoprecipitation experiments demonstrated that NF-kappa B and Sp1 do interact in vivo, Nuclear run-on assays showed that NF-kappa B p65 inhibited transcription of the endogenous alpha 1(I) collagen gene. Together, these results demonstrate that NF-kappa B decreases transcription of the alpha 1(I) collagen gene.