Keratocyte Differentiation Is Regulated by NF-kappa B and TGF beta Signaling Crosstalk

Interleukin-1 (IL-1) and transforming growth factor-beta (TGF beta) are important cytokines involved in corneal wound healing. Here, we studied the effect of these cytokines on corneal stromal cell (keratocyte) differentiation. IL-1 beta treatment resulted in reduced keratocyte phenotype, as evident by morphological changes and decreased expression of keratocyte markers, including keratocan, lumican, ALDH3A1, and CD34. TGF beta 1 treatment induced keratocyte differentiation towards the myofibroblast phenotype. This was inhibited by simultaneous treatment with IL-1 beta, as seen by inhibition of alpha-SMA expression, morphological changes, and reduced contractibility. We found that the mechanism of crosstalk between IL-1 beta and TGF beta 1 occurred via regulation of the NF-kappa B signaling pathway, since the IL-1 beta induced inhibition of TGF beta 1 stimulated keratocyte-myofibroblast differentiation was abolished by a specific NF-kappa B inhibitor, TPCA-1. We further found that Smad7 participated in the downstream signaling. Smad7 expression level was negatively regulated by IL-1 beta and positively regulated by TGF beta 1. TPCA-1 treatment led to an overall upregulation of Smad7 at mRNA and protein level, suggesting that NF-kappa B signaling downregulates Smad7 expression levels in keratocytes. All in all, we propose that regulation of cell differentiation from keratocyte to fibroblast, and eventually myofibroblast, is closely related to the opposing effects of IL-1 beta and TGF beta 1, and that the mechanism of this is governed by the crosstalk of NF-kappa B signaling.