IL-1a regulates the expression of antimicrobial peptides in human keratinocytes

Objectives: Human antimicrobial peptides (AMPs) are produced by keratinocytes in skin and mucosa, and play an important role in a host defense system. We found that calprotectin (S100A8/S100A9), an AMP constitutively expressed in oral mucosa, inhibited a growth of Porphyromonas gingivalis and calprotectin expression is regulated by several inflammatory cytokines in human keratinocytes. However, it is not well known what factors regulate AMP expression. On the other hand, extracellular matrix (ECM) affect gene expression, cellular functions and proliferation. In this study, we investigated whether AMP expression is regulated by several ECMs and interleukin-1a, an autonomous cytokine in human skin and mucosal keratinocytes. Methods: Human skin-derived HaCaT cells and mucosa-derived TR146 cells were inoculated in the dishes coated with ECM including collagen type I(Col I), collagen type IV(Col IV), fibronectin(FN) or laminin(LM), and cultured with or without 20 ng/ml IL-1a. Cells were immunostained with anti-calprotectin antibody and fluorescent dye-conjugated IgG. Total RNA was extracted from the cultured cells and followed by Northern blotting or RT-PCR. The amount of calprotectin protein in the cell fraction was determined by ELISA. Results: Four ECMs(Col I, Col IV, FN and LM) had no effect on the expression of calprotectin in HaCaT and TR146. However, IL-1a increased mRNA expressions of S100A8/S100A9 in both cells cultured on non-coated (control) and ECM-coated dishes, and also increased calprotectin protein in both cells. In the cells cultured on control dish, IL-1a increased mRNA expression of AMPs including S100A7, S100A8/S100A9, lipocalin 2 and b-defensin 2 in HaCaT and TR146. Conclusion: These results suggest that AMP expression in human skin and mucosal keratinocytes is regulated by IL-1a.