Inhibition of ATP-binding cassette transporter downregulates interleukin-1beta-mediated autocrine activation of human dermal fibroblasts

Fibroblasts constitute an important source of cytokines during inflammatory processes in the skin. Interleukin-1 is a potent, pleiotropic cytokine tha t is induced in activated human dermal fibroblasts. Interleukin-1 further i nduces many inflammatory mediators, including the chemokine interleukin-8. As fibroblasts express both interleukin-1 and the interleukin-1 receptor co mplex, the cellular response may be enhanced by autocrine activation. Inter leukin-1 alpha and interleukin-1 beta lack a signal peptide and are translo cated at the plasma membrane using an alternative secretory pathway, which may involve ATP-binding cassette transporter proteins. We hypothesize that inhibition of this pathway prevents secretion of interleukin-1, thereby dow nregulating interleukin-1-dependent autocrine induction of interleukin-8. W e used the ATP-binding cassette 1 transporter inhibitor glybenclamide, whic h has been previously shown to block interleukin-1 beta secretion in human monocytes. Using enzyme-linked immunosorbent assay, we assessed the effect of glybenclamide on interleukin-8 production in human dermal fibroblasts. I n interleukin-1 beta -transfected human dermal fibroblasts, interleukin-8 w as induced through an autocrine activity of interleukin-1 beta. Glybenclami de disabled this activation loop and significantly reduced interleukin-8. I n human dermal fibroblasts that were stimulated with tumor necrosis factor a to reach high interleukin-1 expression levels, glybenclamide similarly su ppressed interleukin-8. In contrast, glybenclamide did not affect interleuk in-8 production in cells stimulated with interleukin-1 only. Glybenclamide did not affect caspase-1 in fibroblasts, which was expressed as an inactive precursor form, irrespective of treatments with tumor necrosis factor alph a and/or glybenclamide. Using overexpressing, interleukin-1-transfected COS -1 cells, inhibition of interleukin-1 alpha and interleukin-1 beta secretio n was directly demonstrated on Western blots. These results are consistent with glybenclamide preventing externalization of interleukin-1 and subseque nt autocrine induction of interleukin-8 in human dermal fibroblasts. Acting through such a mechanism, ATP-binding cassette transporter inhibitors may downregulate inflammation locally.