Suppression of the TNFα-induced increase in IL-1α expression by hypochlorite in human corneal epithelial cells

PURPOSE. In response to injury, activated neutrophils release tumor necrosis factor (TNF)-alpha and myeloperoxidase (MPO). TNFalpha in turn causes human corneal epithelial cells to secrete interleukin (IL)-1alpha, whereas MPO results in formation of HClO/ OCl-. The effect of HClO/OCl- on the expression of the IL-1alpha gene and protein is unknown. The current study was undertaken to examine in immortalized human corneal epithelial cells whether NaOCl alters TNFalpha-induced increases in expression of IL-1alpha gene and protein. METHODS. Semiquantitative RT-PCR and EUSA characterized IL-1alpha gene and protein expression, respectively. TNFalpha-induced nuclear transfer of nuclear factor (NF)-kappaB was measured by electrophoretic mobility shift assay (EMSA). The a isoform of inhibitory protein kappaB (IkappaBalpha) was identified by Western blot analysis. RESULTS. Exposure to NaOCl (0.75 mM) for 10 minutes caused suppression of TNFa-induced increases in IL-1alpha mRNA and protein, declines in NFkappaB nuclear transfer, and a modification of IkappaBalpha, based on a bandshift detected by Western blot analysis. Modified IkappaBalpha became resistant to TNFalpha-induced proteolysis. Methionine sulfoxide reductase A (MsrA, 10 muM) eliminated the NaOCl-induced IkappaBalpha bandshift. CONCLUSIONS. NaOCl oxidizes IkappaBalpha at methionine residues and thereby suppresses dissociation Of IkappaBalpha from NFkappaB. Decreased dissociation could in turn suppress TNFalpha-induced activation of NFkappaB, resulting in declines in expression of IL-1alpha gene and protein. These effects suggest that release of HClO/OCl- in vivo by activated neutrophils may counterbalance TNFalpha-induced NFkappaB-dependent secretion if IL-1alpha and suppress an excessive inflammatory reaction.