Expression of inducible nitric oxide (NO) synthase and arginine-metabolic relative enzymes in retinal pigment epithelial (RPE) cells and the effect of NO on tight junction of RPE cells / 中华眼底病杂志

ABSTRACT

Objective To detect the induction of inducible nitric oxide synthase (iNOS) and nitric oxide (NO) production in immunostimulated retinal pigment epithelial (RPE) cells to seek for the supplying of the arginine, a substrate for NOS; as well as the effects of produced NO on the tight junction of RPE-J cells. Methods Rat′s RPE-J cells were treated with interferon-?(INF-?), tumor necrosis factor-?(TNF-?) and lipopolysaccharide (LPS), and Northern and Western blotting were applied to analyze the expression of the citrulline-NO cycle enzymes and related enzymes and the effect of dexamethasone and cyclic adenosine monophosphate (camp) on the expression of iNOS. Immunocytochemistry reaction and Western blotting were used to evaluate the effect of produced NO on the tight junctions of RPE-J cells. Results iNOS and argininosuccinate synthetase (AS) were highly induced at both mRNA and protection levels in immunostimulated RPE cells while arginiosuccinate lyase (AL) was not induced. NO was produced by cells after stimulation with TNF?, IFN? and LPS. The induction of iNOS mRNA and the production of NO by these immunostimulated cells was further enhanced by cAMP. NO was produced from citrulline as well as from arginine. And the produced NO impaired the tight junction of RPE-J cells, decreased the production of tight junction related protein ZO-1. Conclusion In activated RPE-J cells, citrulline-arginine recycling is important for NO production, and the produced NO weakened the function of tight junction of RPE-J cells.