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JOURNAL ARTICLE
RESEARCH SUPPORT, NON-U.S. GOV'T
Tyrosine kinase inhibitors and antioxidants modulate NF-kappaB and NOS-II induction in retinal epithelial cells.
American Journal of Physiology 1998 July
Bovine retinal pigmented epithelial (RPE) cells express an inducible nitric oxide synthase (NOS-II) after activation with interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS). Experiments were performed to investigate the effects of tyrosine kinase inhibitors (genistein and herbimycin A) and antioxidants [pyrrolidine dithiocarbamate (PDTC) and butyl hydroxyanisol] on NOS-II induction. The LPS-IFN-gamma-induced nitrite release was inhibited in a concentration-dependent manner by these compounds. Analysis by Northern blot showed that this inhibitory effect correlated with a decrease in NOS-II mRNA accumulation. Analysis by electrophoretic mobility shift assay of the activation of the transcription factor nuclear factor-kappaB (NF-kappaB) involved in NOS-II induction demonstrated that LPS alone or combined with IFN-gamma induced NF-kappaB binding. NF-kappaB activation was not changed by the presence of tyrosine kinase inhibitors but was totally prevented by PDTC pretreatment. Immunocytochemistry experiments confirmed the reduction of the nuclear translocation of NF-kappaB only by PDTC. Our results demonstrated the existence in retinal pigmented epithelial cells of different intracellular signaling pathways in NOS-II induction, since tyrosine kinase inhibitors blocked NOS-II mRNA accumulation without inhibiting NF-kappaB activation. Furthermore, the LPS-IFN-gamma-induced NOS-II mRNA accumulation was sensitive to cycloheximide, suggesting that, in addition to NF-kappaB, transcriptional factors that require new protein synthesis are involved in NOS-II induction.
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