TRANSFORMING-GROWTH-FACTOR BETA-1 PREVENTS CYTOKINE-MEDIATED INHIBITORY EFFECTS AND INDUCTION OF NITRIC-OXIDE SYNTHASE IN THE RINM5F INSULIN-CONTAINING BETA-CELL LINE

The aim of this study was to examine lithe growth factor, transforming growth factor beta 1 (TGF beta 1), could prevent induction of nitric oxide synthase and cytokine-mediated inhibitory effects in the insulin -containing, clonal beta cell line RINm5F. Treatment of RINm5F cells f or 24 h with interleukin-1 beta (IL-1 beta) (100 pM) induced expressio n of nitric oxide synthase and inhibited glyceraldehyde-stimulated ins ulin secretion. Combinations of IL-1 beta (100 pM), tumour necrosis fa ctor-alpha (100 pM) and interferon-gamma (100 pM) reduced RINm5F cell viability (determined by the -(4,5-dimethylthiazolyl-2)-2,5-diphenylte trazolium (MTT) reduction assay) and de novo protein synthesis, as mea sured by incorporation of radiolabelled amino acids into perchloric ac id-precipitable protein. Pretreatment of RINm5F cells with TGF beta 1 (10 pM) for 18 or 24 h, prior to the addition of either IL-1 beta or c ombined cytokines, prevented cytokine-induced inhibition of insulin se cretion, protein synthesis and the loss of cell viability. TGF beta 1 pretreatment inhibited cytokine-induced expression and activity of nit ric oxide synthase in RINm5F cells as determined by Western blotting a nd by cytosolic conversion of radiolabelled arginine into labelled cit rulline and nitric oxide. Chemically generated superoxide also induced expression of nitric oxide synthase possibly due to direct activation of the nuclear transcription factor NF kappa B, an effect prevented b y both an antioxidant and TGF beta 1 pretreatment. In conclusion, the mechanism of action of TGF beta 1 in blocking cytokine inhibitory effe cts was by preventing induction of nitric oxide synthase.