Nitric Oxide Production by Human Islets

Although nitric oxide production has been extensively examined in rodents, the generation of nitric oxide by human tissue has been difficult. Nussler et al. (1992) have shown that human hepatocytes produce nitric oxide in response to IL-1, TNF, IFN, and LPS. Nitric oxide, produced by human monocytes treated with LPS, may play an important role in liver disease associated with alcoholism. Monocytes isolated from alcoholics with liver disease produce approximately the same level of nitric oxide as monocytes from control subjects simulated with LPS 

If jS-cell production of nitric oxide participates in IDDM, human islets must produce nitric oxide in response to cytokines. We have shown that a combination of cytokines (lL-1, IFN, and TNF) induce the formation of nitric oxide by isolated human islets (Corbett et al., 1993b). The formation of nitric oxide has been demonstrated by cytokine-induced cGMP accumulation, nitrite formation, and EPR-detectable iron-nitrosyl complex formation (Fig. 12), all of which were prevented by NMMA. The cytokine combination of IFN and lL-1 are required for nitrite production, while TTSIF potentiates IL-1 and IFN-induced nitrite formation by human islets. The cytokine combination of lL-1, TNF, and IFN also influences the physiological function of insulin secretion by human islets. Low concentrations of this cytokine combination slightly stimulate insulin secretion, while high concentrations inhibit insulin secretion, similar to the concentration-dependent effects of lL-1 on rat islet function. NMMA partially prevents the inhibitory effects of this cytokine combination on insulin secretion from human islets, suggesting that nitric oxide may participate in )3-cell dysfunction associated with IDDM. 

We believe that the /3 cell is a source of nitric oxide production by human islets because (1) IL-1 and IFN-induced nitric oxide production by human macrophages has not been clearly demonstrated (2) the cytokine combination of IL-1, IFN, and TNF induces the formation of nitric oxide by human islets either freshly isolated or cultured for 7 days at 25°C (a procedure which removes 80-90% of nonendocrine cells from the islet) and also by islets cryoperserved and (3) NADPH—diaphorase staining reveals that approximately 60-70% of human islet cells treated with cytokines stain for NADPH-diaphorase (J. A. Corbett and M. L. McDaniel, unpublished data). TTiis staining procedure has been shown to colocalize with nitric oxide synthase in a number of cells including rat islets (Corbett et al., 1993c), and nitric oxide synthase has been demonstrated to contain NADPH-diaphorase enzymatic activity (Dawson et al., 1991 Hope et 

Effects of cytokines on the formation of nitric oxide by human islets as determined by EPR spectroscopy. Human islets were treated for 18 hr with 75 U/ml lL-1, 3.5 nM TNF-a, and 750 U/ml IFN-y, the islets were then isolated, and EPR spectroscopy was performed as described previously (Corbett et al., 1993b). Cytokine induced nitric oxide formation is demonstrated by the genetation of an EPR detectable g = 2.04 iton-nitrosyl complex which is prevented by 0.5 mM NMMA. Reproduced with permission from Proc. Nall. Acad. Set. U S.A. (Corbett et al., 1993b). 

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