OBJECTIVE: Ultrafiltration failure is a common problem in continuous ambulatory peritoneal dialysis. Recent work has indicated a role of enhanced expression of nitric oxide synthase (NOS) in ultrafiltration failure. However, the conditions predisposing to increased generation of NO by the peritoneum have not been studied in detail and the cell types potentially involved have not been tested individually. DESIGN: We performed experiments in human peritoneal mesothelial cells (HPMC) in culture. Amino acid-based dialysis solution (Nutrineal; Baxter Deutschland GmbH, Munchen, Germany), L-arginine, and glucose-containing control solutions were used and we observed the effects on the HPMC. We reasoned that amino acid-based dialysis solutions containing L-arginine, the substrate of NOS, might influence mesothelial NO generation. Nitric oxide production was measured in the supernatant using the Griess reaction. We studied the effect of the combined NOS inhibitor L-NMMA and specified the isoform of NOS involved. RESULTS: In serum-free control medium, the cells exhibited baseline generation of nitrite at a rate of 5.4 +/- 0.5 micromol/g protein. Addition of 6 mmol/L L-arginine to the control medium increased nitrite significantly (11.8 +/- 0.66 micromol/g protein, p < 0.002), as did amino acid-based dialysis solution (15.7 +/- 1.3 micromol/g protein, p < 0.002); L-NMMA caused a significant reduction of this nitrite. HPMC expressed eNOS (NOSIII) when grown in L-arginine-supplemented medium, shown by immunocytochemistry and by reverse transcriptase-polymer chain reaction. Biochemical exposure to a calcium ionophore in 1 micromol/L concentration approximately doubled the nitrite production by L-arginine-incubated cells. CONCLUSION: Peritoneal mesothelial cells generate NO in vitro. Generation of NO increased further in response to L-arginine supplementation of the culture medium and to amino acid-containing dialysis solution. Mesothelial cells express eNOS, which was likely involved in the observed peritoneal NO generation.

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