Increased plasma phenylacetic acid in patients with end-stage renal failure inhibits iNOS expression
J. Jankowski, … , W. Zidek, M. Tepel
J. Jankowski, … , W. Zidek, M. Tepel
Published July 15, 2003
Citation Information: J Clin Invest. 2003;112(2):256-264. https://doi.org/10.1172/JCI15524.
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Article Endocrinology

Increased plasma phenylacetic acid in patients with end-stage renal failure inhibits iNOS expression

Abstract

NO prevents atherogenesis and inflammation in vessel walls by inhibition of cell proliferation and cytokine-induced endothelial expression of adhesion molecules and proinflammatory cytokines. Reduced NO production due to inhibition of either eNOS or iNOS may therefore reinforce atherosclerosis. Patients with end-stage renal failure show markedly increased mortality due to atherosclerosis. In the present study we tested the hypothesis that uremic toxins are responsible for reduced iNOS expression. LPS-induced iNOS expression in mononuclear leukocytes was studied using real-time PCR. The iNOS expression was blocked by addition of plasma from patients with end-stage renal failure, whereas plasma from healthy controls had no effect. Hemofiltrate obtained from patients with end-stage renal failure was fractionated by chromatographic methods. The chromatographic procedures revealed a homogenous fraction that inhibits iNOS expression. Using gas chromatography/mass spectrometry, this inhibitor was identified as phenylacetic acid. Authentic phenylacetic acid inhibited iNOS expression in a dose-dependent manner. In healthy control subjects, plasma concentrations were below the detection level, whereas patients with end-stage renal failure had a phenylacetic acid concentration of 3.49 ± 0.33 mmol/l (n = 41). It is concluded that accumulation of phenylacetic acid in patients with end-stage renal failure inhibits iNOS expression. That mechanism may contribute to increased atherosclerosis and cardiovascular morbidity in patients with end-stage renal failure.

Authors

J. Jankowski, M. van der Giet, V. Jankowski, S. Schmidt, M. Hemeier, B. Mahn, G. Giebing, M. Tölle, H. Luftmann, H. Schlüter, W. Zidek, M. Tepel

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Expression of iNOS measured by real-time PCR, protein blotting of iNOS, ...
Expression of iNOS measured by real-time PCR, protein blotting of iNOS, or nitrite formation in RAW 264.7 cells. (a) The iNOS expression was measured by real-time PCR after stimulation by LPS (1 μg/ml) (control) and in the presence of various concentrations of PAA (0.1 mmol/l, 0.5 mmol/l, 1.0 mmol/l, and 5.0 mmol/l). Basal, unstimulated iNOS expression was set to 100% (n = 6). *P < 0.05 compared with control. (b) Representative protein blotting of iNOS and β-actin after 12 hours of stimulation (+) or without stimulation (–) of RAW 264.7 cells with LPS (1 μg/ml) and in the presence of PAA (0.1 mmol/l, 0.5 mmol/l, 1.0 mmol/l, and 5.0 mmol/l). The iNOS protein was detected as a band with a molecular mass of approximately 125 kDa. (c) Signals of iNOS were quantified and normalized to those of β-actin using a bioimaging analyzer. Data represent means of triplicate determinations from each of three protein preparations (n = 3). *P < 0.05 compared with control. (d) Effect of various concentrations of PAA (0.1 mmol/l, 0.5 mmol/l, 1.0 mmol/l, and 5.0 mmol/l) on LPS-induced nitrite production in RAW 264.7 cells. Data are means ± SEM (n = 6). *P < 0.05 compared with control (+).