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Journal of the American Society of Nephrology, Vol 5, 1091-1098, Copyright © 1994 by American Society of Nephrology

REGULAR ARTICLES

Uptake of atrial natriuretic peptide and production of cGMP in cultured human glomerular endothelial cells

DF Green, VA Duruibe, G Blyden, RE Laskey and JJ Bourgoignie
Department of Medicine, University of Miami, School of Medicine, FL 33101.

Interactions between human glomerular endothelial cells and atrial natriuretic peptide (ANP) were studied with 125I-alpha-human-ANP binding and intracellular accumulation of cGMP. Uptake for alpha-hANP (1-28 or 5-28) by homogeneous cultures of human glomerular endothelial cells was dose and time dependent with optimal uptake occurring after 30 min of incubation at 37 degrees C. Scatchard analysis of the specific binding data with a two-compartmental model identified both high (Kd = 0.3 nM)- and low (Kd = 10 nM)-affinity receptors, with a binding site density of 12,000 and 18,060 receptors per cell, respectively. alpha-hANP markedly stimulated glomerular endothelial cell-associated cGMP. After a 2-min incubation, cGMP increased 1.3-fold (from 17.88 +/- 1.29 to 23.33 +/- 3 pmol/mg of protein), in the presence of 1 nM ANP, to more than threefold (from 21 +/- .1 to 80.5 +/- 14.5 pmol/mg of protein) with 1 microM ANP (P < 0.05). In contrast, a 10 microM concentration of the clearance receptor C-ANP4-23 increased cGMP by 1.6 +/- 0.6 fold. ANP stimulation of intracellular cGMP was 100 times more sensitive in human glomerular endothelial than in mesangial cells. In comparison, higher doses of bradykinin were necessary to evoke similar responses in glomerular endothelial cells. In the presence of 10 microM bradykinin, cellular cGMP increased by 1.75 +/- 0.6-fold versus control cells. However, unlike ANP, bradykinin- stimulated cGMP synthesis was significantly inhibited by prior treatment with oxyhemoglobin (10(-5) M), an inhibitor of soluble guanylate cyclase, and NG-nitro-L-arginine (NO2Arg), a specific inhibitor of endothelial-derived relaxing factor (EDRF).(ABSTRACT TRUNCATED AT 250 WORDS)




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Copyright © 2008 by the American Society of Nephrology. Online ISSN: 1533-3450 Print ISSN: 1046-6673