Estimation of EDRF and nitric oxide release using [3H]GTP-labeled human platelets.

Abstract:

:We have developed a new bioassay for endothelium-derived relaxing factor (EDRF) or nitric oxide (NO) using human [3H]guanosine triphosphate (GTP)-labeled platelets. The labeled platelets were preincubated with isobutyl-methylxanthine and co-cultured with endothelial cells and the [3H]cyclic guanosine monophosphate (cGMP) formed was isolated by ion-exchange chromatography. Endothelial cells, either in monolayer cells or in suspension, increased platelet cGMP accumulation dose-dependently, a significant increase being detected with 5000 endothelial cells or more/assay when suspended cells were used. Co-culturing with the same number of skin fibroblasts failed to elevate platelet cGMP. Preincubation of endothelial cells with bradykinin and superoxide dismutase (SOD) synergistically potentiated the increase in platelet cGMP, but was attenuated by Nomega-nitro-L-arginine, with partial restoration by L-arginine but not by D-arginine. These compounds, however, did not affect cGMP accumulation by sodium nitroprusside. Moreover, preincubation of the labeled platelets with the NO synthase inhibitor prior to EDRF assay had no effect. We conclude that [3H]GTP-labeled platelets could provide a simple, sensitive and specific bioassay for estimating EDRF or NO release.

journal_name

Thromb Res

journal_title

Thrombosis research

authors

Kishi Y,Watanabe R,Ashikaga T,Numano F

doi

10.1016/0049-3848(95)00082-3

subject

Has Abstract

pub_date

1995-06-15 00:00:00

pages

483-93

issue

6

eissn

0049-3848

issn

1879-2472

pii

0049-3848(95)00082-3

journal_volume

78

pub_type

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