C-reactive protein decreases endothelial nitric oxide synthase activity via uncoupling

Uma Singh, Sridevi Devaraj, Jeannette Vasquez-Vivar, Ishwarlal Jialal

Research output: Contribution to journalArticle

94 Scopus citations

Abstract

C-reactive protein (CRP), a cardiovascular risk marker, induces endothelial dysfunction. We have previously shown that CRP decreases endothelial nitric oxide synthase (eNOS) expression and bioactivity in human aortic endothelial cells (HAECs). In this study, we examined the mechanisms by which CRP decreases eNOS activity in HAECs. To this end, we explored different strategies such as availability of tetrahydrobiopterin (BH4)-a critical cofactor for eNOS, superoxide (O2 -) production resulting in uncoupling of eNOS and phosphorylation/dephosphorylation of eNOS. CRP treatment significantly decreased levels of BH4 thereby promoting eNOS uncoupling. Pretreatment with sepiapterin, a BH4 precursor, prevented CRP-mediated effects on BH4 levels, superoxide production as well as eNOS activity. The gene expression and enzymatic activity of GTPCH1, the first enzyme in the de novo biosynthesis of BH4, were significantly inhibited by CRP. Importantly, GTPCH1 is known to be regulated by cAMP-mediated pathway. In the present study, CRP-mediated inhibition of GTPCH1 activity was reversed by pretreatment with cAMP analogues. Furthermore, CRP-induced O2 - production was reversed by pharmacologic inhibition and siRNAs to p47 phox and p22 phox. Additionally, CRP treatment significantly decreased the eNOS dimer: monomer ratio confirming CRP-mediated eNOS uncoupling. The pretreatment of cells with NO synthase inhibitor (N-nitro-l-arginine methyl ester [l-NAME]) also prevented CRP-mediated O2 - production further strengthening CRP-mediated eNOS uncoupling. Additionally, CRP decreased eNOS phosphorylation at Ser1177 as well as increased phosphorylation at Thr495. CRP appears to mediate these effects through the Fcγ receptors, CD32 and CD64. To conclude, CRP uncouples eNOS resulting in increased superoxide production, decreased NO production and altered eNOS phosphorylation.

Original languageEnglish (US)
Pages (from-to)780-791
Number of pages12
JournalJournal of Molecular and Cellular Cardiology
Volume43
Issue number6
DOIs
StatePublished - Dec 2007

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Keywords

  • CRP
  • Endothelial nitric oxide synthase
  • Oxidative stress
  • Tetrahydrobiopterin
  • Uncoupling

ASJC Scopus subject areas

  • Molecular Biology
  • Cardiology and Cardiovascular Medicine

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