Received for publication December 14, 2007.
Revised June 12, 2008.
Accepted for publication June 13, 2008.

P2X₇ receptor activation amplifies lipopolysaccharide-induced vascular hyporeactivity via interleukin-1 release

Abstract

Lipopolysaccharide (LPS) stimulates cytoplasmic accumulation of pro-interleukin (IL)-1. Activation of P2X₇ receptors stimulates conversion of pro-IL-1 into mature IL-1, which is then secreted. Because both LPS (in vivo) and IL-1(in vitro) decrease vascular reactivity to contractile agents, we hypothesized that: 1) P2X₇ receptor activation contributes to LPS-induced vascular hyporeactivity; and 2) IL-1 mediates this change. Thoracic aortas were obtained from 12-week-old male C57BL/6 mice. The aortic rings were incubated 24 hours in Dulbecco's Modified Eagle's Medium (DMEM), LPS, benzoylbenzoyl-ATP (BzATP, P2X₇ receptor agonist), LPS plus BzATP, oxidized ATP (oATP, P2X₇ receptor antagonist) or oATP plus LPS plus BzATP. Following the treatment, the rings were either mounted in a myograph for evaluation of contractile activity, or homogenized for IL-1 and inducible nitric oxide synthase (iNOS) protein measurement. In endothelium-intact aortic rings, phenylephrine (PE)-induced contractions were not altered by incubation with LPS or BzATP, but significantly decreased in aortic rings incubated with LPS plus BzATP. Treatment with oATP or IL-1ra (IL-1 receptor antagonist) reversed LPS plus BzATP-induced hyporeactivity to PE. In the presence of N^G-nitro-L-arginine methyl ester (L-NAME) or N-([3-(aminomethyl)phenyl]methyl)ethanimidamide (1400W, selective iNOS inhibitor), the vascular hyporeactivity induced by LPS plus BzATP on PE responses was not observed. BzATP augmented LPS-induced IL-1 release and iNOS protein expression, and these effects were also inhibited by oATP. Moreover, incubation of endothelium-intact aortic rings with IL-1 induced iNOS protein expression. Thus, activation of P2X₇ receptor amplifies LPS-induced hyporeactivity in mouse endothelium-intact aorta, which is associated with IL-1-mediated release of nitric oxide by iNOS.

Key words: P2X7 receptor, inflammation, interleukin-1beta, lipopolysaccharide, nitric oxide, vascular reactivity