Selective up-regulation of P2X₄-receptor gene expression by interferon-γ in vascular endothelial cells

Extracellular nucleotides are involved in the development of vascular inflammation. However, little is known about whether effects of nucleotides are modulated under inflammatory states. We investigated effects of interferon-γ (INF-γ) on ATP-induced responses in vascular endothelial cells. Treatment of human umbilical vein endothelial cells (HUVECs) with IFN-γ for 24 h resulted in an enhancement of the ATP-induced increase in intracellular Ca²⁺ concentration ([Ca²⁺]_i) without affecting the UTP-induced one. The increased Ca²⁺ response to ATP in IFN-γ-treated cells was dependent on the extracellular Ca ²⁺, and was not inhibited by the phospholipase C inhibitor U73122. RT-PCR and Western blotting revealed that HUVECs dominantly expressed P2X ₂ receptor. IFN-γ increased P2X₄-receptor mRNA and protein, accompanied by an increase in ATP-triggered membrane current. IFN-γ did not affect P2X₄-receptor mRNA stability, but increased P2X₄-receptor gene transcription in a cycloheximide- insensitive manner. IFN-γ stimulated phosphorylation of signal transducer and activator of transcription-1 (STAT1). Epigallocatechin gallate (EGCG), an inhibitor of STAT1-mediated signaling, and AG490, a Janus kinase (JAK) inhibitor, impaired P2X₄-receptor mRNA up-regulation by IFN-γ. These results indicate that INF-γ selectively increases P2X ₄-receptor gene expression, leading to an up-regulation of purinergic signaling in vascular endothelial cells.