Ethanol‐Induced Up‐Regulation of Candidate Plasminogen Receptor Annexin II in Cultured Human Endothelial Cells

Abstract

Epidemiological studies indicate that moderate alcohol consumption reduces the risk for coronary heart disease and that this cardioprotective benefit may be mediated, in part, by increased fibrinolysis. Endothelial cells (ECs) synthesize plasminogen activators, tissue-type plasminogen activator (t-PA), urokinase-type plasminogen activator (u-PA), receptors for plasminogen activators, and a receptor for plasminogen, annexin II (Ann-II). These receptors localize and facilitate receptor-bound plasminogen activator-mediated conversion of receptor-bound plasminogen to receptor-bound plasmin on the EC surface, which results in the regulated expression of surface-localized EC fibrinolytic activity. Ethanol is a systemic factor that affects these components, which increases EC fibrinolysis and hence reduces the risk for thrombosis, coronary heart disease, and myocardial infarction (MI). This study was carried out to determine whether low ethanol (0.1% v/v) increased plasminogen receptor, Ann-II antigen (western blot), messenger ribonucleic acid (mRNA) (reverse transcription polymerase chain reaction; RT-PCR) expression, activity (ligand binding/Scatchard analysis), and hence fibrinolysis (plasmin generation) in cultured human ECs. Plasminogen receptor activity increased approximately 2-fold (2.5 vs. 5.6 x 10(6) sites/cell), as evidenced by increased 125I-labeled Glu-plasminogen ligand binding/Scatchard analysis. In addition, western blot analyses indicated an increase in Ann-II antigen, and mRNA levels increased approximately 2-fold (RT-PCR). This increase in Ann-II expression was concomitant with approximately 2- to 3-fold sustained increase (approximately 24 hr) in surface-localized EC fibrinolytic activity. Nuclear transcription run-on assays showed an approximately 5- to 6-fold increase in new 32P-labeled Ann-II mRNA levels, compared with controls (no ethanol). These results demonstrated that low ethanol increased Ann-II antigen/mRNA levels and up-regulated Ann-II gene expression at the transcriptional level. The results further identify and define the contribution and role of the plasminogen receptor, Ann-II, in the ethanol-induced mechanism of increased EC fibrinolysis that may underlie and contribute, in part, to the cardioprotective benefit associated with moderate alcohol consumption.