Epidemiological studies demonstrate that consuming alcohol or red wine moderately, 1-4 drinks/day, reduces the risk for CHD-related mortality. Furthermore, these studies show that this cardioprotection may be due, in part, to increased fibrinolysis. Endothelial cells (ECs) regulate fibrinolysis through the synthesis of plasminogen activators (PAs, t-PA and u-PA) and plasminogen activator inhibitor type-1 (PAI-1), the major physiological regulator of fibrinolysis. In addition ECs serve as the main site of surface-localized fibrinolysis that regulates homeostasis. Thus, ECs and PAI-1 plays a pivotal role both in fibrinolytic homeostasis and in the pathogenesis of CHD and MI. Reduction in plasma PAI-1 levels by systemic factors, such as alcohol and wine polyphenols, will increase fibrinolysis and hence reduce the risk for thrombosis, CHD and eventual MI. Our preliminary studies demonstrate that consuming moderate ethanol or polyphenols (catechin, quercetin) reduces in vivo expression of both PAl-1 protein and mRNA in wild type C57BL/6J mice. Our in vitro experiments show that ethanol down-regulates PAI-1 gene transcription in a time- and dose-dependent manner concomitant with increased expression of fibrinolytic activity in cultured human ECs. Furthermore, we have identified a 251-bp promoter fragment in the PAI-1 gene that mediates this ethanol-induced suppression of PAI-1 expression. Thus, the overall goal of Project 2 is to identify the molecular mechanisms through which ethanol/polyphenols repress PAI-1 gene expression in vivo and in vitro, resulting in increased fibrinolysis. Specific studies will determine the effect of ethanol/polyphenols on the in vivo expression and role of PAI-1 in and deficient PAI-1 mice (Aiml); examine possible cross-talk between PAI-1 and eNOS in regulating vascular function in wild-type and PAI-1 deficient mice (Aim 2); establish the role of vitronectin, LRP, VLDLr, and ?t6133in modulating PAI-1 in cultured ECs (Aim 3); and identify the ethanol/polyphenols responsive cis-element(s) in the PAI-1 gene, and characterize the transcription factor(s) that bind these responsive cis-element(s) (Aim 4). Results from these studies will provide insight into understanding how moderate ethanol/polyphenols repress PAI-1 gene transcription that may contribute to increased fibrinolysis and the cardioprotection attributed to moderate alcohol and red wine consumption.
