Cocaine-Induced Endothelial Dysfunction: Role of RhoA/Rho Kinase Pathway Activation

• Published in: Blood 2012-11, Vol.120 (21), p.2177-2177
• Main Authors: Pereira, Jaime, Saez, Claudia G, Pallavicini, Julio, Pereira-Flores, Karla, Mendoza, Camila, Hernández, Romina, Novoa, Ulises, Ocaranza, María Paz, Massardo, Teresa, Mezzano, Diego, Jalil, Jorge
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V120.21.2177.2177

Abstract 2177 Cocaine abuse is associated with an increased risk of cardiac and cerebrovascular events, such as myocardial infarction, sudden cardiac death, and ischemic stroke. The underlying mechanisms leading to these complications are not fully understood although intravascular thrombus formation and accelerated atherosclerosis are prominent findings. We have recently demonstrated that chronic cocaine use is associated with endothelial dysfunction (Sáez et al. Thromb Res 2011; 128: 18), a key event in the onset and progression of atherosclerosis. There is growing evidence that the RhoA/Rho kinase (ROCK) pathway has an important pathophysiological role in vascular endothelial dysfunction. Accordingly, we hypothesized that cocaine use induces activation of RhoA/ROCK pathway. The main aim of this work was to investigate the activation of RhoA/ROCK pathway through ex vivo, in vivo and in vitro studies. Ex vivo studies. We studied 13 cocaine dependent individuals (aged 19–52 years mean age 37 years) who met DSM-IV criteria for cocaine dependence, seeking treatment for cocaine abuse and age, sex-matched healthy controls (aged 20–49 years, mean age 35 years). Samples were obtained at admission, within 72 hours of drug exposure. Endothelial cell damage was determined by enumerating circulating endothelial cells (CECs). Rho-kinase activity was assessed by the levels of phosphorylated to total myosin light chain phosphatase 1 (MYPT1-P/T) in circulating leukocytes. In vivo studies. Adult male Sprague-Dawley rats were randomly assigned to receive either cocaine (30mg/kg, provided by NIDA, USA) or saline intraperitoneally once daily for 21 days. The levels of aortic phosphorylated MYPT1 (phospho-MYPT1) were assessed by western blot in aorta extracts. In vitro experiments. Human umbilical vein endothelial cells (HUVECs) were cultured under standard conditions and supplemented for 5 hours with plasma from chronic cocaine users, normal plasma, cocaine (10μM) or vehicle. After media removal, HUVECs were either lysed for determination of ROCK activity or co-cultured with resting platelets and immunostained for von Willebrand factor (FVW). Platelet adhesion was evaluated by immunofluorescence microsocopy. Experiments were conducted in the presence or absence of ROCK inhibitors, Y-27632 (10 μM) or atorvastatin (10μM). Cocaine users showed significantly elevated number of CECs compared to the controls (65 ± 6.6 vs 14 ± 3.4 cells/mL, p: 0.0002). In the control subjects, leu