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

Abstract

2177 Background. 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. Objectives. The main aim of this work was to investigate the activation of RhoA/ROCK pathway through ex vivo, in vivo and in vitro studies. Methods. 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). Results. 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, leukocyte mean MYPT1-P/T ratio was 2.2 ± 0.8 whereas in cocaine addicts were significantly increased (9.8 ± 2.8; p 0.015). ROCK activity was higher by 100% (p: 0.019) in the aortic wall of the cocaine-treated rats compared to sham animals. HUVECs supplemented with plasma from cocaine users showed an increase in ROCK activity by 25% (p: 0.039), released significantly higher amount of FVW (p<0.05) and adhered a larger number of platelets (22.6±5 vs 7.9±3 platelets/cell, respectively; p: 0.006) compared with control plasma. Cocaine exposure induced a dramatically higher number of platelets adhered to HUVEC than in vehicle-treated cells (220±73 vs 10.2±1.1 platelets/cell, respectively; p>0.001).ROCK inhibitors, atorvastatin and Y-27632 reduced the release of FVW by HUVECs exposed to plasma from cocaine users by 65% (p: 0.004) and strongly inhibited platelet adhesion (by 75% in plasma-treated cells and by 90% in HUVECs exposed to cocaine, p:< 0.006). Conclusions. We found an increase in Rho kinase activity in peripheral leukocytes of cocaine abusers, in the vessel wall of rats exposed to cocaine and a marked positive effect of ROCK inhibitors on the cellular injury induced by cocaine or plasma from cocaine consumers on endothelial cells. Collectively, these data suggest that activation of RhoA/ROCK pathway plays a key role in cocaine-induced endothelial dysfunction. Inhibition of ROCK may provide therapeutic benefits in a comprehensive treatment for cocaine addiction. Disclosures: No relevant conflicts of interest to declare.

Cocaine Induces Cytoskeletal Changes in Cardiac Myocytes: Implications for Cardiac Morphology

“…A recent study by Lee et al [ 6 ] demonstrated reductions in cardiac fibrosis following oestradial treatment, with this abrogation reported to be mediated by the downregulation of Rho-ROCK signalling, cofilin phosphorylation and F:G actin ratio, demonstrating the importance of the Rho/ROCK/LIMK pathway and actin reorganisation in cytoskeletal mechanisms of fibrogenesis. Supporting this, Pereira et al [ 7 ] examined the role of cocaine in vascular-endothelial dysfunction and reported the enhancement of leukocytic Rho-kinase activity in rat vascular walls, in vitro vascular cell ROCK activity and enhanced platelet adhesion upon treatment with plasma from cocaine abusers. Furthermore, a study by Pradhan et al [ 8 ] demonstrated that enhanced nitric oxide, endothelin-1 production, and in vitro cocaine exposure dependent monocyte adhesion potentiates the role of cocaine in regulating pathological development.…”

Metabolites of Cannabis Induce Cardiac Toxicity and Morphological Alterations in Cardiac Myocytes

“…Numerous studies have reported myocardial dysregulation following cannabis abuse, with myocardial infarction induced by vascular spasm and vasoconstriction being proposed as a mechanism of cannabis induced cardiovascular complications [7,36,66]. Chronic cannabis use has been linked to hepatotoxicity and structural alterations of the liver, with elevated liver markers consistent with hepatic sclerosis [67].…”

“…Cannabis sativa, commonly known as marijuana is one of the most widely used recreational drugs and has elicited significant interest within medical research for the treatment of neuropathic pain, epilepsy, and seizures [1][2][3][4][5][6][7][8]. In recent years, attitudes towards cannabis have evolved, owing to the large-scale decriminalization of medical and recreational cannabis use in countries such as the United States and Canada.…”

Cocaine as a Risk Factor for Heart and Vascular Disease. Part 1

“…При длительном употреблении кокаина продемонстрирована эндотелиальная дисфункция, в том числе в коронарных артериях, которая увеличивает чувствительность к катехоламинам, вызывает поражение эпикардиальных и мелких сосудов и может быть причиной боли в грудной клетке у лиц без значимого стеноза коронарных артерий, употребляющих кокаин. При этом в дополнение к симптоматическому лечению антиишемическими препаратами по поводу вазоконстрикции мелких сосудов может быть целесообразным использование антитромботических препаратов, поскольку кокаин обладает протромботической активностью, и ингибиторов RhoA/ Rho-киназы, которая играет ключевую роль в индуцированной кокаином эндотелиальной дисфункции [34,35].…”