Overwhelming evidence suggests that oxidative-nitrosative/nitrative stress and inflammation are involved in essentially all major pathological processes affecting humans, including those induced by excessive alcohol consumption. The research focus of SOSTI is to understand the cellular and molecular mechanisms underlying oxidative/nitrosative/nitrative stress, inflammation, and their downstream effector pathways using clinically relevant animal models of disease (e.g. ischemia reperfusion injury, cardiomyopathy/heart failure), and to identify novel therapeutic targets against these pathologies. Interplay of oxidative/nitrative stress, inflammation with the endocannabinoid system (ES) in tissue injury. Recently we have been exploring the interplay of oxidative/nitrative stress, inflammation with the ES;an emerging very promising therapeutic target against various inflammatory and other diseases. Our previous studies provided evidence that the activation of cannabinoid 2 receptors (CB2) by selective receptor agonists exerts anti-inflammatory effects and attenuates oxidative/nitrative stress in models of liver, cardiovascular and renal injury. Recently, in collaborations with Dr. Juri Persidsky at Temple we tested the idea that selective CB2 activation in human monocytes suppresses their ability to engage the brain endothelium and migrate across the blood-brain barrier (BBB), preventing consequent injury. We found that CB2 agonists markedly decreased adhesion of ex vivo labeled cells in vivo. In an in vitro BBB model, CB2 activation in monocytes largely attenuated adhesion to and migration across monolayers of primary human brain microvascular endothelial cells and diminished BBB damage. CB2 stimulation in monocytes down-regulated active forms of integrins, lymphocyte function-associated antigen 1 (LFA-1), and very late antigen 4 (VLA-4). Cells treated with CB2 agonists exhibited increased phosphorylation levels of inhibitory sites of the actin-binding proteins cofilin and VASP, which are upstream regulators of conformational integrin changes. Up-regulated by relevant stimuli, Rac1 and RhoA were suppressed by CB2 agonists in monocytes. CB2 stimulation decreased formation of lamellipodia, which play a key role in monocyte migration. Collectively, these results indicate that selective CB2 activation in leukocytes decreases key steps in monocyteBBB engagement, thus suppressing inflammatory leukocyte responses and preventing neuroinflammation. Our upcoming studies will also be directed towards the development and characterization of novel CB2 receptor agonists with improved in vivo pharmacological profile, on better understanding the role of CB2 receptor function in various pathologies (e.g. neuroinflammation), and on development of improved tools for CB2 receptor detection in tissues/cells. We will also focus on the understanding of the mechanisms of the activation of the endocannabinoid system during tissue injury and on the further elucidation of the role of endocannabinoid system (particularly focusing on the endocannabinoid metabolizing enzymes in collaboration with Drs. Cravatt, Nomura and Kunos) in various models of liver disease, cardiomyopathy and nephropathy. Our future collaborative studies with Drs. George Kunos, Bin Gao and Byoung-Joon Song will also explore the role of oxidative/nitrosative stress and ES in various other models of liver and metabolic disorders and fibrosis. The above mentioned studies may identify new pharmacological targets in various forms of tissue injury associated with increased inflammation, oxidative stress, and fibrosis. Our collaborative studies with Dr. Gyorgy Hasko found that A2B AR deletion impaired glucose and lipid metabolism in mice fed chow but not a high-fat diet, which was paralleled by dysregulation of the adipokine system, and increased classical macrophage activation and inhibited alternative macrophage activation. Role of oxidative-nitrative stress, inflammation and apoptosis in ethanol-induced tissue-damage. Recently we have investigated the role of reactive oxygen/nitrogen species-nuclear enzyme poly (ADP-ribose) polymerase 1 (PARP-1) pathway in liver injury and fibrosis. We found increased nitrative stress and PARP activation in liver biopsies of subjects with alcoholic or hepatitis B-induced cirrhosis. Pharmacological inhibition of PARP with structurally distinct inhibitors or genetic deletion of PARP-1 markedly attenuated carbon tetrachloride (CCl4) -induced hepatocyte death, inflammation, and fibrosis. Interestingly, the chronic CCl4 -induced liver injury was also characterized by mitochondrial dysfunction and dysregulation of numerous genes involved in metabolism. Most of these pathological changes were attenuated by PARP inhibitors. PARP inhibition not only prevented CCl4 -induced chronic liver inflammation and fibrosis, but was also able to reverse these pathological processes. PARP inhibitors also attenuated the development of bile duct ligation (BDL)-induced hepatic fibrosis in mice. These results suggest that the reactive oxygen/nitrogen species-PARP pathway plays a key pathogenetic role in the development of liver inflammation, metabolism, and fibrosis. Furthermore, in addition to oncological indications, liver inflammation and liver fibrosis may be additional clinical indications where PARP inhibition may be of translational potential. Chronic excessive drinking may lead to the development of cardiomyopathy and potent immunomodulatory effects. Alcohol-mediated apoptosis of cardiomyocytes has been documented in experimental animals, and there is also evidence of skeletal muscle cell apoptosis in chronic heavy drinkers. The extent of apoptotic damage in the heart is similar in heavy drinkers and in patients with long-standing hypertension and is related to structural damage. Our recent ongoing studies are also focused on the understanding of the mechanisms of ethanol-induced oxidative/nitrative stress, inflammation and cell death in the cardiovascular system and also in the liver during pathological processes (e.g. associated with aging). Through collaboration with Drs. Emanuel Rubin and Gyorgy Hajnoczky at Dept. of Pathology, Thomas Jefferson University, we will also assess tissue samples from alcoholics to investigate the role of oxidative stress and inflammatory signaling in alcohol-induced myocardial and skeletal muscle injury.
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