Alcohol is the most commonly used and abused drug in the United States. Deleterious health effects of alcohol attribute to toxicity of internal organs including irreversible brain tissue injury. Brain tissue from chronic alcoholics exhibits neuronal cell death and degeneration paralleling neuro-cognitive deficits. Alcohol abuse leads to white matter abnormalities that could be associated with blood brain barrier (BBB) dysfunction detected in alcohol abusers or seen in animal model. The BBB formed by brain microvascular endothelial cells (BMVEC), pericytes and astrocytes connect the BMVEC assuring BBB structural tightness. The actin cytoskeleton is linked to tight junctions (TJ) proteins through intracellular zonula occludens (ZO-1-3) and this dynamic complex readily adapts to a variety of physiologic or pathologic conditions. The intracellular signaling processes that regulate phosphorylation of TJ proteins control TJ assembly and BBB integrity. The exact mechanism of BBB dysfunction seen in alcohol abusers remains elusive. We demonstrated the previously unrecognized effects of ethanol (EtOH) on BMVEC, underlying the cause of BBB impairment. We showed that EtOH increases the activity and content of EtOH-metabolizing enzymes, cytochrome P450-2E1 (CYP2E1) and alcohol dehydrogenase in human BMVEC. EtOH metabolism by CYP2E1/ADH in BMVEC enhances production of acetaldehyde (AA) and reactive oxygen species (ROS). We propose that these reactive EtOH metabolites (AA and ROS) activate myosin light chain kinase (MLCK) via IP3R-gated intracellular calcium signaling pathway resulting in phosphorylation of cytoskeletal and TJ proteins as acute effect. Further, the reactive EtOH metabolites activate matrix metalloproteinases -2 and -9 (MMP-2 and -9) via protein tyrosine kinase (PTK) signaling pathway leading to degradation of endothelial specific basement membrane (BM) or extracellular matrix (ECM) proteins. Changes in cytoskeletal and TJ assembly, or degradation of BM/ECM proteins increased BBB permeability and augmented leukocyte migration across the BBB, suggesting break down of BBB function. BBB dysfunction due to alcohol abuse could be associated with in neuro-inflammatory disorders and neurological diseases. Thus, we propose to study the mechanism of alcohol abuse disruption of blood-brain barrier due to oxidative stress stemming from alcohol metabolism in brain endothelial cells, which activates myosin light chain kinase through intracellular calcium release (acute effects) and activation of matrix metalloproteinases via PTK signaling pathway (delayed effects).

National Institute of Health (NIH)
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Exploratory/Developmental Grants (R21)
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Health Services Research Review Subcommittee (AA)
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Regunathan, Soundar
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University of Nebraska Medical Center
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