Acetaldehyde is a toxic metabolite of ethanol consumption. It can disrupt tight junctions between intestinal epithelial cells, allowing toxins into the body. This raises the risk of alcoholic liver disease and certain cancers. This disruption involves dephosphorylation of occludin and other tight junction proteins. PP2A, a serine/threonine phosphatase, is known to play a role in regulating tight junctional integrity. My preliminary data suggests that acetaldehyde destabilizes tight junctions and adherens junctions and promotes epithelial- to-mesenchymal transition by a PP2A-dependent mechanism. The long-range goal of this project is to elucidate the mechanisms involved in the ethanol-induced increase in risk for colon cancer. My first specific aim is to determine the role of PP2A and Thr-dephosphorylation of occludin, a tight junction protein, in acetaldehyde-induced disruption of tight junctions. This will be achieved by examining acetaldehyde-induced disruption of tight junctions with transepithelial resistance, inulin flux, and immunofluorescence confocal microscopy. Protection by fostriecin, a PP2A inhibitor, and by siRNA to knock down PP2A expression, will be used to examine the role of PP2A in disruption of tight junctions and in promotion of cell migration by acetaldehyde. My second specific aim is to determine that acetaldehyde induces epithelial-to-mesenchymal transition(EMT) by a PP2A-dependent mechanism. We will examine acetaldehyde-induced EMT by phase- contrast and immunofluorescence confocal microscopy as well as RT-PCR and Western blot to reveal changes in gene transcription and protein expression. We will use fostriecin and the generation and growth of occludin threonine mutants under 2D and 3D culture conditions to examine the roles of PP2A and threonine dephosphorylation in acetaldehyde-induced EMT.

Public Health Relevance

Acetaldehyde is a toxin produced in the colon when a person drinks alcoholic beverages. This toxin opens the tight junction, a barrier between intestinal cells that protects the body from substances in the colon. Disruption of the barrier increases the risk of liver disease, pancreatitis, and cancer, so I am studying how acetaldehyde breaks connections between tight junction proteins and how to keep these connections intact.

Agency
National Institute of Health (NIH)
Institute
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Type
Predoctoral Individual National Research Service Award (F31)
Project #
5F31AA018243-03
Application #
8120996
Study Section
Health Services Research Review Subcommittee (AA)
Program Officer
Jung, Kathy
Project Start
2009-09-17
Project End
2012-04-30
Budget Start
2011-09-17
Budget End
2012-04-30
Support Year
3
Fiscal Year
2011
Total Cost
$22,747
Indirect Cost
Name
University of Tennessee Health Science Center
Department
Physiology
Type
Schools of Medicine
DUNS #
941884009
City
Memphis
State
TN
Country
United States
Zip Code
38163
Dunagan, Mitzi; Chaudhry, Kamaljit; Samak, Geetha et al. (2012) Acetaldehyde disrupts tight junctions in Caco-2 cell monolayers by a protein phosphatase 2A-dependent mechanism. Am J Physiol Gastrointest Liver Physiol 303:G1356-64