The objective of this proposal is to determine the involvement of the lipid phosphatase PTEN in the formation of steatosis and liver damage during alcoholic liver disease (ALD). Although there are publications concerning the role of PTEN in ALD, these reports are using a static time point in a fluid model and do not address the possibility of both PTEN inactivation via aldehyde modification and changes in PTEN regulation/expression over a period of alcohol exposure. Our working hypothesis states the reactive aldehyde 4-HNE, produced in the liver of chronic ethanol treated mice covalently adducts PTEN thereby reducing enzymatic activity leading to disregulation in PTEN downstream signaling. 4-HNE has already been implicated in the inactivation of proteins during chronic ethanol exposure in the liver. It is also hypothesized that increased PTEN expression leads to changes in downstream pathways ultimately leading to increased steatosis seen in ALD. In order to fulfill the objectives of this research proposal, experiments will be performed using mass spectrometry to identify the sites of 4-HNE modification on PTEN, the ability of these adducts to inhibit enzymatic activity, membrane association and Trx1 association. In addition, using western blotting, immunohistochemistry and mRNA expression, the effects of variable PTEN expression on downstream signaling pathways will be examined following 9-weeks of chronic ethanol exposure.
In the United States today, alcohol induced liver disease is a major cause of morbidity and mortality. Current statistics indicate that among chronic alcohol users, 15% will eventually be diagnosed with alcoholic liver disease. These symptoms include steatosis, alcoholic hepatitis and cirrhosis. The 5 and 10 year survival rates for alcohol induced cirrhosis are 23% and 7% respectively. Completion of the proposed experiments will provide greater insight into the mechanisms of chronic alcohol toxicity especially with respect to steatosis and damage caused by increased oxidative stress and altered PTEN signaling.
|Shearn, Colin T; Mercer, Kelly E; Orlicky, David J et al. (2014) Short term feeding of a high fat diet exerts an additive effect on hepatocellular damage and steatosis in liver-specific PTEN knockout mice. PLoS One 9:e96553|
|Shearn, Colin T; Backos, Donald S; Orlicky, David J et al. (2014) Identification of 5' AMP-activated kinase as a target of reactive aldehydes during chronic ingestion of high concentrations of ethanol. J Biol Chem 289:15449-62|
|Shearn, Colin T; Smathers, Rebecca L; Stewart, Benjamin J et al. (2011) Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) inhibition by 4-hydroxynonenal leads to increased Akt activation in hepatocytes. Mol Pharmacol 79:941-52|
|Shearn, C T; Fritz, K S; Reigan, P et al. (2011) Modification of Akt2 by 4-hydroxynonenal inhibits insulin-dependent Akt signaling in HepG2 cells. Biochemistry 50:3984-96|