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Rasineni K, Donohue TM, Thomes PG, et al. Ethanol-induced steatosis involves impairment of lipophagy, associated with reduced Dynamin2 activity. Hepatol Commun. 2017;1(6):501-512doi: 10.1002/hep4.1063.
Rasineni, K., Donohue, T. M., Thomes, P. G., Yang, L., Tuma, D. J., McNiven, M. A., & Casey, C. A. (2017). Ethanol-induced steatosis involves impairment of lipophagy, associated with reduced Dynamin2 activity. Hepatology communications, 1(6), 501-512. https://doi.org/10.1002/hep4.1063
Rasineni, Karuna, et al. "Ethanol-induced steatosis involves impairment of lipophagy, associated with reduced Dynamin2 activity." Hepatology communications vol. 1,6 (2017): 501-512. doi: https://doi.org/10.1002/hep4.1063
Rasineni K, Donohue TM, Thomes PG, Yang L, Tuma DJ, McNiven MA, Casey CA. Ethanol-induced steatosis involves impairment of lipophagy, associated with reduced Dynamin2 activity. Hepatol Commun. 2017 Aug;1(6):501-512. doi: 10.1002/hep4.1063. Epub 2017 Jul 10. PMID: 29152606; PMCID: PMC5678901.
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Hepatol Commun. 2017 Aug;1(6):501-512. doi: 10.1002/hep4.1063. Epub 2017 Jul 10.

Ethanol-induced steatosis involves impairment of lipophagy, associated with reduced Dynamin2 activity.

Hepatology communications

Karuna Rasineni, Terrence M Donohue, Paul G Thomes, Li Yang, Dean J Tuma, Mark A McNiven, Carol A Casey

Affiliations

  1. The Liver Study Unit, VA Nebraska-Western Iowa Health Care System (VA NWIHCS).
  2. Department of Internal Medicine, University of Nebraska Medical Center.
  3. Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center.
  4. Pathology and Microbiology; College of Medicine; University of Nebraska Medical Center.
  5. The Center for Environmental Toxicology; College of Public Health, University of Nebraska Medical Center Omaha, NE, USA.
  6. Tongji Hospital, Tongji University School of Medicine, Shanghai, China.
  7. Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, Rochester, Minnesota.

PMID: 29152606 PMCID: PMC5678901 DOI: 10.1002/hep4.1063

Abstract

BACKGROUND: Lipid droplets (LDs), the organelles central to alcoholic steatosis, are broken down by lipophagy, a specialized form of autophagy. Here, we hypothesize that ethanol administration retards lipophagy by down-regulating Dynamin 2 (Dyn2), a protein that facilitates lysosome re-formation, contributing to hepatocellular steatosis.

METHODS: Primary hepatocytes were isolated from male Wistar rats fed Lieber-DeCarli control or EtOH liquid diets for 6-8 wk. Hepatocytes were incubated in complete medium (fed) or nutrient-free medium (fasting) with or without the Dyn2 inhibitor Dynasore or the Src inhibitor SU6656. Phosphorylated (active) forms of Src and Dyn2, and markers of autophagy were quantified by Western Blot. Co-localization of LDs-with autophagic machinery was determined by confocal microscopy.

RESULTS: In hepatocytes from pair-fed rats, LD breakdown was accelerated during fasting, as judged by smaller LDs and lower TG content when compared to hepatocytes in complete media. Fasting-induced TG loss in control hepatocytes was significantly blocked by either SU6656 or Dynasore. Compared to controls, hepatocytes from EtOH-fed rats had 66% and 40% lower content of pSrc and pDyn2, respectively, coupled with lower rate of fasting-induced TG loss. This slower rate of fasting-induced TG loss was blocked in cells co-incubated with Dynasore. Microscopic examination of EtOH-fed rat hepatocytes revealed increased co-localization of the autophagosome marker LC3 on LDs with a concomitant decrease in lysosome marker LAMP1. Whole livers and LD fractions of EtOH-fed rats exhibited simultaneous increase in LC3II and p62 over that of controls, indicating a block in lipophagy.

CONCLUSION: Chronic ethanol administration slowed the rate of hepatocyte lipophagy, owing in part to lower levels of phosphorylated Src kinase available to activate its substrate, Dyn2, thereby causing depletion of lysosomes for LD breakdown.

Keywords: Alcoholic fatty liver; Dyn2; autophagy; lipid droplets; lipophagy; lysosomes

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