Endocrine Abstracts

Liver autophagy, as assessed by accumulation of LC3-II and p62 proteins, is positively correlated with the severity of nonalcoholic steatohepatitis (NASH) in humans. Since growth hormone (GH) is negatively associated with liver steatosis and NASH development, and our laboratory has reported that steatosis and NASH develops in a mouse model of adult-onset, hepatocyte-specific GH-Receptor knockdown (aHepGHRkd), we sought to determine if GH directly regulates the hepatocyte autophagy program, and whether this regulation is mediated via STAT5B activation. To this end, adult GHRfl/fl male mice were treated with adeno-associated viral vectors to generate GHR-intact control (AAV8-TBGp-Null), aHepGHRkd (AAV8-TBGp-Cre) or aHepGHRkd mice with constitutive activation of STAT5B (AAV8-TBGp-Cre + AAV8-TBGp-Stat5bCA). Livers were analyzed 7 days post AAV injection, a time when steatosis is observed in aHepGHRkd mice. We first evaluated the level of p62 and LC3-II proteins in liver lysosomal fractions by western blot, from mice euthanized at 1800h (end of the normal sleep cycle, natural fasting), a time of enhanced autophagy. In this condition, we did not observe any differences in p62 or LC3-II protein levels between control, aHepGHRkd or aHepGHRkd+Stat5bCA. Since p62 and LC3-II are involved in the initiation of autophagy, but are rapidly cleared in the autolysosome, we repeated the study after treating the mice with leupeptin, that blocks Cathepsin B, H and L and impairs amphysome-lysosome fusion, leading to a build-up of p62 and LC3-II. In this scenario, we observed that aHepGHRkd resulted in an increase in accumulation of p62 and LC3-II, compared with GHR-intact controls, and Stat5bCA partially reversed this effect. Current literature suggests that autophagy increases in the steatotic liver to protect against NASH progression. Therefore it is possible that the increase in autophagy observed in aHepGHRkd mice is not directly due to loss of GHR signaling but secondary to the development of steatosis, since Stat5bCA also reduces steatosis in aHepGHRkd mice. Therefore, although additional studies are required to determine if GHR/STAT5 plays a direct role in regulating the autophagy process in the liver, our data suggest this remodeling process could be altered in the context of GH signaling alteration during NAFLD development.

Funding: This work was funded by NIH R01DK114326 and VA Merit BX001114 (to RDK)