Liver ASK1 activates autophagy to protect against hepatic fat accumulation, non-alcoholic steatohepatitis and fibrosis

Mon, 04/06/2020 - 13:57

MAP3K9 a MAP3K family member is expressed in the cytoplasm of human liver hepatocytes.

By Jamshed Arslan, Pharm. D., PhD.

The most common chronic liver disorder worldwide is non-alcoholic fatty liver disease (NAFLD). This obesity-linked disorder can manifest as hepatic fat accumulation (steatosis) with inflammation called non-alcoholic steatohepatitis (NASH). NASH can progress to fibrosis, cirrhosis and cancer. Stress conditions and impaired autophagy can contribute to steatosis and fibrosis. Stressors like TNF-α, reactive oxygen species and lipopolysaccharides can activate ASK1, which is a MAP3K family member that regulates autophagy by activating JNK and p38 MAPK. Focusing on NAFLD, a team of scientists in Switzerland and Germany set out to explore the role of liver-specific ASK1 in autophagy, steatosis, NASH and fibrosis. After studying human livers, cultured hepatocytes, and knockout and wild-type mice, they concluded that liver ASK1 suppresses steatosis, NASH and fibrosis, partly by activating autophagy.

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Liver ASK1 Ablation Promotes Hepatic Fat Accumulation, Inflammation and Fibrosis by Impairing Autophagy

To establish the link between liver ASK1 and NAFLD in humans, the researchers analyzed ASK1 mRNA    levels in people with or without diabetes. Liver ASK1 expression was negatively correlated with body-mass index and liver fat, but positively correlated with autophagy markers (ATG5, ATG7, ATG12). Likewise, individuals with higher NASH scores had reduced ASK1 expression, implying a protective role of ASK1 against NAFLD.

To elaborate on the contribution of autophagy, ASK1 was knocked down in cultured hepatocytes by siRNA. Western blotting indicated increased levels of p62 and LC3-II, which was a sign of blocked autophagy. A blunted lipophagy was evident from the increased colocalization of LC3 with fat droplets. This means that liver ASK1 deficiency and the consequent impaired autophagy can lead to NAFLD. The next step was to elucidate whether ASK1 overexpression could actively prevent the liver from NASH and fibrosis.

LC3B was detected by Western blot using LC3B antibody included in Autophagy rabbit polyclonal antibody pack, Novus Biologicals, in mouse embryonic stem cell lysates from ATG5 expressing and ATG5 knockout cells. ATG5 was detected by immunohistochemistry using ATG5 antibody included in Autophagy rabbit polyclonal antibody pack, Novus Biologicals, in human liver hepatocytes.

Autophagy antibody pack NB910-94159 for analysis of various significant autophagy targets includes sample sizes for antibodies targeting LC3 (Rabbit polyclonal anti-LC3B antibody, NB100-2220SS), Beclin1 (Rabbit polyclonal anti-Beclin1 antibody, NB110-87318SS), ATG5 (Rabbit polyclonal anti-ATG5 antibody, NB110-53818SS), ATG9A (Rabbit polyclonal anti-ATG9A antibody, NB110-56893SS), and ATG16L1 (Rabbit polyclonal anti-ATG16L1 antibody, NB110-60928SS). Left: LC3B was analyzed in mouse embryonic stem cell (ESC) lysates derived from control (ATG5+/+) or knockout (ATG5-/-) ESCs. Atg5-/- ES cells from Dr. Noboru Mizushima [Mizushima, N. et al. J. Cell Biol. 152 (2001)] Photo courtesy of Dr. Beth Levine, UT SW Medical Center. Right: Immunostaining of human liver hepatocytes with ATG5 antibody.

ASK1 Overexpression Protects Liver From Steatosis and Fibrosis

The team developed mice that overexpressed hepatic ASK1 to study the regulatory role of ASK1 in NAFLD. The benefit of ASK1 overexpression was profound. For example, these mice had improved glucose tolerance and autophagy as well as reduced liver triglycerides and lipid droplets after 20 weeks of a coconut oil-containing high-fat diet, relative to the controls. ASK1 overexpression protected the mice even from carbon tetrachloride-induced fibrosis, as evidenced from diminished Sirius Red staining for collagen deposition. Likewise, mRNA levels of genes involved in inflammation and fibrosis (Col1a1, Tgfb1) were also reduced in these mice. Overall, the data indicate that liver ASK1 protects against various manifestations of NAFLD, partly by activating autophagy.

Significance of Liver ASK1 Against NAFLD

Pharmacological therapies against liver fibrosis are lacking. By highlighting the protective role of hepatic ASK1 in steatosis, NASH and fibrosis, this study provides a unique therapeutic target in the fight against NAFLD and its complications.

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Jamshed Arslan Jamshed Arslan, Pharm D, PhD   
Dr Arslan is an Assistant Professor at Barrett Hodgson University, Pakistan,
where he uses various pedagogical methods to teach Pharm D students.

Research in focus

Challa, T. D., Wueest, S., Lucchini, F. C., Dedual, M., Modica, S., Borsigova, M., … Konrad, D. (2019). Liver ASK1 protects from non‐alcoholic fatty liver disease and fibrosis. EMBO Molecular Medicine.


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