A hedgehog survival pathway in 'undead' lipotoxic hepatocytes.
OBJECTIVE
Ballooned hepatocytes in non-alcoholic steatohepatitis (NASH) generate sonic hedgehog (SHH). This observation is consistent with a cellular phenotype in which the cell death program has been initiated but cannot be executed. Our aim was to determine whether ballooned hepatocytes have potentially disabled the cell death execution machinery, and if so, can their functional biology be modeled in vitro.


METHODS
Immunohistochemistry was performed on human NASH specimens. In vitro studies were performed using HuH-7 cells with shRNA targeted knockdown of caspase 9 (shC9 cells) or primary hepatocytes from caspase 3(-/-) mice.


RESULTS
Ballooned hepatocytes in NASH display diminished expression of caspase 9. This phenotype was modeled using shC9 cells; these cells were resistant to lipoapoptosis by palmitate (PA) or lysophosphatidylcholine (LPC) despite lipid droplet formation. During lipid loading by either PA or LPC, shC9 cells activate JNK which induces SHH expression via AP-1. An autocrine hedgehog survival signaling pathway was further delineated in both shC9 and caspase 3(-/-) cells during lipotoxic stress.


CONCLUSIONS
Ballooned hepatocytes in NASH downregulate caspase 9, a pivotal caspase executing the mitochondrial pathway of apoptosis. Hepatocytes engineered to reduce caspase 9 expression are resistant to lipoapoptosis, in part, due to a hedgehog autocrine survival signaling pathway.
Division of Gastroenterology and Hepatology, College of Medicine, Mayo Clinic, Rochester, MN 55905, United States.
Keisuke
K
Kakisaka
Sophie C
SC
Cazanave
Nathan W
NW
Werneburg
Nataliya
N
Razumilava
Joachim C
JC
Mertens
Steve F
SF
Bronk
Gregory J
GJ
Gores
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D020170:Caspase 1 / Q000235:genetics; D053453:Caspase 9 / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D015536:Down-Regulation; D005234:Fatty Liver / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D053823:Hedgehog Proteins / Q000235:genetics / Q000378:metabolism*; D022781:Hepatocytes; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008244:Lysophosphatidylcholines / Q000494:pharmacology; D008915:Minute Virus of Mice; D065626:Non-alcoholic Fatty Liver Disease; D010168:Palmitates / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering; D015398:Signal Transduction; D018808:Transcription Factor AP-1 / Q000378:metabolism
D000590:Amino Acid Chloromethyl Ketones; D053823:Hedgehog Proteins; D008244:Lysophosphatidylcholines; D010168:Palmitates; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C509239:SHH protein, human; D018808:Transcription Factor AP-1; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D048031:JNK Mitogen-Activated Protein Kinases; D053453:Caspase 9; D020170:Caspase 1