Protection of rat hepatocytes from apoptosis by inhibition of c-Jun N-terminal kinase.
BACKGROUND
Apoptotic cell death and c-Jun N-terminal kinase (JNK) activation occur after hepatic ischemia/reperfusion injury. In other cell types, JNK activation was shown to be required for apoptosis. This study tested the hypotheses that JNK contributes to hepatocellular apoptosis, and that inhibition of JNK activity improves cell viability.


METHODS
Rat hepatocytes were harvested from Sprague-Dawley rats and pretreated with SP600125, a JNK inhibitor. Subsequently, they were exposed to apoptotic stimuli consisting of either the bile salt glycochenodeoxycholic acid (GCDC) or tumor necrosis factor (TNF)-alpha and actinomycin D.


RESULTS
Western blotting demonstrated specific inhibition of JNK by SP600125. Inhibition of JNK resulted in improved viability measured with crystal violet, decreased in situ DNA nick end labeling positivity, and decreased cleavage of poly (ADP-ribose) polymerase and caspase-3. TNF-alpha and actinomycin D induced apoptosis, upregulated p53, and downregulated expression of the anti-apoptotic protein X-linked inhibitor of apoptosis protein. These effects were abrogated by JNK inhibition.


CONCLUSIONS
These data show that pharmacologic inhibition of JNK activity reduces bile salt or TNF-alpha-induced apoptosis by maintaining expression of anti-apoptotic proteins. The results indicate that JNK is an important component of the apoptosis signaling cascade and suggest a possible therapeutic strategy in certain liver disorders.
Marderstein
Eric L
EL
Department of Surgery, University of Pittsburgh Medical Center, Pittsburgh, PA, USA.
Bucher
Brian
B
Guo
Zhong
Z
Feng
Xuesheng
X
Reid
Kaye
K
Geller
David A
DA
D000818:Animals; D000873:Anthracenes / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003609:Dactinomycin / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism / Q000502:physiology*; D048031:JNK Mitogen-Activated Protein Kinases; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D011506:Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D051636:X-Linked Inhibitor of Apoptosis Protein
D000873:Anthracenes; D004791:Enzyme Inhibitors; D011506:Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D014409:Tumor Necrosis Factor-alpha; D016159:Tumor Suppressor Protein p53; D051636:X-Linked Inhibitor of Apoptosis Protein; D003609:Dactinomycin; C432165:pyrazolanthrone; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases