Preservation of hepatocyte nuclear factor-4alpha is associated with zinc protection against TNF-alpha hepatotoxicity in mice.
Hepatocyte nuclear factor-4alpha (HNF-4alpha), a zinc finger protein, is the most abundant transcription factor in the liver. HNF-4alpha regulates a large number of genes involved in most aspects of hepatocyte functions. The present study was undertaken to determine the role of HNF-4alpha in zinc protection against tumor necrosis factor-alpha (TNF-alpha) hepatotoxicity. Mice were treated with murine TNF-alpha via intravenous injection at 20 mug/kg body wt 30 mins after d-galactosamine (d-Gal) sensitization (800 mg/kg body wt). Two doses of zinc sulfate (5 mg elemental zinc/kg body wt) were administered at 36 and 12 hrs before TNF-alpha treatment via subcutaneous injection. TNF-alpha treatment after sensitization induced liver injury as detected by plasma alanine aminotransferase activity and apoptotic cell death in the liver. Zinc pretreatment attenuated TNF-alpha-induced liver injury. Furthermore, TNF-alpha-induced activations of caspase 3 and caspase 8 in the liver were significantly inhibited by zinc pretreatment. The mRNA and protein levels of HNF-4alpha in the liver were remarkably decreased by TNF-alpha treatment, which was suppressed by zinc. To determine if HNF-4alpha depletion is involved in d-Gal sensitization to TNF-alpha toxicity, mice were administered either d-Gal or TNF-alpha. Immunohistochemistry demonstrated that HNF-4alpha depletion in the liver is associated with d-Gal sensitization but not TNF-alpha treatment. To define the link between HNF-4alpha depletion and TNF-alpha-induced cell death, the effect of silencing the HNF-4alpha gene by siRNA transfection on TNF-alpha cytotoxicity was determined in HepG2 cells. A lactate dehydrogenase cytotoxicity assay showed that neither TNF-alpha nor HNF-4alpha siRNA transfection had a toxic effect, but TNF-alpha treatment after HNF-4alpha siRNA transfection caused HepG2 cell death. These results suggest that zinc protects against TNF-alpha hepatotoxicity, at least partially, through preservation of the zinc finger protein HNF-4alpha.
Zhou
Zhanxiang
Z
Department of Medicine, University of Louisville School of Medicine, Louisville, KY 40202, USA. z0zhou01@louisville.edu
Kang
Xinqin
X
Jiang
Youchun
Y
Song
Zhenyuan
Z
Feng
Wenke
W
McClain
Craig J
CJ
Kang
Y James
YJ
D000410:Alanine Transaminase / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D056486:Chemical and Drug Induced Liver Injury; D051557:Hepatocyte Nuclear Factor 4 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D007275:Injections, Intravenous; D007279:Injections, Subcutaneous; D008099:Liver / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008107:Liver Diseases / Q000517:prevention & control; D051379:Mice; D008815:Mice, Inbred Strains; D020011:Protective Agents / Q000494:pharmacology; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000008:administration & dosage / Q000633:toxicity*; D015032:Zinc / Q000008:administration & dosage / Q000494:pharmacology*; D019287:Zinc Sulfate / Q000008:administration & dosage / Q000494:pharmacology
D051557:Hepatocyte Nuclear Factor 4; D020011:Protective Agents; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D019287:Zinc Sulfate; D000410:Alanine Transaminase; D053148:Caspase 3; D053181:Caspase 8; D015032:Zinc