Hydrogen peroxide-dependent declines in Bcl-2 induces apoptosis in hypoxic liver.
BACKGROUND
Low-flow hypoxia induces xanthine oxidase-dependent hydrogen peroxide production by hepatocytes in the midzone of blood-perfused rat livers and apoptosis in sinusoidal endothelial cells (SECs). As Bcl-2 is a potent inhibitor of apoptotic cell death and is localized mainly in the inner mitochondrial membrane and crista, the purpose of this study was to determine whether cell-specific changes in mitochondrial Bcl-2 levels could account for the hypoxia-induced apoptosis in SECs.


METHODS
A low-flow hypoxia model was generated in isolated rat livers by reducing perfusate inflow pressure from 10 to 2.5 cmH2O for 2 h. Apoptosis was then evaluated using the TdT-mediated dUTP-digoxigenin nick end-labeling (TUNEL) method. Mitochondrial Bcl-2 protein levels were determined in hepatocytes and SECs using cryosectioning immunogold labeling electron microscopy.Results. TUNEL-positive nonparenchymal cells, identified as SECs, were observed predominantly in the midzone of low-flow hypoxic rat livers, whereas few parenchymal cells were stained. Mitochondrial Bcl-2 levels were higher in SECs than in hepatocytes under control conditions, but they declined significantly during hypoxia, though no morphological signs of apoptosis were apparent. In hepatocytes, by contrast, Bcl-2 levels were unaffected by hypoxia. Pretreatment with a specific xanthine oxidase inhibitor, sodium (-)-8-(3-methoxy-4-phenylsulfinylphenyl) pyrazolo [1,5-a]-1,3,5-triazine-4-olate monohydrate, which blocks production of hydrogen peroxide, also blocked both the hypoxia-induced apoptosis and the decline in mitochondrial Bcl-2 in SECs.


CONCLUSIONS
Hydrogen peroxide-dependent declines in Bcl-2 induce apoptosis in SECs in the hypoxic rat liver.
Motoyama
Satoru
S
The Second Department of Surgery, Akita University School of Medicine. 1-1-1 Hondo, 010-8543, Akita City, Japan. motoyama@doc.med.akita-u.ac.jp <motoyama@doc.med.akita-u.ac.jp>
Saito
Satoshi
S
Saito
Reijiro
R
Minamiya
Yoshihiro
Y
Nakamura
Masakatsu
M
Okuyama
Manabu
M
Imano
Hiroshi
H
Ogawa
Jun-ichi
J
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D022781:Hepatocytes / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000860:Hypoxia / Q000503:physiopathology*; D008099:Liver / Q000503:physiopathology*; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism; D016877:Oxidants / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014227:Triazines / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000037:antagonists & inhibitors
D016877:Oxidants; D019253:Proto-Oncogene Proteins c-bcl-2; D014227:Triazines; C072977:BOF 4272; D006861:Hydrogen Peroxide; D014969:Xanthine Oxidase