Induction of inducible nitric oxide synthase in hepatocytes isolated from rats with ischemia-reperfusion injury.
BACKGROUND
Recent evidence indicates that nitric oxide (NO) has a crucial role in hepatic ischemia-reperfusion (I/R) injury. However, little is known about how I/R influences the gene expression of inducible nitric oxide synthase (iNOS) in hepatocytes. Under inflammatory conditions, we compared the induction of iNOS in hepatocytes isolated from normal and I/R-treated rats.


METHODS
Hepatocytes were isolated using the collagenase perfusion method from rats treated with I/R (30-minute ischemia of middle and left lobes, followed by 3-hour reperfusion) or sham operation (control): Primary cultures of rat hepatocytes were incubated with an inflammatory cytokine, interleukin-1beta (IL-1beta), to compare the iNOS induction/NO production between the 2 groups.


RESULTS
Both control and I/R groups had no production of nitrite (a stable metabolite of NO) in the absence of IL-1beta. In the control group, IL-1beta stimulated dose- and time-dependent production of NO. The I/R group showed more than 2-fold increased levels of NO production. Western and Northern blot analyses revealed that the I/R group also showed increased levels of iNOS protein and its messenger RNA.


CONCLUSIONS
These results suggest that I/R directly affects the inducibility of the iNOS gene in hepatocytes by IL-1beta. Increased NO may be associated with protective or toxic effects in hepatic I/R injury.
Yanagida
H
H
Department of Surgery, Kansai Medical University, Osaka, Japan.
Kaibori
M
M
Yamada
M
M
Habara
K
K
Yokoigawa
N
N
Kwon
A-H
AH
Kamiyama
Y
Y
Okumura
T
T
D000818:Animals; D002478:Cells, Cultured; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D022781:Hepatocytes / Q000201:enzymology*; D007375:Interleukin-1 / Q000494:pharmacology; D008102:Liver Circulation; D008297:Male; D019001:Nitric Oxide Synthase / Q000096:biosynthesis / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D015427:Reperfusion Injury / Q000201:enzymology* / Q000473:pathology
D007375:Interleukin-1; D012333:RNA, Messenger; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat