Heat shock response inhibits cytokine-inducible nitric oxide synthase expression in rat hepatocytes.
During sepsis or inflammation, the liver expresses various protective phenotypes such as the acute phase response or the heat shock response (HSR). Inducible nitric oxide synthase (NOS2) is also expressed in the liver in these conditions and may protect the liver under some circumstances and promote injury in others. We have previously reported that the acute phase response and NOS2 expression are differentially regulated, though both can be expressed simultaneously. The HSR is known to prevent expression of other genes, but its effects on NOS2 expression in the liver is unknown. Therefore, we examined how the HSR influences NOS2 expression in primary rat hepatocytes. Sodium arsenite (Ars) or hyperthermia (43 degrees C) induced the synthesis of hsp72 messenger RNA (mRNA) and protein in hepatocytes, indicating activation of the HSR. In the absence of the HSR, combinations of interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNF-alpha), and interferon gamma (IFN-gamma) stimulated high levels of NOS2 mRNA and nitric oxide (NO) synthesis. However, treatment with Ars or heat shock significantly attenuated cytokine-induced NOS2 mRNA and NO production. The addition of the nuclear factor kappaB (NF-kappaB) inhibitor pyrrolidine dithiocarbamate also inhibited NOS2 expression, suggesting a role for NF-kappaB in the cytokine induction of NOS2 in hepatocytes. Cytokines induced the appearance of an NF-kappaB complex as shown in gel retardation assays; however, induction of the HSR by Ars partially prevented cytokine-induced formation of this band while hyperthermia had a more complete inhibition. Furthermore, preinduction of the HSR prevented the activation of the NOS2 promoter construct in hepatocytes transfected with a 1.6 kilobase NOS2 promoter linked to luciferase. These findings show that NO production in stressed cells can be modulated by the HSR, possibly through repression of NOS2 gene transcription via the inhibition of NF-kappaB.
de Vera
M E
ME
Department of Surgery, University of Pittsburgh, PA, USA.
Kim
Y M
YM
Wong
H R
HR
Wang
Q
Q
Billiar
T R
TR
Geller
D A
DA
D000818:Animals; D018053:Arsenites / Q000494:pharmacology; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology*; D004247:DNA / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D006360:Heat-Shock Proteins / Q000096:biosynthesis*; D006358:Hot Temperature; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D008297:Male; D016328:NF-kappa B / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley
D018053:Arsenites; D016207:Cytokines; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D016328:NF-kappa B; D012333:RNA, Messenger; D004247:DNA; D019001:Nitric Oxide Synthase; C015001:arsenite