A DNA microarray study of nitric oxide-induced genes in mouse hepatocytes: implications for hepatic heme oxygenase-1 expression in ischemia/reperfusion.
Nitric oxide (NO) can modulate numerous genes directly; however, some genes may be modulated only in the presence of the inflammatory stimuli that increase the expression of the inducible nitric oxide synthase (iNOS). One method by which to examine changes in NO-mediated gene expression is to carry out a gene array analysis on NO-nai;ve cells. Herein, we report a gene array analysis on mRNA from iNOS-null (iNOS(-/-)) mouse hepatocytes harvested from mice exposed to NO by infection with an adenovirus expressing human iNOS (Ad-iNOS). Of the 6500 genes on this array, only approximately 200 were modulated either up or down by the increased iNOS activity according to our criteria for significance. Several clearly defined families of genes were modulated, including genes coding for proinflammatory transcription factors, cytokines, cytokine receptors, proteins associated with cell proliferation and cellular energetics, as well as proteins involved in apoptosis. Our results suggest that iNOS has a generally anti-inflammatory and anti-apoptotic role in hepatocytes but also acts to suppress proliferation and protein synthesis. The expression of iNOS results in increased expression of stress-related proteins, including heme oxygenase-1 (HO-1). We used HO-1 to confirm that a significant change identified by an analysis could be demonstrated as significant in cells and tissues. The elevation of HO-1 was confirmed at the protein level in hepatocytes in vitro. Furthermore, iNOS(-/-) mice experienced greatly increased liver injury subsequent to intestinal ischemia/reperfusion injury, associated with an inability to upregulate HO-1. This is the first study to address the global gene changes induced by iNOS in any cell type, and the findings presented herein may have clinical relevance for conditions such as septic or hemorrhagic shock in which hepatocytes, NO, and HO-1 play a crucial role.
Zamora
Ruben
R
Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15261, USA. zamorar@pitt.edu
Vodovotz
Yoram
Y
Aulak
Kulwant S
KS
Kim
Peter K M
PK
Kane
John M
JM
Alarcon
Louis
L
Stuehr
Dennis J
DJ
Billiar
Timothy R
TR
D000818:Animals; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D006419:Heme Oxygenase (Decyclizing) / Q000096:biosynthesis; D051547:Heme Oxygenase-1; D022781:Hepatocytes / Q000201:enzymology* / Q000378:metabolism / Q000473:pathology; D008565:Membrane Proteins; D051379:Mice; D018345:Mice, Knockout; D009569:Nitric Oxide / Q000235:genetics*; D019001:Nitric Oxide Synthase / Q000235:genetics* / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D020411:Oligonucleotide Array Sequence Analysis; D015427:Reperfusion Injury / Q000201:enzymology; D014162:Transfection
D008565:Membrane Proteins; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D006419:Heme Oxygenase (Decyclizing); D051547:Heme Oxygenase-1; C496542:Hmox1 protein, mouse