The effect of endotoxin on hepatocyte nuclear factor 1 nuclear protein binding: potential implications on CYP2E1 expression in the rat.
The purpose of this study was to determine if changes in nuclear protein binding of hepatocyte nuclear factor 1 (HNF-1) occur after lipopolysaccharide (LPS) administration. In addition, the time-course of alterations in CYP2E1 regulation were evaluated. Rats were injected with 2.0 mg LPS and euthanized over a 72-h period. Nuclear protein binding to a consensus HNF-1 oligonucleotide was assessed by the electrophoretic mobility shift assay. CYP2E1 activity was analysed using chlorzoxazone as a substrate (60H-CLZ), and CYP2E1 protein concentration was determined by enzyme-linked immunosorbent assay. Endotoxin treatment resulted in decreased nuclear protein binding to an HNF-1 element as early as 1 h after treatment and returned to control levels by 72 h. This reduced binding persisted for 24 h and returned to control values 48 h after LPS administration. In addition, the reduction in binding was primarily attributable to a HNF-1alpha immunoreactive protein. The observed reduction in HNF-1 binding was followed in the time-course by decreases in CYP2E1 activity and protein content with maximal decreases to 50 and 67% of control, respectively, at 48 h after LPS administration. Endotoxin is a potent inducer of the acute phase response (APR). The APR stimulation by endotoxin administration reduced HNF-1alpha binding and decreased the expression of CYP2E1 in the rat liver. The time-course of alterations in HNF-1 and CYP2E1 lend support to the possibility that HNF-1alpha may play a role in the down-regulation of genes that require HNF-1alpha for their constitutive expression. These data serve as an important precedent for future studies evaluating the direct association of decreased HNF-1alpha binding and reduced gene expression after LPS administration.
Roe
A L
AL
Procter & Gamble Pharmaceuticals, Drug Safety Assessment, Health Care Research Center, Mason, OH 45040-9462, USA. roeal@pg.com
Poloyac
S M
SM
Howard
G
G
Shedlofsky
S I
SI
Blouin
R A
RA
D000818:Animals; D015153:Blotting, Western; D002467:Cell Nucleus / Q000737:chemistry; D019392:Cytochrome P-450 CYP2E1 / Q000096:biosynthesis*; D004268:DNA-Binding Proteins; D024202:Electrophoretic Mobility Shift Assay; D004731:Endotoxins / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D051539:Hepatocyte Nuclear Factor 1-beta; D008070:Lipopolysaccharides / Q000494:pharmacology; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D014157:Transcription Factors / Q000378:metabolism*
D004268:DNA-Binding Proteins; D004731:Endotoxins; D051538:Hepatocyte Nuclear Factor 1-alpha; C496489:Hnf1a protein, rat; D008070:Lipopolysaccharides; D009687:Nuclear Proteins; D014157:Transcription Factors; D051537:Hepatocyte Nuclear Factor 1; D051539:Hepatocyte Nuclear Factor 1-beta; C044101:endotoxin, Escherichia coli; D019392:Cytochrome P-450 CYP2E1