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EGF-Induced receptor autophosphorylation in primary hepatocytes isolated from phenobarbitone-treated mice
K Fletcher1, P G Lord, T C Orton
1School of Biochemistry, University of Birmingham, Edgbaston, B15 2TT, United Kingdom.
Abstract:
Phenobarbitone (PB) treatment of mice causes a decrease in the growth factor responsiveness of hepatocytes. Here, epidermal growth factor receptor (EGFR) expression and receptor autophosphorylation was determined in hepatocytes isolated from control and PB-treated mice. There was a decrease in the level of EGFR expression in hepatocytes isolated from mice following PB administration when compared to controls. EGF caused an approximate 20-fold increase of the 170 kD phosphotyrosine band in control hepatocytes, which was inhibited by the EGFR specific tyrosine kinase inhibitor 4, 5-dianilinopthalamide. Following PB treatment, the degree of basal receptor phosphorylation (in the absence of EGF) was significantly greater and therefore the fold rise in EGFR phosphorylation in isolated hepatocytes was lower than in controls. However, the overall extent of EGF-induced receptor phosphorylation was not diminished in hepatocytes isolated from PB-treated mice. Therefore the reduction in responsiveness to growth factors seen in hepatocytes ex vivo or the cessation of proliferation observed in vivo following PB administration is unlikely to be attributed to a decrease in ligand binding and subsequent receptor autophosphorylation.
Insights
Phenobarbitone (PB) treatment reduces epidermal growth factor receptor (EGFR) expression in mouse hepatocytes. However, EGF-induced receptor phosphorylation remains intact, suggesting other mechanisms reduce growth factor responsiveness.
Area of Science:
- Hepatology
- Molecular Biology
- Pharmacology
Background:
- Phenobarbitone (PB) treatment in mice is known to decrease growth factor responsiveness in hepatocytes.
- The precise mechanisms underlying this reduced responsiveness require further investigation.
Purpose of the Study:
- To investigate the effects of PB treatment on epidermal growth factor receptor (EGFR) expression and autophosphorylation in mouse hepatocytes.
- To determine if alterations in EGFR signaling contribute to the reduced growth factor responsiveness observed after PB administration.
Main Methods:
- Hepatocytes were isolated from control and PB-treated mice.
- EGFR expression levels were quantified.
- Receptor autophosphorylation in response to epidermal growth factor (EGF) was assessed, with and without an EGFR-specific tyrosine kinase inhibitor.
Main Results:
- PB treatment led to a significant decrease in EGFR expression in hepatocytes.
- Basal EGFR phosphorylation (without EGF) was elevated in PB-treated hepatocytes.
- Despite lower fold-increase due to higher basal levels, the overall extent of EGF-induced EGFR phosphorylation was not diminished in PB-treated hepatocytes compared to controls.
Conclusions:
- The reduction in growth factor responsiveness and proliferation cessation following PB administration is unlikely due to decreased EGFR expression or impaired EGF-induced receptor autophosphorylation.
- Alternative molecular pathways may be responsible for the observed diminished responsiveness to growth factors in PB-treated hepatocytes.