Abstract
The inducible form of heme oxygenase (HO-1) is increased during oxidative injury, and this may be an important defense mechanism against such injury. Cytochrome P450 2E1 (CYP2E1) generates reactive oxygen species and promotes lipid peroxidation. In this study induction of HO-1 by CYP2E1 and the possible role of mitogen-activated protein kinase (MAPK) in this process were evaluated. HO-1 induction was observed in the livers of chronic alcohol-fed mice or pyrazole-treated rats, conditions known to elevate CYP2E1 levels. Increased levels of HO-1 were observed in HepG2 cells overexpressing CYP2E1 (E47 cells) compared with control HepG2 cells or HepG2 cells expressing CYP3A4. Expression of CYP2E1 in HepG2 cells transcriptionally activated the HO-1 gene, increasing HO-1 mRNA and protein expression and activity of a HO-1 reporter construct. CYP2E1 inhibitors and catalase blocked the increased production of reactive oxygen species as well as HO-1 induction. Increasing oxidative stress by the addition of arachidonic acid or depletion of glutathione further increased HO-1 induction. The phosphorylated form of ERK MAPK but not that of p38 or JNK MAPK was increased in E47 cells compared with the control C34 HepG2 cells. PD98059, a specific inhibitor of ERK MAPK, blocked the activity of a HO-1 reporter in E47 cells but not in C34 cells. These results suggest that increased CYP2E1 activity leads to induction of the HO-1 gene, and the ERK MAPK pathway is important in mediating this process. This induction may serve as an adaptive mechanism to protect the E47 cells against the CYP2E1-dependent oxidative stress.
Highlights
Heme oxygenase (HO)1 is the rate-limiting enzyme in the conversion of heme into biliverdin, carbon monoxide (CO), and free iron (Fe2ϩ) [1]
We present data showing that HO-1 is transcriptionally activated in HepG2 cells overexpressing Cytochrome P450 2E1 (CYP2E1) and that the extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) pathway plays an important role in mediating this activation
HO-1 Protein Expression Is Induced in Liver of Mice Chronically Fed Ethanol— CYP2E1 is induced by ethanol, little is known about changes of HO-1 expression if any in the liver of animals treated with ethanol
Summary
Heme oxygenase (HO) is the rate-limiting enzyme in the conversion of heme into biliverdin, carbon monoxide (CO), and free iron (Fe2ϩ) [1]. CYP2E1-dependent toxicity in the presence of ethanol, arachidonic acid (AA), and AA plus iron has been characterized in these cell lines; these agents did not produce significant toxicity in C34 cells, whereas their addition to E47 cells decreased cell viability and caused either necrosis or apoptosis (14 –17) Toxicity by these agents was enhanced when cellular glutathione (GSH) levels were lowered by treatment with L-buthionine-(S,R)-sulfoximine (BSO). HO-1 Induction by CYP2E1 through ERK MAPK Pathway [18], whereas no toxicity was found in C34 cells or HepG2 cells that expressed CYP3A4 instead of CYP2E1 Antioxidants such as GSH-ethyl ester and N-acetylcysteine partially prevented the apoptosis and necrosis after BSO treatment, whereas diallyl sulfide, a CYP2E1 inhibitor, was fully protective. We present data showing that HO-1 is transcriptionally activated in HepG2 cells overexpressing CYP2E1 and that the ERK MAPK pathway plays an important role in mediating this activation
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