Elsevier

Neuropharmacology

Volume 50, Issue 5, April 2006, Pages 568-575
Neuropharmacology

Regional and cellular distribution of CYP2E1 in monkey brain and its induction by chronic nicotine

https://doi.org/10.1016/j.neuropharm.2005.11.001Get rights and content

Abstract

CYP2E1 is expressed in liver and extrahepatic tissues, including brain. It metabolizes ethanol and other drugs and toxins, such as acetaminophen, chlorzoxazone and tobacco-derived nitrosamines. Hepatic CYP2E1 is inducible by nicotine, and cigarette smoke accelerates chlorzoxazone metabolism. Smokers have higher levels of brain CYP2E1 expression than non-smokers, but the specific regions and cell types which have the higher expression differ from nicotine-induced rat brain. We therefore investigated the expression and distribution of brain CYP2E1 in a non-human primate, the African green monkey, and determined the effect of nicotine treatment. CYP2E1 levels varied among saline-treated monkey brain regions (P < 0.01) and expression was cell-type specific. Chronic nicotine treatment induced CYP2E1 expression in the frontal cortex (1.5-fold, P < 0.05) and cerebellum (1.5-fold, P < 0.01), specifically in cortical pyramidal neurons and cerebellar Purkinje cells but no change was seen in temporal cortex (P = 0.20), hippocampus (P = 0.29), putamen (P = 0.26) and thalamus (P = 0.08). CYP2E1 expression pattern in monkey brain following chronic nicotine treatment is similar to that in smokers, suggesting that nicotine may be the primary component in cigarette smoke that induces CYP2E1. Increased CYP2E1 in brain may contribute to oxidative stress and alter localized metabolism, and resulting pharmacology, of centrally acting drugs metabolized by CYP2E1.

Introduction

Cytochromes P450 (CYP) are mixed function oxidases that biotransform drugs, endogenous compounds, dietary constituents and environmental toxins (Lieber, 1999). These enzymes are expressed in the liver and also in other tissues, including lung, kidney and brain. Although CYP content in the central nervous system (CNS) is relatively low (Warner and Gustafsson, 1994), each CYP appears to be localized to specific regions and cell-types in the brain. It is unlikely that brain CYPs contribute substantially to overall clearance of xenobiotics but they may metabolize a variety of compounds increasing or decreasing their pharmacological effects within specific areas of the brain. Given their localized expression and sensitivity to environmental inducers, they may contribute to the observed interindividual variation in response to centrally acting drugs, their side effects and toxicities (Miksys and Tyndale, 2002).

Cytochrome P450 2E1 (CYP2E1), an ethanol-metabolizing enzyme, is also involved in the metabolism of drugs such as acetaminophen, chlorozoxazone, halothane as well as endogenous substrates such as arachidonic acid, fatty acids, gluconeogenic precursors and estrogenic metabolites (Lieber, 1999, Ohe et al., 2000). It also bioactivates procarcinogens (e.g. tobacco-derived nitrosamines and benzene) and cytotoxins (e.g. carbon tetrachloride) to their reactive intermediates (Lieber, 1999). Expression of human CYP2E1 in mammalian cell lines makes cells highly susceptible to cytotoxicity and DNA damage by nitrosamines (Lin et al., 1998, Lees Murdock et al., 2004). CYP2E1 can also generate reactive oxygen species and substrate-derived radicals that can mediate lipid peroxidation, protein inactivation and DNA damage, leading to cellular injury especially in the presence of CYP2E1 inducers (Cederbaum et al., 2001).

In rat brain, CYP2E1 expression was seen in pyramidal cells of frontal cortex, pyramidal and polymorphic cell layer in hippocampus and glial cells in olfactory bulb and piriform cortex (Hansson et al., 1990, Howard et al., 2003). In a pilot study, human CYP2E1 expression was detected in the brains of nonalcoholic nonsmokers in granular cells of the dentate gyrus, pyramidal cells of hippocampus and pyramidal neurons of frontal cortex (Howard et al., 2003).

Cigarette smoking accelerates chlorzoxazone metabolism in humans, most likely by induction of hepatic CYP2E1 activity (Benowitz et al., 2003). Rat hepatic CYP2E1 is inducible by chronic low doses of nicotine, comparable to levels of nicotine in environmental tobacco smoke exposure in humans (Howard et al., 2001, Micu et al., 2003). In addition, cigarette smoke appears to increase CYP2E1 levels in human brain. Higher CYP2E1 immunoreactivity was seen in the Purkinje cells of cerebellum, granular cells of the dentate gyrus, pyramidal cells of CA2 and CA3 hippocampal regions and pyramidal neurons in the frontal cortex of alcoholic smokers (Howard et al., 2003). Chronic nicotine treatment increases CYP2E1 expression in rat brain in a regiospecific manner (Anandatheerthavarada et al., 1993, Howard et al., 2003); however, the pattern of distribution of nicotine-induced CYP2E1 in the rat differs from that found in human brains from smokers. The differences in CYP2E1 brain expression suggest a species (rodent versus primate) or inducer difference (nicotine versus smoking). This could be a result of differences in regulatory factors or variation in brain structure/function between rodents and primates. Whether induction of primate CYP2E1 occurs due to nicotine in cigarette smoke is not known.

Our first objective was to investigate the regional and cellular distribution of CYP2E1 in African green monkey brain from saline-treated animals in order to obtain information about the basal expression of CYP2E1 in primate brain. The second objective was to examine whether chronic in vivo nicotine treatment induces CYP2E1 in the monkey brain in a regiospecific manner and to compare the distribution pattern in monkey brain to that in rat and human brain.

Part of this work has been published previously in the form of an abstract (Joshi et al., 2005).

Section snippets

Materials

Nicotine bitartrate was purchased from Sigma–Aldrich Canada Ltd. (Oakville, ON, Canada). The protein assay dye reagent concentrate was obtained from Bio-Rad (Hercules, CA, USA). Prestained molecular weight protein markers were purchased from MBI Fermentas (Flamborough, ON, Canada). Microsomes from CYP2E1-expressing lymphoblastoid cells and anti-human CYP2E1 antiserum used in immunocytochemistry were obtained from BD Gentest (Woburn, MA, USA). The rabbit anti-rat CYP2E1 polyclonal antibody used

CYP2E1 protein in monkey brain regions

In addition to endoplasmic reticulum, brain CYP proteins are expressed in several membranes types, including mitochondrial and plasma membranes (Miksys et al., 2000), therefore whole membrane preparations from monkey brain regions were assayed for CYP2E1 immunoreactivity. A quantitative immunoblotting assay was established to measure brain CYP2E1 levels. A dilution curve of membrane proteins from each brain region of a saline-treated monkey was constructed to determine the linear range of

Discussion

This is the first detailed report of the distribution of CYP2E1 among different brain regions and cell types in a primate, the African green monkey. CYP2E1 levels varied approximately 5-fold among the brain regions tested (ANOVA, P < 0.01). Variation between brain regions has also been seen in human brain expression of CYP2B6 (Miksys et al., 2003) and CYP2D6 (Miksys et al., 2002). Regiospecific and species-specific differences in the activities of CYPs have also been seen in rat and monkey lung (

Acknowledgements

The authors would like to thank Drs. Sharon Miksys and Roberta Palmour for assistance with the experiments and Dr. Magnus Ingelman-Sundberg for providing the CYP2E1 polyclonal antibody. This work was funded in part by the Canadian Institutes for Health Research (MOP14173), the Centre for Addiction and Mental Health, a Canada Research Chair (RFT) and the Canadian Society for Clinical Pharmacology and CIHR Strategic Training program in Tobacco Use in Special Populations (M.J.).

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