Comparison of benzo(a)pyrene metabolism in isolated perfused rat lung and liver

The metabolism of 1 mM benzo(a)pyrene was studied in isolated perfused lung and liver of 5,6-benzoflavone-pretreated rats. Benzo(a)pyrene metabolism by the liver was more rapid than by the lung, but total metabolite formation in the lung at the end of a 120-min perfusion period was comparable to that in the liver. Lung perfusate was characterized by high concentrations of free metabolites, with diols outweighing phenols; in liver perfusate free metabolite concentrations were low, and large quantities of metabolites were found as conjugates in the bile at the end of perfusion. The tissue concentrations of free diols and phenols including the precursors of the main DNA-binding secondary metabolites were higher in the lung than in the liver. These findings explain the similar level of covalent binding in perfused lung and liver previously described (Klaus et al. 1982).Abbreviations Used BP benzo(a)pyrene - 9,10-diol 9,10-dihydro9,10-dihydroxy-benzo(a)pyrene - 4,5-diol 4,5-dihydro-4,5-dihydroxy-benzo(a)pyrene - 7,8-diol 7,8-dihydro-7,8-dihydroxy-benzo(a)pyrene - 9-OH 9-hydroxy-benzo(a)pyrene - 3-OH 3-hydroxybenzo(a)pyrene - tetrols 7,8,9,10-tetrahydro-7,8,9,10-tetrahydroxy-benzo(a)pyrenes - BF 5,6-benzoflavone - TLC thin-layer chromatography - HPLC high-pressure liquid chromatography     

Influence of pretreatment with tobacco smoke condensate or 3-methylcholanthrene on [14C] benzo (A) pyrene metabolism in the isolated perfused rabbit lung

Pretreatment 24 h before sacrifice with i.p. tobacco smoke condensate (TSC) or 3-methylcholanthrene (3MC) increased the rate of disappearance of [¹⁴C]benzo(a)pyrene (BP) from an isolated perfused rabbit lung model. Both pretreatments significantly increased the amount of most metabolites formed. This study indicates that rabbits tend to resemble rats, mice and hamsters in that increased rates of pulmonary BP metabolism are a consequence of exposure to TSC.     

The distribution of benzo(a)pyrene and its metabolites in isolated perfused lung and liver

Distribution of benzo(a)pyrene (BP) and its metabolites in an isolated perfused lung was compared with distribution in an isolated perfused liver. The uptake of BP into the lung was slower than into the liver. A methylcholanthrene (MC)-induced lung metabolized BP as efficiently as MC-induced liver. The lung seemed to accumulate BP-metabolites while the liver excreted them into the bile and perfusion medium. It is suggested that distribution of metabolites may be significant in determining the carcinogenic effect of BP in various tissues.     

Effect of benzo(a)pyrene exposure on fluoranthene metabolism by mouse adipose tissue microsomes

The present study has been undertaken to examine whether exposure to benzo(a)pyrene (BaP), a polycyclic aromatic hydrocarbon (PAH) compound, influences the metabolism of fluoranthene (FLA), another PAH compound. Microsomes were isolated from the adipose tissue of mice that received 50?μg/kg BaP and incubated with FLA (3?µM) alone; FLA in combination with BaP at equimolar concentrations, and a control group that received nothing. Post-incubation, samples were extracted with ethyl acetate and analyzed for FLA metabolites by reverse-phase HPLC with fluorescence detection. The rate of FLA metabolism (pmol of metabolite/min/mg protein) was increased when microsomes from BaP-treated mice were exposed to FLA alone and FLA in combination with BaP, compared to controls. On the other hand, the difference in FLA metabolic rate between microsomes that were exposed to FLA + BaP was higher than the ones that received FLA. The microsomes from BaP-pre-treated mice produced a considerably higher proportion of FLA 2, 3-diol, and 2, 3 D FLA when microsomes were incubated with FLA. There were no differences in the FLA metabolite types formed when BaP-pre-treated mice were co-incubated with BaP and FLA than with FLA alone. The enhanced biotransformation of FLA as a result of prior and concomitant exposure to BaP may have implications for assessment of risks arising from human exposure to PAH mixtures.     

Aryl hydrocarbon (benzo[a]pyrene) hydroxylases in liver from rats of different age, sex and nutritional status. Distinction of two types by 7,8-benzoflavone.

Two types of aryl hydrocarbon (benzo[a]pyrene) hydroxylase have been distinguished in liver from rats of different sex and age by their sensitivity to the synthetic flavonoid, 7,8-benzoflavone. One type, which is stimulated by the 7,8-benzoflavone, is found in newborn rats and predominates in the liver of adult male rats. This type is inducible by phenobarbital. A second type, which is inhibited by 7,8-benzoflavone, comprises a larger fraction in the liver of adult female rats and is inducible by polycyclic hydrocarbons in immature and mature animals of either sex. The presence of this form in adult female liver is also indicated by the kinetics of the hydroxylase reaction. Removal of solid food for 18 hr not only decreases hepatic aryl hydrocarbon hydroxylase activity in female rats, but also lowers the degree of inhibition by 7,8-benzoflavone. Kinetic data suggest that at low concentrations 7,8-benzoflavone acts as a competitive inhibitor but at higher concentrations inhibits the hydroxylation reaction by a more complex mechanism.     

Comparative metabolism of benzo(a)pyrene by ovarian microsomes of various species

Knowledge of the ability of the female reproductive system to metabolize polycyclic aromatic hydrocarbons (PAHs) is critical to the diagnosis and management of female infertility and for risk assessment purposes. The PAHs are a family of widespread pollutants that are released into the environment from automobile exhausts, cigarette smoke, burning of refuse, industrial emissions, and hazardous waste sites. In exposed animals, PAHs become activated to reactive metabolites that interfere with target organ function and as a consequence cause toxicity. The extent of susceptibility to PAH exposure may depend on the ability of animals to metabolize these chemicals. The present study has been undertaken to assess whether any differences exist among mammals in the metabolism of benzo(a)pyrene (BaP), a prototypical PAH compound. Microsomes isolated from the liver and ovaries of rats, mice, goats, sheep, pigs, and cows were incubated with 5 μM BaP. Postincubation, samples were extracted with ethyl acetate and analyzed for BaP/metabolites by reverse‐phase HPLC with fluorescence detection. The rate of metabolism (pmol of metabolite/min/mg protein) was found to be more in liver than in ovary in all the species studied (P < 0.05). The differences in metabolite concentrations were statistically significant (P < 0.0001) among the various species in both organs studied. Multiple species comparison also revealed that the differences were statistically significant (P < 0.001) between rodents (rat and mouse) and higher mammals (ewe, sow, and cow). Even among the higher mammals, in a majority of the cases, the differences in metabolite concentrations were significantly different (P < 0.001) both in ovary and liver. The BaP metabolites identified were 4,5‐diol; 7,8‐diol; 9,10‐diol; 3‐hydroxy BaP; and 9‐hydroxy BaP. The rodent microsomes produced considerably higher proportion of BaP 4,5‐diol and 9,10‐diol than did cow, sow, goat, and sheep. However, microsomes from higher mammals converted a greater proportion of BaP to 3‐hydroxy and 9‐hydroxy BaP, the detoxification products of BaP. Overall, our results revealed a great variation among species to metabolize BaP. © 2008 Wiley Periodicals, Inc. Environ Toxicol, 2009.     

Comparison of benzo(a)pyrene metabolism by testicular homogenate and the isolated perfused testis of rat following 2,3,7,8-tetrachlorodibenzo-P-dioxin treatment

Benzo(a)pyrene (BP) metabolism was studied in the cell free testicular homogenate and in the isolated perfused rat testis 72 h following tetrachlorodibenzo-P-dioxin (TCDD). The BP concentration for both metabolic systems was 2 × 10^–7 M. BP metabolites were extracted from testicular homogenate, perfusate and testicular tissue and subjected to high-pressure liquid Chromatographic analysis. The ratio of various BP metabolites in the cell free homogenates ranged from 3.5 to 164 times those of the isolated perfused testis, and the total BP metabolites in the cell free system of either control or TCDD-induced testis were 16 times that of the intact isolated perfused testis. The major BP metabolites in the organic extractable phase from the isolated perfused testis and the testicular homogenate were BP dihydrodiols and BP phenols, respectively.The ratio of water soluble metabolites to organic soluble metabolites in the homogenate and the isolated perfused testis is 1.1 and 3.0 respectively. Therefore, in the intact isolated testis, water soluble BP metabolites are formed three times greater than those of the organic soluble BP metabolites, and thus suggests that specific conjugating enzyme activities in the intact testis are greater than those of the homogenates. The magnitude of various BP metabolites in either the homogenates or the isolated perfused testis of TCDD treated rats ranged from 1.4 to 2.2 times their respective controls, except 4,5-dihydroxy-4,5-dihydrobenzo(a)pyrene in the isolated perfused testis was not altered by TCDD treatment. In conclusion, the isolated perfused rat testis is metabolically active and capable of biotransforming PAH. This system better reflects metabolic capability of the intact organ in the rats than do cell free homogenate system. The isolated perfused testis system retains the integrity of the complex biological organization of tissues, cell types, and enzymes in testicular metabolism and may provide data that may aid in the prediction of germ cell mutation as well as toxicity.The Abbreviations Used are PAH polycyclic aromatic hydrocarbons - BP benzo(a)pyrene - 9,10-diol 9,10-dihydroxy-9,10-dihydrobenzo(a)pyrene - 7,8-diol 7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene - 4,5-diol 4,5-dihydroxy-4,5-dihydrobenzo(a)pyrene - 3-OH 3-hydroxybenzo(a)pyrene - 9-OH 9-hy-droxybenzo(a)pyrene - 7-OH 7-hydroxybenzo(a)pyrene - 12-OH 12-hydroxybenzo(a)pyrene - 1,6-quinone benzo(a)pyrene-1,6-dione - 3,6-quinone benzo(a)pyrene 3,6-dione - 6,12-quinone benzo(a)pyrene 6,12-dione - DMBA 7,12-dimethylbenzanthracene - DMN dimethylnitrosamine - 7,8-diol 9,10-epoxide 5,7-t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene - HPLC high pressure liquid chromatography - TCDD 2,3,7,8-tetrachlorodibenzo-P-dioxin - HEPES N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid - NADP⁺ nicotinamide adenine dinucleotide - AHH aryl hydrocarbon hydroxylase - EH epoxide hydrolase - GSH-T glutathione transferase - HPRT hypoxanthine phosphoribosyltransferase     

Comparative metabolism and excretion of benzo(a)pyrene in 2 species of ictalurid catfish.

Differential susceptibility of polycyclic aromatic hydrocarbon (PAH)-mediated liver cancer exists in two related species of Ictalurid catfish. Two hypotheses are addressed in this study to explain this difference. Specifically, the relatively insensitive channel catfish 1) do not produce mutagenic PAH metabolites, and/or 2) they more quickly eliminate PAHs due to greater Phase II enzyme activities than the more sensitive brown bullhead. Livers and bile were collected from each species 6, 24, 72, and 168 h after a single 10 mg/kg i.p. benzo(a)pyrene (BaP) exposure. BaP treatment had no significant effect on cytosolic 1-chloro-2,4-dinitrobenzene or ethacrynic acid (EA)-glutathione-S:- transferase (GST) and cis-stilbene oxide-microsomal epoxide hydrolase (EH) activities of either species. Channel catfish EH and GST activities were 1.2-fold higher than brown bullhead activities (p = 0.058 and p < 0.002, respectively). HPLC-APCI-MS of extracted bile and bile enzymatically digested to detect glucuronyl transferase (GT), GST, and sulfotransferase (ST) conjugated metabolites indicated no species differences in elimination or profiles of total biliary metabolites. GT conjugates predominated; ST and GST conjugates were minimal. BaP-diones accounted for the majority of metabolites in both species. Overall, these results indicated that brown bullhead preferentially formed BaP-7,8-dihydrodiol, a precursor to the DNA-reactive BaP-7, 8-dihydrodiol-9,10-epoxide (BPDE), which may be linked to the increased PAH susceptibility in this species.     

The effect of long term treatment with disulfiram on content of cytochrome P-450 and on benzo(a)pyrene monooxygenase activity in microsomes isolated from the rat small intestinal mucosa

Disulfiram (DS) was administered perorally once a day to rats for 30 days to investigate the effects on cytochrome P-450 content and benzo(a)pyrene (BP) monooxygenase activity in microsomes isolated from the small intestinal mucosa. 50 mg or 100 mg DS/kg body weight caused a dose-related increase in BP monooxygenase activity, whereas the content of cytochrome P-450 was increased at the higher dose only. Similar absorption characteristics of cytochrome P-450 and turnover rates for BP on the basis o f cytochrome P-450 was observed among the different microsomal preparations. The addition of DS or diethyldithiocarbamate (DDTC) to incubates of intestinal microsomes inhibited BP monooxygenase activity. Microsomes isolated from DS-treated rats were however less sensitive to in vitro inhibition by DS.     

Increasing exposure levels cause an abrupt change in the absorption and metabolism of acutely inhaled benzo(a)pyrene in the isolated, ventilated, and perfused lung of the rat.

The carcinogenic polycyclic aromatic hydrocarbons (PAHs) are active primarily at the site of entry to the body. Lung cancer following inhalation of PAH-containing aerosols such as tobacco smoke is one likely example. A suggested mechanism for this site preference is a slow passage of the highly lipophilic PAHs through the thicker epithelia of the conducting airways, accompanied by substantial local metabolism in airway epithelium. However, it is likely that the airway epithelium will become saturated with PAHs at surprisingly low exposure levels. The purpose of this research was to quantify the level of saturation for inhaled benzo(a)pyrene (BaP) in the isolated, perfused lung (IPL) of the rat. BaP was coated onto carrier particles of silica 3.5 microm diameter at three different levels. The DustGun aerosol generator was then used to deliver respectively 2.2, 36, and 8400 ng of BaP to the IPL with the carrier particles in less than 1 min. For 77 min after the exposure, single-pass perfusate was collected from the lungs. Lungs were then removed and, with the perfusate, analyzed for BaP and metabolites. Results show that the absorption and metabolism of inhaled BaP in the lungs was highly dose dependent. At low exposure levels absorption of BaP in the mucosa was proportional to the concentration in the air/blood barrier and proceeded with substantial local metabolism. At higher exposure levels the capacity of the epithelium to dissolve and metabolize BaP became saturated, and the absorption rate remained constant until crystalline BaP had dissolved, and the process proceeded with much smaller fractions of BaP metabolites produced in the mucosa. This phenomenon may explain the well-known difficulties of inducing lung cancer in laboratory animals with inhalants containing carcinogenic PAHs, where similar lifespan exposures are used as humans may experience but with much higher dose rates.     

Influence of estradiol on the benzo(a)pyrene hydroxylase activity induced by hexachlorocyclohexane

Basal BP-hydroxylase activity was measured in male Swiss mice from the age of 3 weeks to 20 months. Maximal enzyme activity was at the age of 5 months. Comparison of the inducibility of BP-hydroxylase by HCH was also investigated in male and female mice of different ages. Male mice showed higher induction of BP-hydroxylase by HCH than females of the same ages. Sterilization of female mice enhanced enzyme induction. Estradiol exhibited competitive inhibition of BP-hydroxylase activity. After treatment with HCH for 8 months, female mice had a lower tumour incidence than males, and this paralleled a lower induction of BP-hydroxylase.     

DNA adduct formation in precision-cut rat liver and lung slices exposed to benzo[a]pyrene.

Chemical-DNA adducts provide an integrated measure of exposure, absorption, bioactivation, detoxification, and DNA repair following exposure to a genotoxic agent. Benzo[a]pyrene (BaP), a prototypical polycyclic aromatic hydrocarbon (PAH), can be bioactivated by cytochrome P-450s (CYPs) and epoxide hydrolase to genotoxic metabolites which form covalent adducts with DNA. In this study, we utilized precision-cut rat liver and lung slices exposed to BaP to investigate tissue-specific differences in chemical absorption and formation of DNA adducts. To investigate the contribution of bioactivating CYPs (such as CYP1A1 and CYP1B1) on the formation of BaP-DNA adducts, animals were also pretreated in vivo with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin) prior to in vitro incubation of tissue slices with BaP. Furthermore, the tissue distribution of BaP and BaP-DNA adduct levels from in vivo studies were compared with those from the in vitro tissue slice experiments. The results indicate a time- and concentration-dependent increase in tissue-associated BaP following exposure of rat liver and lung tissue slices to BaP in vitro, with generally higher levels of BaP retained in lung tissue. Furthermore, rat liver and lung slices metabolized BaP to reactive intermediates that formed covalent adducts with DNA. Total BaP-DNA adducts increased with concentration and incubation time. Adduct levels (fmol adduct/microg DNA) in lung slices were greater than liver at all doses. Liver slices contained one major and two minor adducts, while lung slices contained two major and 3 minor adducts. The tissue-specific qualitative profile of these adducts in tissue slices was similar to that observed from in vivo studies, further validating the use of this model. Pretreatment of animals with TCDD prior to in vitro incubation with BaP potentiated the levels of DNA adduct formation. TCDD pretreatment altered the adduct distribution in lung but not in liver slices. Together, the results suggest that tissue-specific qualitative and quantitative differences in BaP-DNA adducts could contribute to the lung being a target tissue for BaP carcinogenesis. Furthermore, the results validate the use of precision-cut tissue slices incubated in dynamic organ culture as a useful model for the study of chemical-DNA adduct formation.     

A toxicokinetic study to elucidate 3‐hydroxybenzo(a)pyrene atypical urinary excretion profile following intravenous injection of benzo(a)pyrene in rats

The toxicokinetics of benzo(a)pyrene (BaP) and 3‐hydroxybenzo(a)pyrene (3‐OHBaP) were assessed in 36 male Sprague–Dawley rats injected intravenously with 40 µmol kg¹ of BaP to explain the reported atypical urinary excretion profile of 3‐OHBaP. Blood, liver, kidney, lung, adipose tissue, skin, urine and feces were collected at t = 2, 4, 8, 16, 24, 33, 48, 72 h post‐dosing. BaP and 3‐OHBaP were measured by high‐performance liquid chromatography/fluorescence. A biexponential elimination of BaP was observed in blood, liver, skin and kidney (t_½ of 4.2–6.1 h and 12.3–14.9 h for initial and terminal phases, respectively), while a monoexponential elimination was found in adipose tissue and lung (t_½ of 31.2 and 31.5 h, respectively). A biexponential elimination of 3‐OHBaP was apparent in blood, liver and skin (t_½ of 7.3–11.7 h and 15.6–17.8 h for initial and terminal phases, respectively), contrary to adipose tissue, lung and kidney. In adipose tissue and lung, a monophasic elimination of 3‐OHBaP was observed (t_½ of 27.0 h and 24.1 h, respectively). In kidney, 3‐OHBaP kinetics showed a distinct pattern with an initial buildup during the first 8 h post‐dosing followed by a gradual elimination (t_½ of 15.6 h). In the 72‐h post‐treatment, 0.21 ± 0.09% (mean ± SD) of dose was excreted as 3‐OHBaP in urine and 12.9 ± 1.0% in feces while total BaP in feces represented 0.40 ± 0.16% of dose. This study allowed the identification of the kidney as a retention compartment governing 3‐OHBaP atypical urinary excretion. Copyright © 2010 John Wiley & Sons, Ltd.     

Synergistic effects of piperine and curcumin in modulating benzo(a)pyrene induced redox imbalance in mice lungs

The objective of the present study was to evaluate the effects of curcumin alone and with adjuvant piperine against benzo(a)pyrene (BaP) induced oxidative stress in lungs of male Swiss albino mice. Mice were pretreated either with curcumin (100?mg/kg body weight), or piperine (20?mg/kg body weight), and in combination of both for one week, followed by single dose of benzo(a)pyrene (125?mg/kg body weight) treatment. Treatment with benzo(a)pyrene resulted in increased levels of lipid peroxides (LPO), protein carbonyl content (PCC) and with consequent decrease in the levels of tissue antioxidants like superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx) and reduced glutathione (GSH), which however, were increased significantly following curcumin treatment, but the increase was more pronounced when piperine was used as an adjuvant. BaP treatment alone did not alter significantly the GST activity. Pretreatment with curcumin increased the GST activity in BaP treated group, which was enhanced further upon synergistic treatment with piperine and curcumin. Therefore, combined administration of curcumin and piperine shall prove to be more effective in attenuating BaP induced toxicity.     

Induction of hepatocellular carcinoma in rat liver by initial treatment with benzo(a)pyrene after partial hepatectomy and promotion by phenobarbital

The hepatocarcinogenicity of benzo(a)pyrene(BP) in the rat was examined. Rats were treated with BP after partial hepatectomy and then kept on a diet containing phenobarbital (PB) as a promoter. Tumors including a hepatocellular carcinoma developed in the rat liver by week 52.     

The biliary excretion and enterohepatic circulation of benzo(a)pyrene and its metabolites in the rat

The enterohepatic circulation of benzo(a)pyrene (BP) has been investigated in the rat with a view to determining the availability of potentially toxic metabolities to tissues within this cycle. Some 60% of the dose of [¹⁴C]-BP (3 μmoles kg^?1, i.v.) is excreted in bile in 6 hr, with less than 3% in urine. The biliary metabolities are mainly polar conjugates; only 8% of the ¹⁴C in 2 hr bile samples can be directly extracted into ethyl acetate. However, following hydrolysis by β-glucuronidase some 40% of the ¹⁴C is extractable at pH 7. The extract consisted of polar metabolites (polyhydroxylated and/or conjugated; 37.5%), BP 4,5-diol (16.8%), BP 3,6-quinone (5.9%). 9-hydroxy BP (5.4%) and 3-hydroxy BP (5.3%) as indicated by co-chromatography with authentic standards on reversed phase HPLC, together with several unidentified metabolites. The proximate carcinogen BP 7, 8-diol was not detected. Biliary metabolites of [¹⁴C]-BP undergo enterohepatic recirculation in the rat; following the intraduodenal infusion of bile containing metabolites of [¹⁴C]-BP into bile duct cannulated rats, approximately 20% of the dose is absorbed and excreted in bile in 30 hr, with only 1% in urine. The pattern of metabolites in this bile is very similar to that in bile from rats administered [¹⁴C]-BP i.v. Following a single i.v. dose of [¹⁴C]-BP (3 μmoles kg^?1) to rats with re-entrant bile duct cannulae, which allowed intermittent collection of bile over a period of several days with minimal interference to the enterohepatic circulation, the proximate carcinogen BP 7, 8-diol was detected in recirculating bile. Biliary metabolites of BP, which have recently been shown to be mutagenic, can thus traverse the intestine to undergo enterohepatic circulation in the rat.     

Effect of NO2 and SO2 inhalation on benzo(a)pyrene metabolism in rat lung

Male Wistar rats were continuously exposed to NO₂ (14.4 ppm), SO₂ (46.5 ppm) and to a mixture of both gases and their effect on lung microsomal aryl hydrocarbon (benzo(a)pyrene) hydroxylase (AHH) activity was determined. The pre-exposed animals were administered methylcholanthrene (MC) to investigate the exposure effect on enzyme inducibility and pattern of B(a)P metabolites. NO₂ significantly increased AHH activity but no marked change was noted with SO₂. Induction of AHH by MC was markedly inhibited by SO₂, only slightly by mixture of NO₂-SO₂ but not with NO₂ alone.     

Differences in benzo(a)pyrene metabolism between cultured human and murine bronchial cells after pre-treatment with benz(a)anthracene

Primary cultures of human and murine (strain C3Hz) bronchial epithelial cells were pretreated with benz(a)anthracene (BA) (10 microM). 16 h later the formation of phenolic as well as dihydrodiol metabolites of benzo(a)pyrene (BP) was measured. Whereas murine cultures showed enhanced metabolism towards both phenolic and dihydrodiol compounds, in the human cultures only phenolic BP-metabolites were increased. In view of their precursor role in the formation of biologically active diol-epoxides, formation of dihydrodiol-derivatives can be considered as a key factor in determining susceptibility to polycyclic aromatic hydrocarbon (PAH)-induced carcinogenesis. Therefore the observations of this study indicate that animal model systems for PAH carcinogenesis in man have to be selected on the basis of comparable metabolite patterns.     

3-Methylcholanthrene and benzo(a)pyrene modulate cardiac cytochrome P450 gene expression and arachidonic acid metabolism in male Sprague Dawley rats

Background and purpose:

There is a strong correlation between cytochrome P450 (P450)-dependent arachidonic acid metabolism and the pathogenesis of cardiac hypertrophy. Several aryl hydrocarbon receptor (AhR) ligands were found to alter P450-dependent arachidonic acid metabolism. Here, we have investigated the effect of 3-methylcholanthrene (3-MC) and benzo(a)pyrene (BaP), two AhR ligands, on the development of cardiac hypertrophy.

Experimental approach:

Male Sprague Dawley rats were injected (i.p.) daily with either 3-MC (10 mg·kg⁻¹) or BaP (20 mg·kg⁻¹) for 7 days. Then hearts were removed, and the heart to body weight ratio and the gene expression of the hypertrophic markers and P450 genes were determined. Levels of arachidonic acid metabolites were determined by liquid chromatography-electron spray ionization-mass spectrometry.

Key results:

Both 3-MC and BaP increased the heart to body weight ratio as well as the hypertrophic markers, atrial natriuretic peptide and brain natriuretic peptide. 3-MC and BaP treatment increased the gene expression of CYP1A1, CYP1B1, CYP2E1, CYP4F4, CYP4F5 and soluble epoxide hydrolase. Both 3-MC and BaP treatments increased the dihydroxyeicosatrienoic acids (DHETs) : epoxyeicosatrienoic acids (EETs) ratio and the 20-hydroxyeicosatetraenoic acid (20-HETE) : total EETs ratio. Treatment with benzo(e)pyrene, an isomer of BaP that is a poor ligand for the AhR, did not induce cardiac hypertrophy in rats, confirming the role of AhR in the development of cardiac hypertrophy. Treatment with the ω-hydroxylase inhibitor, HET0016, significantly reversed BaP-induced cardiac hypertrophy.

Conclusions and implications:

3-MC and BaP induce cardiac hypertrophy by increasing the ratio of DHETs : EETs and/or the ratio of 20-HETE : total EETs, through increasing soluble epoxide hydrolase activity.