mRNA Expression Levels Of CYP1A1 Research Articles

Effects of Curcuma comosa Roxb. Extract on the Expression of CYP450s in Immortalized Hepatocyte-like Cells (imHC)

Hepatocytes are the most abundant liver cells that produce biotransformation enzymes, including phase I metabolism enzymes and cytochrome P450s (CYP450s). These enzymes are induced or suppressed by several drugs and chemicals. Here, immortalized hepatocyte-like cells (imHC) were applied as a novel hepatocyte to study the CYP450 enzyme expression induced by a Thai folk herb, Curcuma comosa Roxb. (C. comosa). The results demonstrated that the mRNA expression levels were dependent on cell cultivation conditions, the extract's concentration, and the duration of treatment. In the 3-day cultivation, 1 µg/mL C. comosa extract upregulated the mRNA expression levels of CYP1A1, CYP1A2, CYP2B6, CYP2C9, CYP2D6, CYP2E1 and CYP3A4 in imHC, after treatment for 48 h, which was comparable to the expression levels of those genes in HepG2 cells. In addition, C. comosa increased CYP3A4 protein expression in both imHC and HepG2 cells. Furthermore, 1 µg/mLC. comosa could neutralize erythromycin's inhibitory effect, a specific CYP3A4 inhibitor, on CYP3A4 enzyme activity. In conclusion, imHC exhibited several detoxification enzymes, and this cell could be used to replace hepatic carcinoma cells in hepatotoxic studies. The function of C. comosa as a CYP enzyme-inducing agent indicates a promising treatment for the prevention of drug-induced hepatotoxicity by activating detoxification enzymes.

The palliative effect of camel milk on hepatic CYP1A1 gene expression and DNA damage induced by fenpropathrin oral intoxication in male rats

The present study investigated the alleviating role of camel milk (CM) in the mitigation of fenpropathrin (FNP) type II pyrethroid induced oxidative stress, alterations of hepatic (CYP1A1) mRNA expression pattern, and DNA damage using the alkaline comet assay (SCGE) in male rats. Sixty male Sprague-Dawley rats were separated into six groups (n = 10): 1st control (C), 2nd corn oil (CO), 3rd (CM): gavaged CM 2ml/rat, 4th (FNP): gavaged FNP 7.09 mg/kg body weight (BW), 5th (FNP pro/co-treated): gavaged CM firstly for 15 days, then CM + FNP by the same mentioned doses and route, 6th (FNP + CM co-treated): gavaged FNP firstly followed by CM by the same mentioned doses and route. Rats were orally gavaged three times per week, day after day for 60 days. FNP exposure significantly reduced serum glutathione (GSH) levels, but significantly increased serum levels of superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA), protein carbonyl (PCO), and 8hydroxy2deoxyguanosine (8OH2dG). Additionally, FNP exposure significantly up-regulated the mRNA expression levels of hepatic CYP1A1 and increased the SCGE indices in whole blood, liver, and spleen tissues of exposed male rats. Administration of CM significantly regulated the FNP induced oxidative stress, reduced hepatic CYP1A1 mRNA expression levels and values of comet assay indices particularly in the (CM + FNP pro/co-treated) group compared to the (FNP + CM co-treated) group. In conclusion, our results indicate, for the first time, that FNP retains an in vivo genotoxic potential at a dose of (1/10 LD50) and up-regulated hepatic CYP1A1 mRNA expression in male rats. Additionally, CM supplements may improve the genotoxic outcomes, oxidative stress, and altered CYP1A1 mRNA expression induced by FNP particularly in the pro/concurrent-treatment compared to the concurrent treatment alone.

Otoprotective Effects of Zingerone on Cisplatin-Induced Ototoxicity.

Previous studies have described the effects of zingerone (ZO) on cisplatin (CXP)-induced injury to the kidneys, liver, and other organs but not to the cochlea. This study aimed to investigate the effects of ZO on CXP-induced ototoxicity. Eight-week-old Sprague–Dawley rats were used and divided into a control group, a CXP group, and a CXP + ZO group. Rats in the CXP group received 5 mg/kg/day CXP intraperitoneally for five days. Rats in the CXP + ZO group received 5 mg/kg/day CXP intraperitoneally for five days and 50 mg/kg/day ZO intraperitoneally for seven days. Auditory brainstem response thresholds (ABRTs) were measured before (day 0) and after (day 10) drug administration. Cochlear histology was examined using hematoxylin and eosin (H&E) staining and cochlear whole mounts. The expression levels of cytochrome P450 (CYP)1A1, CYP1B1, inducible nitric oxide synthase (iNOS), nuclear factor kappa B (NFκB), tumor necrosis factor alpha (TNFα), and interleukin 6 (IL6) were estimated using quantitative reverse transcription-polymerase chain reaction. The expression levels of heme oxygenase 1 (HO1) and caspase 3 were analyzed via Western blotting. The auditory thresholds at 4, 8, and 16 kHz were attenuated in the CXP + ZO group compared with the CXP group. The mRNA expression levels of CYP1A1, CYP1B1, iNOS, NFκB, TNFα, and IL6 were lower in the CXP + ZO group than in the CXP group. The protein expression levels of HO1 and caspase 3 were lower in the CXP + ZO group than in the CXP group. Cotreatment with ZO exerted otoprotective effects against CXP-induced cochlear injury via antioxidative and anti-inflammatory activities involving CYPs, iNOS, NFκB, and TNFα.

Histopathological, biochemical and molecular studies on the toxic effect of used engine oil on the health status of Oreochromis niloticus

The accidental spilling of petroleum oils into natural water resources expose fishes in the effluent area to serious problems.. Oreochromis niloticus were used in the current study as a model to investigate the toxicity of used engine oil and to evaluate the protective role of vitamin C against this toxicity. The oil concentration used in this study was previously determined to be 0.25 ml/l by 96 h-LC50. After 21 days of engine oil exposure, haematological and biochemical analyses revealed significant reduction in RBCs counts, haemoglobin concentrations and total proteins. However, ALT, AST and glucose levels were significantly increased by the end of the experiment indicating the damaging effects of the oil on fish tissues. Oxidative stress biomarkers were also measured; liver CAT activity was significantly decreased in the oil exposed group compared to control group, while MDA levels were significantly elevated. Histopathological examination showed the presence of several alterations in hepatic and branchial tissues in exposed group compared to the control group. Significant elevations in CYP1 A1 mRNA expression levels in hepatic tissue were also detected in the group exposed to used engine oil compared to the control group. However, supplementation of fishexposed to used engine oil with vitamin Csignificantly enhance the biochemical, oxidative and histological parameters.

Cytochrome P450 1A1 and 1B1 promoter CpG island methylation regulates rat liver injury induced by isoniazid.

DNA methylation is an important component of epigenetics that is involved in the occurrence and development of a variety of diseases. The present study aimed to clarify the relationship between cytochrome P450 (CYP)1A1 and CYP1B1 promoter CpG island methylation and isoniazid-induced liver injury in rats, and to explore the possible mechanism, rats were given an intragastric dose of isoniazid (55 mg·kg−1·d−1). High performance liquid chromatography was used to analyze the DNA methylation level of the whole genome in liver tissue. Methylation-specific polymerase chain reaction (PCR) was used to detect the methylation level of CpG islands in the promoter region of CYP1A1 and CYP1B1. Reverse transcription-quantitative PCR was used to determine the mRNA expression levels of CYP1A1, CYP1B1, toll-like receptor 4 (TLR4), extracellular signal-regulated kinase (ERK) 2, peroxisome proliferator-activated receptor (PPAR) -γ, interleukin (IL)-6 and tumor necrosis factor (TNF)-α. The expression levels of CYP1A1 and CYP1B1 proteins were measured by ELISA, and malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were analyzed by colorimetric method. Liver tissue pathology, an indicator of liver function, indicated rat liver injury at 10 days following isoniazid treatment. Whole-genome methylation levels were gradually reduced, and methylation at day 7 post-treatment was significantly lower than the control group. CYP1A1 and CYP1B1 promoter CpG island methylation level was significantly increased at 3 days post-treatment. CYP1A1 and CYP1B1 mRNA expression levels were significantly reduced from day 7 and 10, respectively. These results suggested that CpG island hypermethylation of the CYP1A1 and CYP1B1 promoters regulate the low expression of genes involved in the occurrence of isoniazid-induced liver injury. With the alterations of CYP1A1 and CYP1B1 expression, the mRNA expression levels of TLR4, ERK, MDA, IL-6 and TNF-α were upregulated, and the expression of SOD and PPAR-γ were downregulated. These data demonstrated that alterations in methylation patterns may involve changes in the TLR4-ERK signaling pathway and PPAR-γ, which may alter the expression of MDA, SOD, IL-6 and TNF-α, leading to liver injury.

Toxic Effects of the Major Components of Diesel Exhaust in Human Alveolar Basal Epithelial Cells (A549).

We investigated the toxicity of benzo[a]pyrene (B[a]P), 1-nitropyrene (1-NP) and 3-nitrobenzanthrone (3-NBA) in A549 cells. Cells were treated for 4 h and 24 h with: B[a]P (0.1 and 1 μM), 1-NP (1 and 10 μM) and 3-NBA (0.5 and 5 μM). Bulky DNA adducts, lipid peroxidation, DNA and protein oxidation and mRNA expression of CYP1A1, CYP1B1, NQO1, POR, AKR1C2 and COX2 were analyzed. Bulky DNA adducts were induced after both treatment periods; the effect of 1-NP was weak. 3-NBA induced high levels of bulky DNA adducts even after 4-h treatment, suggesting rapid metabolic activation. Oxidative DNA damage was not affected. 1-NP caused protein oxidation and weak induction of lipid peroxidation after 4-h incubation. 3-NBA induced lipid peroxidation after 24-h treatment. Unlike B[a]P, induction of the aryl hydrocarbon receptor, measured as mRNA expression levels of CYP1A1 and CYP1B1, was low after treatment with polycyclic aromatic hydrocarbon (PAH) nitro-derivatives. All test compounds induced mRNA expression of NQO1, POR, and AKR1C2 after 24-h treatment. AKR1C2 expression indicates involvement of processes associated with reactive oxygen species generation. This was supported further by COX2 expression induced by 24-h treatment with 1-NP. In summary, 3-NBA was the most potent genotoxicant, whereas 1-NP exhibited the strongest oxidative properties.

Relationship of blood aryl hydrocarbon receptor mRNA and cytochrome P450 1A1 mRNA expression with corrected QT interval among residents exposed to arsenic via drinking water

To investigate the relationship between blood aryl hydrocarbon receptor(AhR)mRNA and cytochrome P450(CYP)1A1 mRNA expression and corrected QT interval among residents exposed to arsenic via drinking water. Arsenic exposure area in Bayannao'er city of Nei Monggol Autonomous Region was selected as the survey point, and the residents living more than 10 years in this area were investigated from December 2012 to January 2015.A total of 233 residents were divided into four groups according to drinking water arsenic concentration (ranged from 0.8 to 824.7 μg/L): control group (drinking water arsenic concentration <10.0 μg/L, n=55), low exposure group (drinking water arsenic concentration 10.0-99.9 μg/L, n=47), middle exposure group (drinking water arsenic concentration 100.0-199.9 μg/L, n=45), high exposure group (drinking water arsenic concentration ≥200.0 μg/L, n=86). Epidemiological investigation was performed.Real-time PCR technology was used to detect the expression levels of blood AhR mRNA and CYP1A1 mRNA, and the relationship between expression levels of blood AhR mRNA and CYP1A1 mRNA and corrected QT interval was analyzed. (1) Blood AhR mRNA and CYP1A1 mRNA expression levels were similar among control group, low exposure group and middle exposure group (all P>0.05) while blood AhR mRNA (3.18×10(-3)(2.42×10(-3), 4.45×10(-3)) vs.2.30×10(-3)(1.53×10(-3), 3.20×10(-3)), P<0.05) and CYP1A1 mRNA (1.58×10(-3)(0.80×10(-3), 2.73×10(-3))vs.1.00×10(-3)(0.59×10(-3), 2.09×10(-3)), P<0.05) expression levels were significantly higher in high dose group than in control group.(2) AhR mRNA expression level was similar between residents with longer corrected QT interval and residents with normal corrected QT interval (2.89×10(-3)(1.90×10(-3), 3.71×10(-3)) vs.2.58×10(-3)(1.85×10(-3), 3.57×10(-3)), P>0.05). CYP1A1 mRNA expression level was significantly higher in residents with longer corrected QT interval than in residents with normal corrected QT interval (1.50×10(-3)(0.78×10(-3), 2.63×10(-3)) vs.1.16×10(-3)(0.64×10(-3), 2.36×10(-3)), P<0.05). (3) AhR mRNA expression level was similar between residents with sinus tachycardia or bradycardia or normal heart rate(2.89×10(-3)(1.71×10(-3), 4.45×10(-3)) vs.2.64×10(-3)(1.86×10(-3), 3.52×10(-3)), P>0.05). CYP1A1 mRNA expression level was significantly higher in residents with sinus tachycardia or bradycardia than in residents with normal heart rate (1.47×10(-3)(0.87×10(-3), 2.77×10(-3)) vs.1.24×10(-3)(0.64×10(-3), 2.31×10(-3)), P<0.05). Long-term exposure to arsenic is associated with upregulated blood AhRmRNA and CYP1A1 mRNA expression.Blood CYP1A1 mRNA expression, but not AhRmRNA expression, is associated with prolonged corrected QT interval.

Gender differences in detoxification metabolism of polycyclic aromatic hydrocarbon (chrysene) in scallop Chlamys farreri during the reproduction period

This study aims to investigate the effects of chrysene (CHR) on biotransformation and detoxification responses of mature scallop Chlamys farreri during the reproduction period. Scallops were exposed to 0.2, 0.8 and 3.2μg/L CHR for 21days; at day 10 scallops were induced to spawn. At days 1, 3, 6, 10, 11, 15 and 21, enzymatic activities of 7-ethoxyresorufin O-deethylase (EROD) and glutathione-s-transferase (GST), related mRNA expression levels of CYP1A1, GST-pi and P-glycoprotein (P-gp) in digestive glands and CHR bioaccumulation in tissues were examined by separately analyzing male and female scallops. During the pre-spawn period, CHR concentrations of the treated groups in tissues except the hemolymph increased rapidly. Levels of enzymatic activities and related gene expressions were all induced by the exposure to CHR for females and males. GST activity and GST-pi mRNA expression showed a good time- and dose-dependent relationship only in males, and P-gp mRNA expression exhibited a dose-dependent manner in both sexes. During the post-spawn period, spawning caused significant reductions of bioaccumulation in tissues but the gill and hemolymph. Enzymatic activities and related gene expressions were for females significantly depressed at day 21 at 0.8 or 3.2μg/L CHR. Overall, females accumulated more CHR than males, while males were more sensitive than females to CHR exposure in gene expressions and enzyme activities. P-gp mRNA expression seemed to be a potential biomarker for PAH exposure. These results will offer the information on CHR biotransformation in this species, and ensure the influence of gender and reproductive status on PAH detoxification metabolism.

Antioxidant enzymatically modified isoquercitrin or melatonin supplementation reduces oxidative stress-mediated hepatocellular tumor promotion of oxfendazole in rats

To clarify whether enzymatically modified isoquercitrin (EMIQ) or melatonin (MLT) supplementation reduces oxidative stress-mediated hepatocellular tumor-promoting effect of oxfendazole (OX), a benzimidazole anthelmintic, male rats were administered a single intraperitoneal injection of N-diethylnitrosamine (DEN) and were fed a diet containing OX (500 ppm) for 10 weeks with or without EMIQ (2,000 ppm) or MLT (100 ppm) in the drinking water after DEN initiation. One week after the commencement of the administration of OX, rats were subjected to two-thirds of partial hepatectomy. The number of GST-P-positive foci promoted by OX was significantly inhibited by the combined antioxidant EMIQ or MLT administration, and the area of GST-P-positive foci was inhibited by the administration of MLT. Real-time RT-PCR analysis revealed decreases in mRNA expression levels of cytochrome P450, family 2, subfamily b, polypeptide 2 (Cyp2b2) and malic enzyme 1 (Me1) in the DEN-OX-EMIQ and DEN-OX-MLT groups and decreases in mRNA expression levels of Cyp1a1 and aldo-keto reductase family 7, member A3 (Akr7a3) in the DEN-OX-MLT group compared to those in the DEN-OX group. In in vitro ROS production assay, inhibited production of NADPH-dependent ROS was observed by the treatment with EMIQ or MLT. These results suggest that coadministration of EMIQ or MLT suppresses the hepatocellular tumor-promoting activity of OX in rats through the decrease in ROS production by the activation of CYPs.

Aryl hydrocarbon receptor (AHR) and AHR nuclear translocator (ARNT) expression in Baikal seal ( Pusa sibirica) and association with 2,3,7,8-TCDD toxic equivalents and CYP1 expression levels

Most toxic effects of 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) and related planar halogenated aromatic hydrocarbons (PHAHs) are mediated by ligand-activated aryl hydrocarbon receptor (AHR) signaling pathway. To understand the regulation mechanism of AHR and AHR nuclear translocator (ARNT) expression in wild Baikal seal ( Pusa sibirica) population contaminated by PHAHs, the present study investigated hepatic mRNA expression levels of AHR and its heterodimer, ARNT genes, in association with biological index (age, gender and body weight), PHAH accumulation and expression levels of cytochrome P450 (CYP) 1A and 1B. While there was no gender difference, the AHR mRNA expression levels were increased with ages ( p = 0.014) and body weights ( p = 0.015), indicating that AHR expression might be affected by these biological factors. The AHR mRNA expression levels exhibited significant positive correlations with total TEQs and most of individual congener TEQs derived from polychorinated dibenzo- p-dioxins, dibenzofurans and non- ortho coplanar polychorinated biphenyls (PCBs), indicating the transcriptional up-regulation of AHR expression by these congeners. On the other hand, there was no significant correlation between individual TEQs from mono -ortho coplanar PCBs and AHR expression. These results imply the structure-related transcriptional activity of AHR among PHAHs congeners. AHR mRNA levels showed positive correlations with both CYP1A protein ( p = 0.039) and CYP1A1 mRNA expression levels ( p = 0.046). In contrast to AHR expression, neither the total nor individual congener TEQs influenced ARNT at the transcriptional level. ARNT mRNA showed significant negative correlations with CYP1A/1B protein ( p = 0.027 and p = 0.006) and CYP1A1 mRNA expression levels ( p = 0.039), implying the existence of different transcriptional regulation between AHR and ARNT genes and negative regulation by CYP1A/1B-mediated signaling pathways. The present findings may render significant insight on the basic mechanisms underlying regulation of AHR and ARNT expressions associated with biological factors and PHAH exposure in wild mammalian populations.

Molecular cloning and mRNA expression of cytochrome P4501A1 and 1A2 in the liver of common minke whales ( Balaenoptera acutorostrata)

This study presents full-length cDNA sequences of CYP1A1 and 1A2, in common minke whale ( Balaenoptera acutorostrata) from the North Pacific. Both CYP1A1 and CYP1A2 cDNAs had an open reading frame of 516 amino acid residues, and predicted molecular masses were 58.3 kDa and 58.1 kDa, respectively. The deduced full-length amino acid sequence of CYP1A1 revealed higher identities with those of sheep (86%) and pig (87%), and that of CYP1A2 was most closely related to human (82%) and monkey CYP1A2 (82%) among species from which CYP1A2 has been isolated so far. Differences in certain conserved and functional amino acid residues of CYP1A1 and 1A2 between common minke whale and other mammalian species indicate the possibility of their specific metabolic function. Concentrations of organochlorine compounds (OCs) including PCBs and DDTs analyzed in common minke whale liver showed no significant correlation with hepatic mRNA expression levels of CYP1A1 and CYP1A2, indicating no induction of these enzymes by such OCs.

Evidence for a new human CYP1A1 regulation pathway involving PPAR-α and 2 PPRE sites

Cytochrome P450 1A1 catalyzes the degradation of endobiotics (estradiol, fatty acids, and so on) and the bioactivation of numerous environmental procarcinogens, such as arylamines and polycyclic aromatic hydrocarbons, that are found in food. Several peroxisome proliferators and arachidonic acid derivatives enhance cytochrome P450 1A1 activity, but the mechanisms involved remain unknown. The aim of this work was to study the role of peroxisome proliferator-activated receptors in cytochrome P450 1A1 gene induction. The role of peroxisome proliferator-activated receptor transcription factors in cytochrome P450 1A1 induction was assessed by means of enzymatic activities, quantitative real-time polymerase chain reaction, gene reporter assays, mutagenesis, and electrophoretic mobility shift assay. We show that peroxisome proliferator-activated receptor-alpha agonists (WY-14643, bezafibrate, clofibrate, and phthalate) induce human cytochrome P450 1A1 gene expression, whereas 2,4-thiazolidinedione, a specific peroxisome proliferator-activated receptor-gamma agonist, represses it. The induction of cytochrome P450 1A1 transcripts by WY-14643 was associated with a marked increase of ethoxyresorufin O -deethylase activity (10-fold at 200 mumol/L). Transfection of peroxisome proliferator-activated receptor-alpha complementary DNA enhanced cytochrome P450 1A1 messenger RNA induction by WY-14643, although WY-14643 failed to activate xenobiotic responsive element sequences. Two peroxisome proliferator response element sites were located at positions -931/-919 and -531/-519 of the cytochrome P450 1A1 promoter. Their inactivation by directed mutagenesis suppressed the inductive effect of WY-14643 on cytochrome P450 1A1 promoter activation. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay experiments showed that the 2 cytochrome P450 1A1 peroxisome proliferator response element sites bind the peroxisome proliferator-activated receptor-alpha/retinoid X receptor-alpha heterodimer. We describe here a new cytochrome P450 1A1 induction pathway involving peroxisome proliferator-activated receptor-alpha and 2 peroxisome proliferator response element sites, indicating that peroxisome proliferator-activated receptor-alpha ligands, which are common environmental compounds, may be involved in carcinogenesis.

Aryl hydrocarbon receptor/dioxin receptor in human monocytes and macrophages.

Aryl hydrocarbon receptor (AhR) belongs to the bHLH/PAS transcription factor family and is activated by various polycyclic or halogenated aromatic hydrocarbons, e.g. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 3-methylcholanthrene (3MC). In the present study, we showed that in U937 cells and human macrophages AhR, with its partner cofactor Arnt, is expressed and CYP1A1 mRNA expression is induced in the presence of AhR ligand 3MC. Moreover, we showed that AhR, associating with Arnt, binds to target DNA sequences and activates transcription. Since part of AhR is activated into DNA binding species in the absence of exogenous ligand and competitive AhR antagonist alpha-naphthoflavone inhibits this activation process with reducing CYP1A1 mRNA expression levels, the presence of endogenous ligand is indicated.