Cytochrome P450 1A1 Expression Research Articles

Generation and application of CES1-knockout Tet-Off-regulated CYP3A4 and UGT1A1-expressing Caco-2 cells

Caco-2 cells, a human colorectal adenocarcinoma cell line, are widely used to model small intestinal epithelial cells in the drug development process because they can predict drug absorption with high accuracy. However, Caco-2 cells have several issues. First, Caco-2 cells have little expression of cytochrome P450 3A4 (CYP3A4), which is a major drug-metabolizing enzyme in the human intestine. We previously developed Caco-2 cells whose expression of CYP3A4 can be controlled using doxycycline (Dox) (CYP3A4-Caco-2 cells) (Ichikawa et al., Sci. Rep, 2021). However, since the Tet-On system was used to regulate CYP3A4 expression in these cells, there was concern about drug-drug interactions. The second issue is that in the human small intestine, carboxylesterase 2 (CES2) is more highly expressed than carboxylesterase 1 (CES1), while in Caco-2 cells CES1 is more highly expressed. The third issue is the low level expression of uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1), a phase II drug-metabolizing enzyme. In this study, we used genome-editing technology to establish CES1-knockout Caco-2 cells whose CYP3A4 and UGT1A1 expression can be regulated by the Tet-Off system. These cell lines would be useful in pharmaceutical researches, including intestinal toxicological studies, as an in vitro model for orally administered drugs.

Thyme and Oregano Oil Potential Therapeutics against Malathion Toxicity through Biochemical, Histological, and Cytochrome P450 1A2 Activities in Male Wistar Rats.

The widespread use of malathion may offer several hazards to humans and animals; additionally, many medicinal plants provide what is known as a broad antitoxicity treatment. This study was carried out to investigate hazardous biochemical and histological reactions to MOP and evaluate the effectiveness of TEO and OEO essential oils in restoring normal physiological conditions after MOP exposure by measuring enzyme-specific activity for Cytochrome P450 1A2 (CYP1A2). One hundred and twenty rats were divided into six groups of twenty animals each: (i) C - MOP served as the control group, (ii) C + MOP treated with 5 mg/kg/BW of Malathion-D10, (iii) TEO treated with 100 mg/kg/BW of oregano essential oil, (iv) TEO treated with 100 mg/kg/BW of thyme essential oil, (v) MOP + OEO treated with 5 mg/kg/BW of Malathion-D10 and 100 mg/kg/BW of oregano essential oil, and (vi) MOP + TEO treated with 5 mg/kg/BW of Malathion-D10 and 100 mg/kg/BW of thyme essential oil. The results indicated the protective effects of OEO and TEO against MOP-induced weight loss. Additionally, there was a significant improvement in ALT, AST, and ALK-Ph after being treated with OEO and TEO, either alone or after MOP exposure. Also, treatment with OEO and TEO ameliorated these oxidative stress parameters, indicating their antioxidative properties. A histopathological examination of liver tissues showed reduced hepatocellular damage and improved liver architecture in the OEO and TEO, both alone and in combination with MOP, and protective effects were more pronounced in the TEO-treated groups. However, the results indicated that TEO was more effective than OEO in increasing CYP1A2 expression and alleviating MOP-induced toxicity. Specifically, TEO showed higher protein expression and therapeutic action in reducing liver damage. In conclusion, these findings suggest that OEO and TEO may be potent therapeutic agents against MOP toxicity, offering protective effects by enhancing CYP1A2 activity and mitigating organ damage. Such knowledge would be an important step toward developing potentially unique treatment options for natural antitoxins.

A Natural Chalcone Cardamonin Inhibited Transformation of Aryl Hydrocarbon Receptor Through Binding to the Receptor Competitively.

Chalcones are widely present in most plants and have various health beneficial functions. This study investigates the suppressive effect of 13 natural and synthetic chalcones on transformation of aryl hydrocarbon receptor (AhR) induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and 3-methylcholanthrene (3-MC) in a cell-free system, Hepa-1c1c7 cells, and liver of ICR mice. In the cell-free system, cardamonin dose-dependently inhibits AhR transformation. Chalcones with substitution on 2' and/or 6' position is important for the suppressive effect, while the substitution on 4' position is negatively for the effect. Moreover, cardamonin and 2'-hydroxychalcone competitively inhibit the binding of [3H]-3-MC to the AhR. In Hepa-1c1c7 cells, cardamonin inhibits AhR transformation and expression of cytochrome P4501A1 (CYP1A1) in a dose-dependent manner through suppressing TCDD-induced phosphorylation of both AhR and AhR nuclear translocator, heterodimerization of them, and nuclear translocation of AhR. In the liver of mice, oral administered cardamonin also inhibits 3-MC-induced AhR translocation and expression of CYP1A1. Among used chalcones, a natural chalcone cardamonin competitively binds to AhR and suppresses its transformation. Thus, cardamonin is an effective food factor for suppression of the dioxin-caused biochemical alterations and toxicities.

High Expression of AhR and Environmental Pollution as AhR-Linked Ligands Impact on Oncogenic Signaling Pathways in Western Patients with Gastric Cancer-A Pilot Study.

The vast majority of gastric cancer (GC) cases are adenocarcinomas including intestinal and diffuse GC. The incidence of diffuse GC, often associated with poor overall survival, has constantly increased in Western countries. Epidemiological studies have reported increased mortality from GC after occupational exposure to pro-carcinogens that are metabolically activated by cytochrome P450 enzymes through aryl hydrocarbon receptor (AhR). However, little is known about the role of AhR and environmental AhR ligands in diffuse GC as compared to intestinal GC in Western patients. In a cohort of 29, we demonstrated a significant increase in AhR protein and mRNA expression levels in GCs independently of their subtypes and clinical parameters. AhR and RHOA mRNA expression were correlated in diffuse GC. Further, our study aimed to characterize in GC how AhR and the AhR-related genes cytochrome P450 1A1 (CYP1A1) and P450 1B1 (CYP1B1) affect the mRNA expression of a panel of genes involved in cancer development and progression. In diffuse GC, CYP1A1 expression correlated with genes involved in IGF signaling, epithelial-mesenchymal transition (Vimentin), and migration (MMP2). Using the poorly differentiated KATO III epithelial cell line, two well-known AhR pollutant ligands, namely 2-3-7-8 tetrachlorodibenzo-p-dioxin (TCDD) and benzo[a]pyrene (BaP), strongly increased the expression of CYP1A1 and Interleukin1β (IL1B), and to a lesser extend UGT1, NQO1, and AhR Repressor (AhRR). Moreover, the increased expression of CYP1B1 was seen in diffuse GC, and IHC staining indicated that CYP1B1 is mainly expressed in stromal cells. TCDD treatment increased CYP1B1 expression in KATO III cells, although at lower levels as compared to CYP1A1. In intestinal GC, CYP1B1 expression is inversely correlated with several cancer-related genes such as IDO1, a gene involved in the early steps of tryptophan metabolism that contributes to the endogenous AhR ligand kynurenine expression. Altogether, our data provide evidence for a major role of AhR in GC, as an environmental xenobiotic receptor, through different mechanisms and pathways in diffuse and intestinal GC. Our results support the continued efforts to clarify the identity of exogenous AhR ligands in diffuse GC in order to define new therapeutic strategies.

Assessment of the Aryl Hydrocarbon Receptor-Mediated Effects of Aromatic Sensitizers in Paper Recycling Effluent Employing Zebrafish Embryos and in Silico Docking.

Aromatic sensitizers and related substances (SRCs), which are crucial in the paper industry for facilitating color-forming and color-developing chemical reactions, inadvertently contaminate effluents during paper recycling. Owing to their structural resemblance to endocrine-disrupting aromatic organic compounds, concerns have arisen about potential adverse effects on aquatic organisms. We focused on SRC effects via the aryl hydrocarbon receptor (AHR), employing molecular docking simulations and zebrafish (Danio rerio) embryo exposure assessments. Molecular docking revealed heightened binding affinities between certain SRCs in the paper recycling effluents and zebrafish Ahr2 and human AHR, which are pivotal components in the SRC toxicity mechanism. Fertilized zebrafish eggs were exposed to SRCs for up to 96 h post fertilization; among these substances, benzyl 2-naphthyl ether (BNE) caused morphological abnormalities, such as pericardial edema and shortened body length, at relatively low concentrations (1 μM) during embryogenesis. Gene expression of cytochrome P450 1A (cyp1a) and ahr2 was also significantly increased by BNE. Co-exposure to the AHR antagonist CH-223191 only partially mitigated BNE's phenotypic effects, despite the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin being relatively well restored by CH-223191, indicating BNE's AHR-independent toxic mechanisms. Furthermore, some SRCs, including BNE, exhibited in silico binding affinity to the estrogen receptor and upregulation of cyp19a1b gene expression. Therefore, additional insights into the toxicity of SRCs and their mechanisms are essential. The present results provide important information on SRCs and other papermaking chemicals that could help minimize the environmental impact of the paper industry. Environ Toxicol Chem 2024;43:2176-2188. © 2024 SETAC.

The augment effects of magnesium hydride on the lipid lowering effect of atorvastatin: an in vivo and in vitro investigation.

There is strong evidence connecting increased serum lipid levels to cardiovascular disorders, including atherosclerosis. Statins is prescribed as the primary medication to decrease lipid levels. Recent research has demonstrated that hydrogen possesses anti-inflammatory and antioxidant properties by modulating the expression of peroxisome proliferator-activated receptor gamma coactivator-1α, ultimately leading to the preservation of lipid homeostasis. Magnesium hydride (MgH2) is a prolonged stable hydrogen storage medium, which can be utilized to investigate its synergistic lipid-lowering effect with statins and its detailed molecular mechanism, both in vivo and in vitro. To ascertain the safety and efficacy of MgH2, we executed a comprehensive research of its influence on both physiological and pathological metrics. We noted a substantial diminution in lipid levels when MgH2 was integrated with atorvastatin, as attested by oil red staining. Furthermore, we scrutinized the regulatory effect of MgH2 on cytochrome P450 3A, which is a metabolic enzyme of statins, and discovered that it could be reduced by the MgH2. Concluding from our results, we propose that MgH2 inhibits the expression of cytochrome P450 3A in the liver and exerts an auxiliary lipid-lowering effect by increasing the blood concentration of statins. By augmenting our comprehension of MgH2's role in ameliorating lipid metabolism, we aspire to develop more promising therapies in the future.

3D spheroid HepaRG and fluorescent biphasic tracer for CYP3A4-mediated antibiotic interaction monitoring in sepsis.

Cytochrome P450 3A4 (CYP3A4) is a crucial enzyme in the metabolism of xenobiotics, particularly in drug metabolism interactions (DDIs), making it a significant factor in clinical drug use. However, current assay techniques are both laborious and costly, making it difficult to construct a high-throughput monitoring method that can be used in conjunction with the clinic. This poses certain safety hazards for drug combination. Therefore, it is crucial to develop a synchronized monitoring method for the inhibition and induction of CYP3A4. In this study, we utilized 3D culture technology to develop a HepaRG cells spheroid model. The CYP450 and transporter expression, the albumin secretion, and urea synthesis capacity characteristics were analyzed. The NEN probe was utilized as a tracer molecule for CYP3A4. The fluorescence intensity of metabolites was characterized by laser confocal technique to determine the inhibition and expression of CYP3A4 in the HepaRG cell spheroid model by the antibiotics for sepsis. The results indicate that the HepaRG sphere model successfully possessed the physiological phenotype of the liver, which could be used for drug interaction monitoring. Through positive drug testing, NEN probe was able to achieve bidirectional characterization of CYP3A4 induction and inhibition. The monitoring method described in this paper was successfully applied to drug interaction monitoring of commonly used antibiotics in sepsis patients, which is a convenient and rapid monitoring method. The proposed method offers a new strategy for monitoring CYP3A4-mediated drug-drug interactions with a high-throughput assay, which will help to improve the safety of clinical drug combination.

A novel approach to assess the health risk of aryl hydrocarbon receptor-bound contaminants via inhalation exposure using CYP1A1 expression as a biomarker

Polycyclic aromatic hydrocarbons (PAHs) and dioxins are potential causes of multiple diseases by activating the aryl hydrocarbon receptor (AhR) pathway. Health risk assessment of chemicals primarily relies on the relative potency factor (RPF), although its accuracy may be limited when solely using EC50 values. The induction of cytochrome P4501A1 (CYP1A1) serves as a biomarker for AhR activation and is an integrator of dioxin-like toxicity. Here, we present a method for evaluating the risks associated with AhR activation using mathematical models of dose-CYP1A1 induction. The dose-effect curves for certain PAHs and dioxins, including Ant, BghiP, 1,2,3,4,7,8-HxCDD, and others, exhibited a non-classical S-shaped form. The toxic equivalent factor (TEF) profiles revealed a broad range of toxic equivalent factor values. The TEFs for PAHs ranged from approximately 0.01 to 6, with higher values being observed when the concentration was less than 10−10 M, with the exceptions of Ace, Phe, and BghiP. Most congeners of dioxins got the lowest TEF value at around 10−10 M, ranging from 0.04 to 1.00. The binding affinity of AhR to ligands did not display a strong correlation with the EC50 of CYP1A1 expression, suggesting that the AhR-mediated effects of PAHs and dioxins are not fixed but instead fluctuate with the dose. Air samples acquired from a parking area were used to compare the proficiency of RPF and our current approach. In the current method, naphthalene and chrysene were the primary contributors of PAHs to AhR-mediated risks in parking lots air samples, respectively. However, the contributions of naphthalene and chrysene could be disregarded in the RPF approach.

Induction of hepatic CYP3A4 expression by cholesterol and cholic acid: Alterations of gene expression, microsomal activity, and pharmacokinetics.

Human cytochrome P450 3A4 (CYP3A4) is a drug-metabolizing enzyme that is abundantly expressed in the liver and intestine. It is an important issue whether compounds of interest affect the expression of CYP3A4 because more than 30% of commercially available drugs are metabolized by CYP3A4. In this study, we examined the effects of cholesterol and cholic acid on the expression level and activity of CYP3A4 in hCYP3A mice that have a human CYP3A gene cluster and show human-like regulation of the coding genes. A normal diet (ND, CE-2), CE-2 with 1% cholesterol and 0.5% cholic acid (HCD) or CE-2 with 0.5% cholic acid was given to the mice. The plasma concentrations of cholesterol, cholic acid and its metabolites in HCD mice were higher than those in ND mice. In this condition, the expression levels of hepatic CYP3A4 and the hydroxylation activities of triazolam, a typical CYP3A4 substrate, in liver microsomes of HCD mice were higher than those in liver microsomes of ND mice. Furthermore, plasma concentrations of triazolam in HCD mice were lower than those in ND mice. In conclusion, our study suggested that hepatic CYP3A4 expression and activity are influenced by the combination of cholesterol and cholic acid invivo.

Induction of human hepatic cytochrome P-450 3A4 expression by antifungal succinate dehydrogenase inhibitors

Succinate dehydrogenase inhibitors (SDHIs) are widely-used fungicides, to which humans are exposed and for which putative health risks are of concern. In order to identify human molecular targets for these agrochemicals, the interactions of 15 SDHIs with expression and activity of human cytochrome P-450 3A4 (CYP3A4), a major hepatic drug metabolizing enzyme, were investigated in vitro. 12/15 SDHIs, i.e., bixafen, boscalid, fluopyram, flutolanil, fluxapyroxad, furametpyr, isofetamid, isopyrazam, penflufen, penthiopyrad, pydiflumetofen and sedaxane, were found to enhance CYP3A4 mRNA expression in human hepatic HepaRG cells and primary human hepatocytes exposed for 48 h to 10 µM SDHIs, whereas 3/15 SDHIs, i.e., benzovindiflupyr, carboxin and thifluzamide, were without effect. The inducing effects were concentrations-dependent for boscalid (EC50=22.5 µM), fluopyram (EC50=4.8 µM) and flutolanil (EC50=53.6 µM). They were fully prevented by SPA70, an antagonist of the Pregnane X Receptor, thus underlining the implication of this xenobiotic-sensing receptor. Increase in CYP3A4 mRNA in response to SDHIs paralleled enhanced CYP3A4 protein expression for most of SDHIs. With respect to CYP3A4 activity, it was directly inhibited by some SDHIs, including bixafen, fluopyram, fluxapyroxad, isofetamid, isopyrazam, penthiopyrad and sedaxane, which therefore appears as dual regulators of CYP3A4, being both inducer of its expression and inhibitor of its activity. The inducing effect nevertheless predominates for these SDHIs, except for isopyrazam and sedaxane, whereas boscalid and flutolanil were pure inducers of CYP3A4 expression and activity. Most of SDHIs appear therefore as in vitro inducers of CYP3A4 expression in cultured hepatic cells, when, however, used at concentrations rather higher than those expected in humans in response to environmental or dietary exposure to these agrochemicals.

Epigenetic Activation of Cytochrome P450 1A2 Sensitizes Hepatocellular Carcinoma Cells to Sorafenib.

Cytochrome P450 1A2 (CYP1A2) is a known tumor suppressor in hepatocellular carcinoma (HCC), but its expression is repressed in HCC and the underlying mechanism is unclear. In this study, we investigated the epigenetic mechanisms of CYP1A2 repression and potential therapeutic implications. In HCC tumor tissues, the methylation rates of CYP1A2 CpG island (CGI) and DNA methyltransferase (DNMT) 3A protein levels were significantly higher, and there was a clear negative correlation between DNMT3A and CYP1A2 protein expression. Knockdown of DNMT3A by siRNA significantly increased CYP1A2 expression in HCC cells. Additionally, treating HCC cells with decitabine (DAC) resulted in a dose-dependent upregulation of CYP1A2 expression by reducing the methylation level of CYP1A2 CGI. Furthermore, we observed a decreased enrichment of H3K27Ac in the promoter region of CYP1A2 in HCC tissues. Treatment with the trichostatin A (TSA) restored CYP1A2 expression in HCC cells by increasing H3K27Ac levels in the CYP1A2 promoter region. Importantly, combination treatment of sorafenib with DAC or TSA resulted in a leftward shift of the dose-response curve, lower IC50 values, and reduced colony numbers in HCC cells. Our findings suggest that hypermethylation of the CGI at the promoter, mediated by the high expression of DNMT3A, and hypoacetylation of H3K27 in the CYP1A2 promoter region, leads to CYP1A2 repression in HCC. Epigenetic drugs DAC and TSA increase HCC cell sensitivity to sorafenib by restoring CYP1A2 expression. Our study provides new insights into the epigenetic regulation of CYP1A2 in HCC and highlights the potential of epigenetic drugs as a therapeutic approach for HCC. SIGNIFICANCE STATEMENT: This study marks the first exploration of the epigenetic mechanisms underlying cytochrome P450 (CYP) 1A2 suppression in hepatocellular carcinoma (HCC). Our findings reveal that heightened DNA methyltransferase expression induces hypermethylation of the CpG island at the promoter, coupled with diminished H3K27Ac levels, resulting in the repression of CYP1A2 in HCC. The use of epigenetic drugs such as decitabine and trichostatin A emerges as a novel therapeutic avenue, demonstrating their potential to restore CYP1A2 expression and enhance sorafenib sensitivity in HCC cells.

Jujuboside A Attenuates Polycystic Ovary Syndrome Based on Estrogen Metabolism Through Activating AhR-mediated CYP1A2 Expression.

Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders in women. This study aimed to investigate the therapeutic effects and mechanism of Jujuboside A on PCOS using a dehydroepiandrosterone (DHEA)-induced PCOS mouse model. Estrogen and androgen homeostasis was evaluated in serum from both clinical samples and PCOS mice. The stages of the estrous cycle were determined based on vaginal cytology. The ovarian morphology was observed by stained with hematoxylin and eosin. Moreover, we analyzed protein expression of cytochrome P450 1A1 (CYP1A1), cytochrome P450 1A2 (CYP1A2) and aryl hydrocarbon receptor (AhR) in ovary and KGN cells. Molecular docking, immunofluorescence, and luciferase assay were performed to confirm the activation of AhR by Jujuboside A. Jujuboside A effectively alleviated the disturbance of estrogen homeostasis and restored ovarian function, leading to an improvement in the occurrence and progression of PCOS. Furthermore, the protective effect of JuA against PCOS was dependent on increased CYP1A2 levels regulated by AhR. Our findings suggest that Jujuboside A improves estrogen disorders and may be a potential therapeutic agent for the treatment of PCOS.

Circ0087385 promotes DNA damage in benzo(a)pyrene-induced lung cancer development by upregulating CYP1A1.

Increasing environmental genotoxic chemicals have been shown to induce epigenetic alterations. However, the interaction between genetics and epigenetics in chemical carcinogenesis is still not fully understood. Here, we constructed an in vitro human lung carcinogenesis model (16HBE-T) by treating human bronchial epithelial cells with a typical significant carcinogen benzo(a)pyrene (BaP). We identified a novel circular RNA, circ0087385, which was overexpressed in 16HBE-T and human lung cancer cell lines, as well as in lung cancer tissues and serum exosomes from lung cancer patients. The upregulated circ0087385 after exposure to BaP promoted DNA damage in the early stage of chemical carcinogenesis and affected the cell cycle, proliferation, and apoptosis of the malignantly transformed cells. Overexpression of circ0087385 enhanced the expression of cytochrome P450 1A1 (CYP1A1), which is crucial for metabolically activating BaP. Interfering with circ0087385 or CYP1A1 reduced the levels of ultimate carcinogen benzo(a)pyrene diol epoxide (BPDE) and BPDE-DNA adducts. Interfering with CYP1A1 partially reversed the DNA damage induced by high expression of circ0087385, as well as decreased the level of BPDE and BPDE-DNA adducts. These findings provide novel insights into the interaction between epigenetics and genetics in chemical carcinogenesis which are crucial for understanding the epigenetic and genetic toxicity of chemicals.

The long noncoding RNA HNF1A-AS1 with dual functions in the regulation of cytochrome P450 3A4

Cytochrome P450 3A4 (CYP3A4) is the most important and abundant drug-metabolizing enzyme in the human liver. Inter-individual differences in the expression and activity of CYP3A4 affect clinical and precision medicine. Increasing evidence indicates that long noncoding RNAs (lncRNAs) play crucial roles in the regulation of CYP3A4 expression. Here, we showed that lncRNA hepatocyte nuclear factor 1 alpha–antisense 1 (HNF1A-AS1) exerted dual functions in regulating CYP3A4 expression in Huh7 and HepG2 cells. Mechanistically, HNF1A-AS1 served as an RNA scaffold to interact with both protein arginine methyltransferase 1 and pregnane X receptor (PXR), thereby facilitating their protein interactions and resulting in the transactivation of PXR and transcriptional alteration of CYP3A4 via histone modifications. Furthermore, HNF1A-AS1 bound to the HNF1A protein, a liver-specific transcription factor, thereby blocking its interaction with the E3 ubiquitin ligase tripartite motif containing 25, ultimately preventing HNF1A ubiquitination and protein degradation, further regulating the expression of CYP3A4. In summary, these results reveal the novel functions of HNF1A-AS1 as the transcriptional and post-translational regulator of CYP3A4; thus, HNF1A-AS1 may serve as a new indicator for establishing or predicting individual differences in CYP3A4 expression.

Etoricoxib enhances aryl hydrocarbon receptor activity

Etoricoxib is a nonsteroidal anti-inflammatory drug (NSAID) that possesses properties that include reducing inflammation and relieving pain and fever. Etoricoxib is an oral medication that selectively inhibits cyclooxygenase-2 with high efficacy. Controversies about its cardiovascular side effects have long existed. The aryl hydrocarbon receptor (AhR) is a cytoplasmic receptor that plays a key role in the metabolism of xenobiotics and many physiological functions. 2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) is a tryptophan metabolite and endogenous AhR agonist. Activation of AhR by its ligand induces upregulation of AhR-targeted cytochrome P450 (CYP) 1A1 expression. We found that etoricoxib (10–60 μM) induced CYP1A1 mRNA and protein expressions and the transcriptional activity of AhR mediated by the aryl hydrocarbon response element (AHRE) in both mouse Hepa-1c1c7 and human HepG2 cells. Its induction did not appear in AhR signaling-deficient cells, and was inhibited by the AhR antagonist, CH-223191. Etoricoxib was able to induced the translocalization of AhR from cytosol into nucleus. Etoricoxib also worked synergistically with ITE to further increase the expression of CYP1A1 mRNA and protein in human cells. The synergistic effect was higher in cells with than cells without overexpression of AhR. In summary, etoricoxib is an agonist of AhR in both mouse and human cells. Etoricoxib has a synergistic effect on ITE-induced CYP1A1 expression in human cells. The effect of etoricoxib on AhR and ITE on endothelial cells and cardiomyocytes should be further elucidated to in hope to clarify the mechanism of increased cardiovascular events in COX-2 inhibitors and etoricoxib.

CYP1A1 immunohistochemistry is highly specific for angiofibroma of soft tissue among morphological mimics.

Angiofibroma of soft tissue (AFST) is a benign, morphologically distinctive tumour type that harbours recurrent AHRR::NCOA2 fusions in 60-70% of cases and shows a non-specific immunophenotype, expressing EMA in roughly half of cases. The AHRR::NCOA2 fusion results in increased expression of cytochrome P450 1A1 (CYP1A1); a recent study demonstrated CYP1A1 immunohistochemistry (IHC) to be moderately sensitive and highly specific for AFST. In this study, we sought to validate these findings in a larger independent cohort of 30 AFST, as well as 215 morphological mimics, including 30 solitary fibrous tumours, 29 myxoid liposarcomas, 28 low-to-intermediate grade myxofibrosarcomas (MFS), 20 atypical spindle cell lipomatous tumours (ASCLT), 20 cellular angiofibromas, 10 cases each of spindle cell lipoma, neurofibroma, malignant peripheral nerve sheath tumour, superficial angiomyxoma, cellular myxoma, soft tissue perineurioma and deep fibrous histiocytoma, and nine cases each of low-grade fibromyxoid sarcoma and mammary-type myofibroblastoma. We found CYP1A1 IHC to be 70% sensitive for AFST, with granular cytoplasmic staining in 21 of 30 tumours, and 98% specific, with staining in only five morphological mimics: two deep fibrous histiocytomas, one MFS, one cellular angiofibroma and one ASCLT. These findings confirm that CYP1A1 is 70% sensitive, consistent with the prevalence of AHRR::NCOA2 fusions that up-regulate this protein, and that it is highly specific among morphological mimics.

Mulberry leaf supplementation inhibits skatole deposition by regulating gut microbiota and upregulating liver cytochrome P450 1A1 expression in finishing pigs

Skatole, a strong fecal odor substance, is generated through microbial degradation of tryptophan in the animal hindgut. It easily accumulates in adipose tissue and affects meat quality. In this study, the effect of mulberry leaf supplementation on skatole in finishing pigs was studied. In a 35-day trial, 20 finishing pigs (barrows and gilts) were fed with a basal diet or basal diet with 6% mulberry leaves. Growth performance of the pigs (n = 10) was automatically recorded by a performance-testing feeder system and 8 pigs in each treatment were slaughtered and sampled for the remaining tests. Skatole and short-chain fatty acids were detected using HPLC and gas chromatography, respectively. Fecal microbiota were analyzed using 16S rRNA gene sequencing. The metabolomics analysis of feces and serum was performed with UHPLC-MS/MS. The major cytochrome P450 (CYP) enzymes that catalyze skatole degradation in the liver were tested by using RT-PCR and Western blot. Effects of major bioactive compounds in mulberry leaves on the CYP genes were verified in the hepatic cell line HepG2 in an in vitro test (n = 3). In finishing pigs, mulberry leaf supplementation had no significant effect on the average daily gain, average daily feed intake, and feed conversion ratio (P > 0.05), but reduced skatole levels in feces, serum, and backfat (P < 0.05), and increased acetic acid levels in feces (P = 0.027). Mulberry leaf supplementation decreased the relative abundance of the skatole-producing bacteria Megasphaera and Olsenella (P < 0.05). Indole-3-acetic acid, the intermediate that is essential for skatole production, was significantly reduced in feces by mulberry leaf supplementation (P < 0.05) and was positively correlated with skatole content in feces (P = 0.004). In pigs treated with mulberry leaves, liver CYP1A1 expression was increased (P < 0.05) and was negatively correlated with skatole content in backfat (P = 0.045). The in vitro test demonstrated that mulberry leaf polyphenols and polysaccharides could directly stimulate CYP1A1 expression in hepatic cells. These findings suggest that mulberry leaf supplementation reduces skatole production and deposition in finishing pigs by regulating the gut microbiota and promoting skatole degradation in liver.

Ablation of Dual-Specificity Phosphatase 6 Protects against Nonalcoholic Fatty Liver Disease via Cytochrome P450 4A and Mitogen-Activated Protein Kinase

Dual-specificity phosphatase 6 (DUSP6) is a specific phosphatase for mitogen-activated protein kinase (MAPK). This study used a high-fat diet (HFD)-induced murine nonalcoholic fatty liver disease model to investigate the role of DUSP6 in this disease. Wild-type (WT) and Dusp6-haploinsufficiency mice developed severe obesity and liver pathology consistent with nonalcoholic fatty liver disease when exposed to HFD. In contrast, Dusp6-knockout (KO) mice completely eliminated these phenotypes. Furthermore, primary hepatocytes isolated from WT mice exposed to palmitic and oleic acids exhibited abundant intracellular lipid accumulation, whereas hepatocytes from Dusp6-KO mice showed minimal lipid accumulation. Transcriptome analysis revealed significant down-regulation of genes encoding cytochrome P450 4A (CYP4A), known to promote ω-hydroxylation of fatty acids and hepatic steatosis, in Dusp6-KO hepatocytes compared with that in WT hepatocytes. Diminished CYP4A expression was observed in the liver of Dusp6-KO mice compared with WT and Dusp6-haploinsufficiency mice. Knockdown of DUSP6 in HepG2, a human liver-lineage cell line, also promoted a reduction of lipid accumulation, down-regulation of CYP4A, and up-regulation of phosphorylated/activated MAPK. Furthermore, inhibition of MAPK activity promoted lipid accumulation in DUSP6-knockdown HepG2 cells without affecting CYP4A expression, indicating that CYP4A expression is independent of MAPK activation. These findings highlight the significant role of DUSP6 in HFD-induced steatohepatitis through two distinct pathways involving CYP4A and MAPK.

Persister cell phenotypes contribute to poor patient outcomes after neoadjuvant chemotherapy in PDAC

Neoadjuvant chemotherapy can improve the survival of individuals with borderline and unresectable pancreatic ductal adenocarcinoma; however, heterogeneous responses to chemotherapy remain a significant clinical challenge. Here, we performed RNA sequencing (n = 97) and multiplexed immunofluorescence (n = 122) on chemo-naive and postchemotherapy (post-CTX) resected patient samples (chemoradiotherapy excluded) to define the impact of neoadjuvant chemotherapy. Transcriptome analysis combined with high-resolution mapping of whole-tissue sections identified GATA6 (classical), KRT17 (basal-like) and cytochrome P450 3A (CYP3A) coexpressing cells that were preferentially enriched in post-CTX resected samples. The persistence of GATA6hi and KRT17hi cells post-CTX was significantly associated with poor survival after mFOLFIRINOX (mFFX), but not gemcitabine (GEM), treatment. Analysis of organoid models derived from chemo-naive and post-CTX samples demonstrated that CYP3A expression is a predictor of chemotherapy response and that CYP3A-expressing drug detoxification pathways can metabolize the prodrug irinotecan, a constituent of mFFX. These findings identify CYP3A-expressing drug-tolerant cell phenotypes in residual disease that may ultimately inform adjuvant treatment selection.

StemRegenin-1 Attenuates Endothelial Progenitor Cell Senescence by Regulating the AhR Pathway-Mediated CYP1A1 and ROS Generation.

Endothelial progenitor cell (EPC)-based stem cell therapy is a promising therapeutic strategy for vascular diseases. However, continuous in vitro expansion for clinical studies induces the loss of EPC functionality due to aging. In this study, we investigated the effects of StemRegenin-1 (SR-1), an antagonist of aryl hydrocarbon receptor (AhR), on replicative senescence in EPCs. We found that SR-1 maintained the expression of EPC surface markers, including stem cell markers, such as CD34, c-Kit, and CXCR4. Moreover, SR-1 long-term-treated EPCs preserved their characteristics. Subsequently, we demonstrated that SR-1 showed that aging phenotypes were reduced through senescence-associated phenotypes, such as β-galactosidase activity, SMP30, p21, p53, and senescence-associated secretory phenotype (SASP). SR-1 treatment also increased the proliferation, migration, and tube-forming capacity of senescent EPCs. SR-1 inhibited the AhR-mediated cytochrome P450 (CYP)1A1 expression, reactive-oxygen species (ROS) production, and DNA damage under oxidative stress conditions in EPCs. Furthermore, as a result of CYP1A1-induced ROS inhibition, it was found that accumulated intracellular ROS were decreased in senescent EPCs. Finally, an in vivo Matrigel plug assay demonstrated drastically enhanced blood vessel formation via SR-1-treated EPCs. In summary, our results suggest that SR-1 contributes to the protection of EPCs against cellular senescence.