CYP1B1 Expression Research Articles

Hepatic enzyme induction and its potential effect on thyroid hormone metabolism in the metamorphosing tadpole of Xenopus laevis (African clawed frog).

Hepatic enzyme induction, an inherent defense system against xenobiotics, is known to simultaneously affect endocrine system functions in mammals under specific conditions, particularly thyroid hormone (TH) regulation. While this phenomenon has been studied extensively, the pathway leading to this indirect thyroid effect in mammals has unclear applicability to amphibians, despite the importance of amphibian species in assessing thyroid-disruptive chemicals. Here, we investigated the effects of three well-known mammalian enzyme inducers-β-naphthoflavone (BNF), pregnenolone carbonitrile (PCN), and sodium phenobarbital (NaPB)-on the gene expression of phase-I and phase-II metabolizing enzymes in Xenopus laevis tadpoles. Waterborne exposure to BNF and PCN significantly induced the expression of both phase-I (cytochrome P450, CYP) and phase-II enzymes (UDP-glucuronosyltransferase, UGT and sulfotransferase, SULT), but in different patterns, while NaPB exposure induced CYP2B expression without affecting phase-II enzymes in tadpoles, in contrast to mammals. Furthermore, an ex vivo hepatic enzyme activity assay confirmed that BNF treatment significantly increased phase-II metabolic activity (glucuronidation and sulfation) toward TH. These results suggest the potential for certain mammalian enzyme inducers to influence TH clearance in X. laevis tadpoles. Our findings provide insights into the profiles of xenosensing activity and enzyme induction in amphibians, which can facilitate a better understanding of the mechanisms of indirect effects on the thyroid system via hepatic enzyme induction in nonmammalian species.

The uremic toxin indoxyl sulfate decreases osteocyte RANKL/OPG and increases wnt inhibitor RNA expression that is reversed by PTH

Abstract Renal osteodystrophy (ROD) leads to increased fractures, potentially due to underlying low bone turnover in chronic kidney disease (CKD). We hypothesized that indoxyl sulfate (IS), a circulating toxin elevated in CKD and ligand for the aryl hydrocarbon receptor (AhR), may target the osteocytes leading to bone cell uncoupling in ROD. The IDG-SW3 osteocytes were cultured for 14 days (early) and 35 days (mature osteocytes) and incubated with 500 μM of indoxyl sulfate after dose finding studies to confirm AhR activation. Long-term incubation of IS for 14 days led to decreased expression of Tnfsf11/Tnfrsf11b ratio (RANKL/OPG), which would increase osteoclast activity, and increased expression of Wnt inhibitors Sost and Dkk1, which would decrease bone formation in addition to decreased mineralization and alkaline phosphatase (ALP) activity. When osteocytes were incubated with IS and the AhR translocation inhibitor CH223191, mineralization and ALP activity were restored. However, the Tnfsf11/Tnfrsf11b ratio and Sost, Dkk1 expression were not altered compared to IS alone, suggesting more complex signaling. In both early and mature osteocytes, co-culture with PTH and IS reversed the IS-induced upregulation of Sost and Dkk1, and IS enhanced the PTH-induced increase of the Tnfsf11/Tnfrsf11b ratio. Co-culture of IS with PTH additively enhanced the AhR activity assessed by Cyp1a1 and Cyp1b1 expression. In summary, IS in the absence of PTH, increased osteocyte mRNA Wnt inhibitor expression in both early and mature osteocytes and decreased mRNA expression ofTnfsf11/Tnfrsf11b ratio and mineralization in early osteocytes. These changes would lead to decreased resorption and formation resulting in low bone remodeling. These data suggest IS may be important in the underlying low turnover bone disease observed in CKD when PTH is not elevated. In addition, when elevated PTH, IS interacts to further increase Tnfsf11/Tnfrsf11b ratio for osteoclast activity in both early and mature osteocytes which would worsen bone resorption.

The Expression of Genes CYP1A1, CYP1B1, and CYP2J3 in Distinct Regions of the Heart and Its Possible Contribution to the Development of Hypertension.

It is believed that alterations in the functioning of the cytochrome P450 (CYP), which participates in metabolic transformations of endogenous polyunsaturated fatty acids (PUFAs) (with the formation of cardioprotective or cardiotoxic products), affects the development of age-related cardiovascular diseases and reduces the effectiveness of some cardioselective drugs. For example, CYP2J2 activation or CYP1B1 inhibition protects against the cardiovascular toxicity of anticancer drugs. It is currently unclear whether CYPs capable of metabolizing arachidonic acid and ω-3 PUFAs to vasodilatory and vasoconstrictive derivatives are expressed in all heart regions. The work was performed on senescence-accelerated OXYS rats featuring elevated blood pressure, OXYSb rats (an OXYS substrain with normal blood pressure), and Wistar rats as a "healthy" control. The mRNA level was determined in the right and left ventricles, the right and left atria, and the aorta of 1-, 3-, and 12-month-old rats. We showed that all heart regions express CYPs capable of metabolizing arachidonic acid and ω-3 PUFAs and revealed significant differences between heart regions both in the mRNA level of genes CYP1B1, CYP2J3, and CYP1A1 and in the time course of expression changes with age. We noticed that expression levels of these CYPs in the heart regions and aorta differ between hypertensive OXYS rats, normotensive OXYSb rats, and healthy Wistar rats but could not detect any clear-cut patterns associated with the hypertensive status of OXYS rats.

Expression of pregnenolone-synthesizing enzymes CYP11A1 and CYP1B1 in the human, rat, and mouse brain

The central nervous system (CNS) is capable of synthesizing steroids for modulating essential functions such as neurotransmission, neuroplasticity, and neuroinflammation. These locally synthesized steroids, called neurosteroids, are produced through the conversion of cholesterol into the major steroid precursor pregnenolone, followed by downstream metabolism to form various steroids such as progesterone and allopregnanolone. Given that changes in neurosteroids are implicated in many neurological and psychiatric disorders, understanding the neurosteroidogenesis pathway is crucial. Recent studies have demonstrated an alternative pathway for the biosynthesis of pregnenolone, which is classically produced by CYP11A1 but was found instead to be made by CYP1B1 in human glial cells. However, numerous studies have demonstrated Cyp11a1 expression and activity in rodent brain tissue and brain cells. To elucidate whether species differences exist for the pregnenolone synthesis enzyme in human and rodent brains, we sought to directly compare the expression levels of CYP11A1 and CYP1B1 in human, rat, and mouse CNS tissues. We found that CYP1B1 mRNA expression was significantly higher than that of CYP11A1 in almost all CNS brain regions in human, rat, and mouse. The exception is in the mouse cerebral cortex, where Cyp11a1 RNA was more abundant than Cyp1b1. However, Cyp11a1 protein was clearly detectable in rodent CNS while completely undetectable in human brain. In contrast, the presence of CYP1B1 protein can be observed in both human and rodent brains. These results suggest that CYP1B1 is likely the dominant pregnenolone synthesis enzyme in the human brain, while rodent brains may use both Cyp11a1 and Cyp1b1.

Serotonin Availability Shapes the Effects of Phenelzine on Inflammatory Response and Gene Expression in Macrophages.

Background Serotonin (5-HT) is a neurotransmitter with extensive physiological influence in the central nervous system (CNS) and various behavioral and biological functions, including immune regulation through 5-HT receptors (5-HTRs) expressed by immune cells. A variety of serotonin-modulating drugs have been developed to treat neurological disorders. Phenelzine, a drug indicated for the management of treatment-resistant depression, is a potent, non-selectiveinhibitor of monoamine oxidase (MAO), the enzyme that metabolizes serotonin to 5-hydroxyindole acetic acid (5-HIAA). Given the emerging evidence of a bidirectional link between depression and inflammation, as well as the potential therapeutic applications of serotonin-modulating drugs in autoimmune diseases and cancer, our study investigated the pro-inflammatory and anti-inflammatory factors influenced by serotonin and phenelzine. Methodology We conducted experiments on RAW264.7 macrophages, exposing them to various combinations and concentrations of serotonin, 5-HIAA, and phenelzine. We assessed the relative gene expression of monoamine oxidase-A (MAO-A), CYP1B1, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) using real-time PCR and measured the production of IL-6, TNF-α, and IL-10 cytokines using enzyme-linked immunosorbent assays (ELISA). Results Our findings revealed that phenelzine can downregulate genes associated with the production of reactive oxygen and nitrogen species, reduce aryl hydrocarbon receptor (AHR)-related gene expression induced by serotonin, and enhance the production of inflammatory cytokines. These effects were significantly influenced by the concentration of available serotonin. Conclusions Our study demonstrates that various mechanisms, including AHR activation, modulation of reactive oxygen and nitrogen speciesproduction, and others, in addition to the increased availability of serotonin due to phenelzine treatment, can significantly influence the inflammatory state.

Induction of drug metabolizing enzyme and drug transporter expression by antifungal triazole pesticides in human HepaSH hepatocytes

Triazole pesticides are widely used fungicides, to which humans are rather highly exposed. They are known to activate drug-sensing receptors regulating expression of hepatic drug metabolizing enzymes and drug transporters, thus suggesting that the hepatic drug detoxification system is modified by these agrochemicals. To investigate this hypothesis, the effects of 9 triazole fungicides towards expression of drug metabolizing enzymes and transporters were characterized in cultured human HepaSH cells, that are human hepatocytes deriving from chimeric humanized liver TK-NOG mice. Most of triazoles used at 10 μM were found to act as inducers of cytochrome P-450 (CYP) 1A1, CYP1A2, CYP2B6, CYP3A4 and UDP-glucuronosyltransferase 1A1 mRNA levels and of CYP3A4 protein; some triazoles also enhanced mRNA expression of the canalicular transporters P-glycoprotein/MDR1, multidrug resistance-associated protein 2 and breast cancer resistance protein. Triazoles however concomitantly inhibited CYP2B6 and CYP3A4 activities and thus appeared as dual regulators of these CYPs, being both inducers of their expression and inhibitors of their activity. The inducing effect however predominated, at least for bromuconazole, propiconazole and tebuconazole. Bromuconazole was moreover predicted to enhance CYP2B6 and CYP3A4 expression in humans exposed to this fungicide in a chronic, acute or occupational context. These data demonstrate that key-actors of the human hepatic detoxification system are impacted by triazole pesticides, which may have to be considered for the risk assessment of these agrochemicals. They additionally highlight that the use of human HepaSH cells as surrogates to primary human hepatocytes represents an attractive and promising way for studying hepatic effects of environmental chemicals.

Honokiol Exhibits Anti-Tumor Effects in Breast Cancer by Modulating the miR-148a-5p-CYP1B1 Axis.

Breast cancer (BC) is the most frequently diagnosed malignancy in female patients. There is a significant lack of therapeutic strategies for BC, particularly triple-negative breast cancer (TNBC). Honokiol (HNK), a lignin extracted from the Magnolia genus plant, has demonstrated numerous pharmacological effects. Therefore, this study aims to investigate the antitumor effect of HNK on BC cells and employ high-throughput sequencing to elucidate its potential mechanism. We found that HNK significantly inhibited proliferation and induced apoptosis on BC cell lines in a dose-dependent manner. Moreover, HNK treatment suppressed migration and colony formation and initiated the intrinsic apoptotic pathway specifically in MDA-MB-231 cells. High-throughput sequencing and bioinformatics analysis revealed that miR-148a-5p expression was significantly up-regulated, whereas CYP1B1 expression was down-regulated following HNK treatment. Importantly, survival analysis based on TCGA database showed high miR-148a-5p expression was correlated with a better prognosis for BC patients. Inhibition of miR-148a-5p by inhibitor not only increased cell viability but also attenuated apoptosis induced by HNK.Finally, a strong synergistic effect between HNK and paclitaxel was observed in vitro. In conclusion, our study validated the antitumor efficacy of HNK against human BC cells and elucidated its underlying mechanism through high-throughput sequencing, thereby providing compelling evidence for further exploration of the potential clinical application of HNK towards the treatment of BC.

CYP1B1-RMDN2 Alzheimer’s disease endophenotype locus identified for cerebral tau PET

Determining the genetic architecture of Alzheimer’s disease pathologies can enhance mechanistic understanding and inform precision medicine strategies. Here, we perform a genome-wide association study of cortical tau quantified by positron emission tomography in 3046 participants from 12 independent studies. The CYP1B1-RMDN2 locus is associated with tau deposition. The most significant signal is at rs2113389, explaining 4.3% of the variation in cortical tau, while APOE4 rs429358 accounts for 3.6%. rs2113389 is associated with higher tau and faster cognitive decline. Additive effects, but no interactions, are observed between rs2113389 and diagnosis, APOE4, and amyloid beta positivity. CYP1B1 expression is upregulated in AD. rs2113389 is associated with higher CYP1B1 expression and methylation levels. Mouse model studies provide additional functional evidence for a relationship between CYP1B1 and tau deposition but not amyloid beta. These results provide insight into the genetic basis of cerebral tau deposition and support novel pathways for therapeutic development in AD.

CYP1B1 promotes PARPi-resistance via histone H1.4 interaction and increased chromatin accessibility in ovarian cancer

IntroductionOvarian cancer is the most lethal gynecological cancer and presents significant therapeutic challenges. The discovery of synthetic lethality between PARP inhibitors (PARPi) and homologous recombination deficiency marked a new era in treating BRCA1/2-mutated tumors. However, PARPi resistance remains a major clinical challenge. MethodsRNA sequencing was used to identify genes altered by PARPi treatment and LC-MS was used to detect proteins interacting with CYP1B1. Resistance mechanisms were explored through ATAC-seq and gene expression manipulation. Additional techniques, including micrococcal nuclease digestion assays, DAPI staining, and fluorescence microscopy, were used to assess changes in nuclear morphology and chromatin accessibility. ResultsThe gradual exposure of Olaparib has developed a PARPi-resistant cell line, A2780-OlaR, which exhibits significant upregulation of CYP1B1 at both RNA and protein levels. Down-regulating CYP1B1 expression or using specific inhibitors decreased the cellular response to Olaparib. Linker histone H1.4 was identified as associated with CYP1B1. ATAC-seq showed differential chromatin accessibility between A2780-OlaR and parental cells, indicating that the downregulation of H1.4 was associated with increased chromatin accessibility and higher cell viability after Olaparib treatment. ConclusionOur findings reveal a novel role for CYP1B1 in driving PARPi resistance through distinct molecular mechanisms in A2780-OlaR. This study highlights the importance of chromatin accessibility in PARPi efficacy and suggests the CYP1B1/H1.4 axis as a promising therapeutic target for overcoming drug resistance in ovarian cancer, offering potentially therapeutic benefits.

Long-term persistent exposure to cigarette smoke induces AhR driven corneal endothelial dysfunction in mice

Epidemiological studies show cigarette smoking enhances corneal endothelial dysfunction, but mechanisms remain unclear. Our study reveals that prolonged smoke exposure activates the aryl hydrocarbon receptor (AhR), increasing CYP1B1 expression and accelerating senescence and fibrosis in corneal endothelium, potentially reflecting adaptive responses to maintain corneal resilience. Although these molecular modifications indicate early endothelial dysfunction, no pathological changes were observed. The findings indicate that while chronic cigarette smoke exposure triggers initial molecular alterations and endothelial dysfunction, the progression to Fuchs endothelial corneal dystrophy likely requires additional environmental or genetic factors beyond smoke exposure alone.

EFFECTS OF ARYL HYDROCARBON RECEPTOR LIGAND TCDD ON HUMAN TROPHOBLAST CELL DEVELOPMENT.

The primary interface between mother and fetus, the placenta, serves two critical functions: extraction of nutrients from the maternal compartment and facilitation of nutrient delivery to the developing fetus. This delivery system also serves as a barrier to environmental exposures. The aryl hydrocarbon receptor (AHR) is an important component of the barrier. AHR signaling is activated by environmental pollutants and toxicants that can potentially affect cellular and molecular processes, including those controlling trophoblast cell development and function. In this study, we investigated the impact of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), an effective AHR ligand, exposure on human trophoblast cells. Human trophoblast stem (TS) cells were used as in vitro model system for investigating the downstream consequences of AHR activation. The actions TCDD were investigated in human TS cells maintained in the stem state or in differentiating TS cells. TCDD exposure stimulated the expression of CYP1A1 and CYP1B1 in human TS cells. TCDD was effective in stimulating CYP1A1 and CYP1B1 expression and altering gene expression profiles in human TS cells maintained in the stem cell state or induced to differentiate into extravillous trophoblast cells (EVT) or syncytiotrophoblast (ST). These actions were dependent upon the presence of AHR. TCDD exposure did not adversely affect maintenance of the TS cell stem state or the ability of TS cells to differentiate into EVT cells or ST. However, TCDD exposure did promote the biosynthesis of 2 methoxy estradiol (2ME), a biologically active catechol estrogen, with the potential to modify the maternal-fetal interface. Human trophoblast cell responses to TCDD were dependent upon AHR signaling and possessed the potential to shape development and function of the human placentation site.

Abstract Mo023: Inhibition of Cyp1a Protects Mice against Anthracycline Cardiomyopathy

Background: Anthracyclines such as doxorubicin (Dox) are highly effective anti-tumor agents, but their use is limited by dose-dependent cardiomyopathy. Our laboratory previously reported that activation of cytochrome P450 family 1 (Cyp1) enzymes contributes to acute doxorubicin (Dox) cardiotoxicity in zebrafish and in mice, and that potent Cyp1 inhibitors prevent cardiotoxicity. However, the role of Cyp1 enzymes in chronic Dox cardiomyopathy, as well as the mechanisms underlying cardioprotection associated with Cyp1 inhibition, have not been fully elucidated. Objective: This study aims to define the role of Cyp1 inhibition in mouse models that more accurately recapitulates the chronic cardiomyopathy seen in patients. Methods: The function of Cyp1 enzymes was evaluated using a small molecule Cyp1 inhibitor in wild-type (WT) mice, or Cyp1-null mice ( Cyp1a1/1a2 -/- , Cyp1b1 -/- , and Cyp1a1/1a2/1b1 -/- ). Low-dose Dox was administered by serial intraperitoneal or intravenous injections, respectively. Expression of Cyp1 isoforms was measured by RT-qPCR, and myocardial tissue was isolated from the left ventricle for RNA sequencing. Cardiac function was evaluated by transthoracic echocardiography. Results: In WT mice, Dox treatment was associated with a decrease in Cyp1a2 and an increase in Cyp1b1 expression in the heart and in the liver. Co-treatment of WT mice with Dox and the potent Cyp1 inhibitor YW-130 protected against cardiac dysfunction compared to Dox treatment alone. Cyp1a1/1a2 -/- and Cyp1a1/1a2/1b1 -/- mice were protected from Dox cardiomyopathy compared to WT mice. Male, but not female, Cyp1b1 -/- mice had increased cardiac dysfunction following Dox treatment compared to WT mice. RNA sequencing of myocardial tissue showed upregulation of Fundc1 and downregulation of Ccl21c in Cyp1a1/1a2 -/- mice treated with Dox, implicating changes in mitophagy and chemokine-mediated inflammation as possible mechanisms of Cyp1a-mediated cardioprotection. Conclusions: Taken together, this study highlights the potential therapeutic value of Cyp1a inhibition in mitigating anthracycline cardiomyopathy.

Exploring the role of air pollution on the development of atherosclerosis disorder in human endothelial cells

Abstract Funding Acknowledgements Type of funding sources: Public Institution(s). Main funding source(s): QU - NSPP Introduction Atherosclerosis, the main cause of cardiovascular and cerebrovascular diseases, is a global health threat. Rising cardiovascular disease prevalence, unexplained by traditional risk factors, prompts exploration of the impact of occupational exposure to environmental toxins as a risk factor. benzo[a]pyrene (BaP) is a polyaromatic compound that mediate its toxicity through activation of the Aryl hydrocarbon receptor (AhR). The molecular mechanisms of involvement of environmental toxins and the role of AhR/CYP1A pathway in atherosclerosis development remain not fully investigated. Purpose The current study explores the AhR pathway activation in endothelial cells upon exposure to environmental toxins in order to elucidate the molecular mechanisms of atherosclerotic cardiovascular diseases and plaque formation through investigating pro-inflammatory (CYP1A1, CYP1B1, TNF-a) and pro-apoptotic markers (BCL-2, Caspase-3). Methods EA. hy926 cell line underwent treatment with increasing concentrations of BaP (0-20 microM), as well as 10nm of TCDD, to confirm the results. DSMO was used as a control. Total RNA and protein were isolated to investigate mRNA expression and protein expression of AhR, CYP1A1, CYP1B1, as well as caspase-3, Bcl-2, and TNF-α using RT-PCR and Western Blot analysis. Statistical analysis was done using one-way ANOVA. Results The results showed that BaP induce significant dose-dependent upregulation of AhR and target genes (CYP1A1, CYP1B1). At 20uM, expression of AhR (3-fold), CYP1A1 (2.5-fold) and CYP1B1 (2-fold) was significantly high as compared to their respective controls and low doses. As for apoptotic markers, Bcl-2 was significantly downregulated at the highest concentrations 10uM and 20uM (0.7- and 0.6-fold, respectively). Caspase-3 showed no significant change but exhibited upregulation across all doses. TCDD 10nM significantly upregulated AhR (2.0-fold), CYP1A1 (1.5-fold), CYP1B1 (1.7-fold). Bcl-2 downregulated (0.6-fold), TNF-α significantly upregulated (1.8-fold) at 10nM. Caspase-3 increased 2-folds but was not statistically significant. Conclusion This study illustrated that AhR inducers, such as BaP, can accelerate the pathological progress of atherosclerosis by evoking oxidative stress and inflammation response molecules in human endothelial cells, demonstrated by the reduced expression of anti-apoptotic markers and the increased expression of inflammatory markers.

#1110 The AHR antagonist CH223191 alleviates kidney injury and renal fibrosis in adenine-induced chronic kidney disease

Abstract Background and Aims The aryl hydrocarbon receptor (AhR) is a transcription factor that can be activated by a variety of endogenous and exogenous ligands, and is involved in varieties of biological processes. It has been confirmed that the expression of AhR in kidney is elevated in chronic kidney disease (CKD). However, the specific role of AhR signaling pathway in CKD and its molecular mechanism remain unclear. This study aims to investigate the role and underlying mechanism of AhR in CKD, providing potential targets for the treatment of CKD. Method Male C57/B6L mice were randomly divided into control group, model group and treatment group. The model group was fed 0.2% adenine diet for 14 days to establish CKD model, while the treatment group was treated with AhR antagonist CH223191. AhR agonist kynurenine and antagonist CH223191 was applied in mouse tubular epithelial (TCMK-1) cells to investigate the underlying mechanisms of AhR in vitro. The renal function, histological changes and renal fibrosis in mice were evaluated to explore the role of AhR in adenine-induced CKD. Histological changes were evaluated by PAS and MASSON stainings, immunohistochemistry and immunofluorescence. Western blot and qPCR were used to measure protein/gene transcription levels. Results The levels of serum creatinine (p < 0.0001) and urea nitrogen (p < 0.0001) in the model group were significantly increased, with obvious renal tubule dilation and renal fibrosis by PAS and MASSON staining. CH223191 treatment significantly reduced serum creatinine (p=0.0012) and urea nitrogen ( < 0.0001) levels in mice, and alleviated renal fibrosis histologically. Q-PCR and Western-Blot results showed that the renal expression level of AhR in the treatment group was decreased, while the expression levels of fibronectin (Fn) (p < 0.0001), α-smooth muscle actin (α-SMA) (p < 0.0001), type I collagen (p=0.0007) and transforming growth factor-β (TGF-b) (p=0.0027) were significantly decreased. In addition, the expression levels of N-GAL (p < 0.0001) and KIM-1 ( < 0.0001) molecular markers associated with kidney injury were reduced, suggesting that inhibiting AhR expression in vivo can reduce adenine-mediated kidney injury and renal fibrosis in CKD. In vitro experiments indicated that kynurenic acid could induce the expression of AhR and AhR downstream regulatory gene CYP1b1 and upregulated fibrosis-related genes in tubular epithelial cells, while CH223191 treatment downregulated the fibrosis-related genes in tubular epithelial cells. Conclusion AhR mediates renal injury and renal fibrosis in adenine-induced CKD mice. AhR antagonist CH223191 exerts a renal protective effect by reducing the expression of AhR in renal tubular epithelial cells, thereby alleviating renal injury and renal fibrosis.

Exposure of cinnamyl alcohol in co-culture of BEAS-2B and dendritic cells: Interaction between CYP450 and cytokines.

The prevalence of fragrances in various hygiene products contributes to their sensorial allure. However, fragrances can induce sensitization in the skin or respiratory system, and the mechanisms involved in this process are incompletely understood. This study investigated the intricate mechanisms underlying the fragrance's effects on sensitization response, focusing on the interplay between CYP450 enzymes, a class of drug-metabolizing enzymes, and the adaptive immune system. Specifically, we assessed the expression of CYP450 enzymes and cytokine profiles in culture of BEAS-2B and mature dendritic cells (mDC) alone or in co-culture stimulated with 2 mM of a common fragrance, cinnamyl alcohol (CA) for 20 h. CYP1A1, CYP1A2, CYP1B1, CYP2A6, and CYP2A13 were analyzed by RT-PCR and IL-10, IL-12p70, IL-18, IL-33, and thymic stromal lymphopoietin (TSLP) by Cytometric Bead Array (CBA). Through RT-PCR analysis, we observed that CA increased CYP1A2 and CYP1B1 expression in BEAS-2B, with a further increased in BEAS-2B-mDC co-culture. Additionally, exposure to CA increased IL-12p70 levels in mDC rather than in BEAS-2B-mDC co-culture. In regards to IL-18, level was higher in BEAS-2B than in BEAS-2B-mDC co-culture. A positive correlation between the levels of IL-10 and CYP1B1 was found in mDC-CA-exposed and between IL-12p70 and CYP1A1 was found in BEAS-2B after CA exposure. However, IL-12p70 and CYP1A2 as well as IL-18, IL-33, and CYP1A1 levels were negative, correlated mainly in co-culture control. These correlations highlight potential immunomodulatory interactions and complex regulatory relationships. Overall, exposure to CA enhances CYP450 expression, suggesting that CA can influence immune responses by degrading ligands on xenosensitive transcription factors.

Cytochrome P450 1B1 is critical in the development of TNF-α, IL-6, and LPS-induced cellular hypertrophy.

Heart failure (HF) is preceded by cellular hypertrophy (CeH) which alters expression of cytochrome P450 enzymes (CYPs) and arachidonic acid (AA) metabolism. Inflammation is involved in CeH pathophysiology, but mechanisms remain elusive. This study investigates the impacts of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and lipopolysaccharides (LPS) on the development of CeH and the role of CYP1B1. AC16 cells were treated with TNF-α, IL-6, and LPS in the presence and absence of CYP1B1-siRNA or resveratrol. mRNA and protein expression levels of CYP1B1 and hypertrophic markers were determined using PCR and Western blot analysis, respectively. CYP1B1 enzyme activity was determined, and AA metabolites were analyzed using liquid chromatography-tandem mass spectrometry. Our results show that TNF-α, IL-6, and LPS induce expression of hypertrophic markers, induce CYP1B1 expression, and enantioselectively modulate CYP1B1-mediated AA metabolism in favor of mid-chain HETEs. CYP1B1-siRNA or resveratrol ameliorated these effects. In conclusion, our results demonstrate the crucial role of CYP1B1 in TNF-α, IL-6, and LPS-induced CeH.

Myostatin deletion alters hormone metabolism shifts in obese skeletal muscle of C57/Bl6 mice

Objective: Myostatin deletion improves cardiometabolic disease outcomes observed in obesity without any changes in activity in mice. Hormone balance is dysregulated in obesity, resulting in low testosterone and high estradiol levels, which have been shown to correlate with higher risk for developing associated cardiometabolic disease outcomes. However, the basis of these alterations and how they impact factors that improve obese cardiometabolic outcomes are unknown. Methods: Control and obese mice ( db/db) with or without myostatin deletion to induce hypermuscularity were used. Plasme from mice were analyzed for hormone levels via radioimmunoassay (RIA). Aromatase inhibition was performed via dosing 10mg/kg/day via water. Quantitative PCR was used cytochrome P450 enzymes involved in estrogen metabolism in the liver, skeletal muscle, and visceral white adipose tissue. C2C12 overexpression of CYP1B1 and treatment with hydroxyestrogens in vitro was performed and respective enzyme mRNA expression assessed. Results: Deletion of myostatin increased muscle mass by >30% and resulted in and improved glucose tolerance in the context of obesity without any changes in activity. Obesity decreased plasma testosterone 30% and increased estrogen 20% with no change in gonadal weight, which was improved in the absence of myostatin. CYP1B1, an enzyme that converts estradiol to toxic hydroxy estrogens, showed 2.5-fold expression in obese skeletal muscle, which is reduced in the absence of myostatin, with no change in other cytochrome P450s involved in estrogen metabolism. Inhibition of aromatase reduces estrogen and improves testosterone plasma concentration, indicating the excess estrogen in obesity is derived primarily from testosterone conversion. Aromatase expression was increased in obesity but unaffected by changes in muscle mass, this indicates an alteration to downstream estrogen metabolism. In C2C12 cells, overexpression of myostatin recapitulated changes in CYP1B1 expression seen in vivo, suggesting a direct link between the myokine and sex hormone metabolism. In addition, treatment of C2C12 cells with CYP1B1 derived hydroxyestrogens caused a tandem increase in SCD1 expression, indicating a causal link between CYP1B1 activity in obesity to increased triglyceride synthesis. Conclusion: In summary, obesity causes a shift in sex hormone levels towards estrogen production, which is improved by myostatin deletion. A potential mechanism responsible could be altered estrogen metabolism, leading to decreased estrogen levels without restoration of androgen levels. These data suggest CYP1B1 drives the conversion of estrogen to hydroxestrogens, known to harbor carcinogenic and pro-lipidemic influences, in skeletal muscle which may also contribute to cardiometabolic disease. Whether targeting CYP1B1 improves cardiometabolic function in obesity remains to be determined. D.W. Stepp and D.J. Fulton are supported by NIH 1R01HL147159. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Polycyclic aromatic hydrocarbon (PAH) activation of AhR inhibits glutathione S-transferase (GST) activity in primary human small airway epithelial cells (hSAECs)

Background: PAH represents the organic phase of particulate matter (PM) in air pollution emitted via the incomplete combustion of various compounds (i.e. diesel fuel, wood, and plastics). It is widely regarded that PAH activation of aryl hydrocarbon receptor (AhR) plays an important role in pro-inflammatory signaling, however the mechanisms remain unknown. In contrast- endogenous AhR agonists, such as 6-Formylindolo[3,2-b]carbazole (FICZ), exert pleiotropic protective effects against inflammation, fibrosis, and oxidative stress. The objective of our research program is to delineate the differences between activation of AhR by PAH (with adverse effects) compared to its beneficial homeostatic activation by endogenous FICZ. PM may induce oxidative stress in part through inhibition of antioxidant defense mechanisms. GSTs are a superfamily of enzymes that plays an important role in protecting cells from oxidative damage (by conjugating reduced glutathione to substrates under pro-oxidizing conditions). We hypothesize that PAH activation of AhR contributes to oxidative stress via inhibition of GST activity in primary hSAECs. Methods: Primary hSAECs were subjected to varying dosages of PAH [0, 250, 500, 1000, 5000 ng/mL] to model airborne PM exposure levels, and to increasing doses of FICZ [0, 0.50, 10, 100, 1000 nM] to model normal and excessively high endogenous ligand activation of AhR. We determined the relative potencies of PAH and FICZ by measuring CYP1A1 and CYP1B1 gene expression. Oxidative stress was measured using DHE-labeling and GST activity was measured with enzymatic kits (Sigma Aldrich). AhR activity was pharmacologically inhibited using 10μM CH223191. Statistical analysis was conducted using SigmaPlot 14.0 (Systat Software). Results: PAH and FICZ maximally stimulate hSAEC CYP1A1 and CYP1B1 expression at 1000ng/mL and 100nM, respectively, and are therefore defined as equipotent concentrations used in our studies. In a representative experiment in hSAECs, PAH (1000ng/mL) significantly increased hSAEC oxidative stress vs FICZ (100nM) n=6, p<0.01), while significantly decreasing GST specific activity vs equipotent FICZ activation of AhR in primary hSAECs from a 66-year-old female without lung disease (26.6±0.874 vs 28.3±.611 nmol/mL/min); n=6 from 2 independent assays; p=0.032. Similar differential effects of PAH vs FICZ activation of AhR on GST were seen in additional experiments with male hSAECs and Calu-3 cells (male). Importantly, CH223191 inhibition of GST enzymatic activity suggests that AhR plays an important role in protecting cells from oxidative stress. Conclusions: Our studies indicate that airborne PAH exposure can have detrimental health effects on the lung via aberrant AhR signaling. Significance: Understanding activity of AhR signaling can leads to novel therapeutic options for redox disorders in the lung. Funding: VA 5I01BX001777-05 (RP) and R01HL137033 (MH). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Genetic Signatures for Distinguishing Chemo-Sensitive from Chemo-Resistant Responders in Prostate Cancer Patients.

Prostate cancer remains a significant public health concern in sub-Saharan Africa, particularly impacting South Africa with high mortality rates. Despite many years of extensive research and significant financial expenditure, there has yet to be a definitive solution to prostate cancer. It is not just individuals who vary in their response to treatment, but even different nodules within the same tumor exhibit unique transcriptome patterns. These distinctions extend beyond mere differences in gene expression levels to encompass the control and networking of individual genes. Escalating chemotherapy resistance in prostate cancer patients has prompted increased research into its underlying mechanisms. The heterogeneous nature of transcriptomic organization among men makes the pursuit of universal biomarkers and one-size-fits-all treatments impractical. This study delves into the expression of drug resistance-associated genes, ABCB1 and CYP1B1, in cancer cells. Employing bioinformatics, we explored the molecular pathways and cascades linked to drug resistance following upregulation of these genes. Samples were obtained from archived prostate cancer patient specimens through pre-treatment biopsies of two categories: good vs. poor responders, with cDNAs synthesized from isolated RNAs subjected to qPCR analysis. The results revealed increased ABCB1 and CYP1B1 expression in tumor samples of the poor responders. Gene enrichment and network analysis associated ABCB1 with ABC transporters and LncRNA-mediated therapeutic resistance (WP3672), while CYP1B1 was linked to ovarian steroidogenesis, tryptophan metabolism, steroid hormone biosynthesis, benzo(a)pyrene metabolism, the sulindac metabolic pathway, and the estrogen receptor pathway, which are associated with drug resistance. Both ABCB1 and CYP1B1 correlated with microRNAs in cancer and the Nuclear Receptors Meta-Pathway. STRING analysis predicted protein-protein interactions of ABCB1 and CYP1B1 with Glutathione S-transferase Pi, Catechol O-methyltransferase, UDP-glucuronosyltransferase 1-6, Leucine-rich Transmembrane and O-methyltransferase (LRTOMT), and Epoxide hydrolase 1, with scores of 0.973, 0.971, 0.966, 0.966, and 0.966, respectively. Furthermore, molecular docking analysis of the chemotherapy drug, docetaxel, with CYP1B1 and ABCB1 revealed robust molecular interactions, with binding energies of -20.37 and -15.25 Kcal/mol, respectively. These findings underscore the susceptibility of cancer patients to drug resistance due to increased ABCB1 and CYP1B1 expression in tumor samples from patients in the poor-responders category that affects associated molecular pathways. The potent molecular interactions of ABCB1 and CYP1B1 with docetaxel further emphasize the potential basis for chemotherapy resistance.

Effects of dietary supplementation of different ratios of koumine and gelsemin on the liver immunity of common carp (Cyprinus carpio, FFRC)

Gelsemium elegans Benth. (GEB) exhibits immunoregulatory and growth-promoting effects in animals. However, owing to its complex composition, the specific mechanism remains to be explored. Koumine (KM) and gelsemine (GS) are the two alkaloids with the highest content and strongest biological activity in GEB. In this experiment, four ratios (0,0 mg/kg, 2,2 mg/kg, 2,1 mg/kg, and 2:0.67 mg/kg) of KM: GS from natural GEB were incorporated into the diet of common carp (Cyprinus carpio, FFRC). A 10-week breeding experiment was conducted to explore the immune and metabolic effects of these different KM and GS ratios on the liver of C. carpio, analyzing histology, antioxidant enzyme activity, immune gene expression, and transcriptomics. Hepatocyte edema and apoptosis were observed in the treatment groups. Acid phosphatase activity in the treatment group was significantly higher than that in the control group (P < 0.05), and malondialdehyde activity was lower (P < 0.05) in the treatment group, demonstrating a dose effect of GS. Superoxide dismutase and total antioxidant capacity activities increased gradually with a decreasing GS ratio (P < 0.05). Compared to the control group, changes in the GS ratio regulated the expression of immune genes. The relative expression of CYP1B1, TNF-β, and Myd-88 increased with decreasing GS ratio in the combination. Conversely, the relative expression of CYP1A1, TNF-α, GPX, IGF1, GST, and TLR-2 decreased with decreasing GS ratio in the combination. A total of 81.33 GB of clean data, 1370 DEGs, and 9258 co-expressed genes were obtained from the transcriptome data. These coexpressed genes were grouped into 244 KEGG pathways. Non-alcoholic fatty liver disease, Fatty acid degradation, Oxidative phosphorylation, and the PPAR signaling pathway were among the most abundant signaling pathways identified. This information contributes to a better understanding of the underlying mechanisms of KM and GS on liver immunity and lays a theoretical foundation for their application.