Inhibition of cytochrome P450 epoxygenase promotes endothelium-to-mesenchymal transition and exacerbates doxorubicin-induced cardiovascular toxicity.

BACKGROUND: Doxorubicin (DOX) is a potent chemotherapy widely used in treating various neoplastic diseases. However, the clinical use of DOX is limited due to its potential toxic effect on the cardiovascular system. Thus, identifying the pathway involved in this toxicity may help minimize chemotherapy risk and improve cancer patients' quality of life. Recent studies suggest that Endothelial-to-Mesenchymal transition (EndMT) and endothelial toxicity contribute to the pathogenesis of DOX-induced cardiovascular toxicity. However, the molecular mechanism is yet unknown. Given that arachidonic acid and associated cytochrome P450 (CYP) epoxygenase have been involved in endothelial and cardiovascular function, we aimed to examine the effect of suppressing CYP epoxygenases on DOX-induced EndMT and cardiovascular toxicity in vitro and in vivo. METHODS AND RESULTS: To test this, human endothelial cells were treated with DOX, with or without CYP epoxygenase inhibitor, MSPPOH. We also investigated the effect of MSPPOH on the cardiovascular system in our zebrafish model of DOX-induced cardiotoxicity. Our results showed that MSPPOH exacerbated DOX-induced EndMT, inflammation, oxidative stress, and apoptosis in our endothelial cells. Furthermore, we also show that MSPPOH increased cardiac edema, lowered vascular blood flow velocity, and worsened the expression of EndMT and cardiac injury markers in our zebrafish model of DOX-induced cardiotoxicity. CONCLUSION: Our data indicate that a selective CYP epoxygenase inhibitor, MSPPOH, induces EndMT and endothelial toxicity to contribute to DOX-induced cardiovascular toxicity.

Unlocking the secrets: exploring the influence of the aryl hydrocarbon receptor and microbiome on cancer development.

Gut microbiota regulates various aspects of human physiology by producing metabolites, metabolizing enzymes, and toxins. Many studies have linked microbiota with human health and altered microbiome configurations with the occurrence of several diseases, including cancer. Accumulating evidence suggests that the microbiome can influence the initiation and progression of several cancers. Moreover, some microbiotas of the gut and oral cavity have been reported to infect tumors, initiate metastasis, and promote the spread of cancer to distant organs, thereby influencing the clinical outcome of cancer patients. The gut microbiome has recently been reported to interact with environmental factors such as diet and exposure to environmental toxicants. Exposure to environmental pollutants such as polycyclic aromatic hydrocarbons (PAHs) induces a shift in the gut microbiome metabolic pathways, favoring a proinflammatory microenvironment. In addition, other studies have also correlated cancer incidence with exposure to PAHs. PAHs are known to induce organ carcinogenesis through activating a ligand-activated transcriptional factor termed the aryl hydrocarbon receptor (AhR), which metabolizes PAHs to highly reactive carcinogenic intermediates. However, the crosstalk between AhR and the microbiome in mediating carcinogenesis is poorly reviewed. This review aims to discuss the role of exposure to environmental pollutants and activation of AhR on microbiome-associated cancer progression and explore the underlying molecular mechanisms involved in cancer development.

Regulation of the aryl hydrocarbon receptor in cancer and cancer stem cells of gynecological malignancies: An update on signaling pathways.

Gynecological malignancies are a female type of cancers that affects the reproductive system. Cancer metastasis or recurrence mediated by cellular invasiveness occurs at advanced stages of cancer progression. Cancer Stem Cells (CSCs) enrichment in tumors leads to chemoresistance, which results in cancer mortality. Exposure to environmental pollutants such as polycyclic aromatic hydrocarbons is associated with an increased the risk of CSC enrichment in gynecological cancers. One of the important pathways that mediates the metabolism and bioactivation of these environmental chemicals is the transcription factor, aryl hydrocarbon receptor (AhR). The present review explores the molecular mechanisms regulating the crosstalk and interaction of the AhR with cancer-related signaling pathways, such as apoptosis, epithelial-mesenchymal transition, immune checkpoints, and G-protein-coupled receptors in several gynecological malignancies such as ovarian, uterine, endometrial, and cervical cancers. The review also discusses the potential of targeting the AhR pathway as a novel chemotherapy for gynecological cancers.

Epigenetic Regulation of Cancer Stem Cells by the Aryl Hydrocarbon Receptor Pathway.

Compelling evidence has demonstrated that tumor bulk comprises distinctive subset of cells generally referred as cancer stem cells (CSCs) that have been proposed as a strong sustainer and promoter of tumorigenesis and therapeutic resistance. These distinguished properties of CSCs have raised interest in understanding the molecular mechanisms that govern the maintenance of these cells. Numerous experimental and epidemiological studies have demonstrated that exposure to environmental toxins such as the polycyclic aromatic hydrocarbons (PAHs) is strongly involved in cancer initiation and progression. The PAH-induced carcinogenesis is shown to be mediated through the activation of a cytosolic receptor, aryl hydrocarbon receptor (AhR)/Cytochrome P4501A pathway, suggesting a possible direct link between AhR and CSCs. Several recent studies have investigated the role of AhR in CSCs self-renewal and maintenance, however the molecular mechanisms and particularly the epigenetic regulations of CSCs by the AhR/CYP1A pathway have not been reviewed before. In this review, we first summarize the crosstalk between AhR and cancer genetics, with a particular emphasis on the mechanisms relevant to CSCs such as Wnt/ß-catenin, Notch, NF-κB, and PTEN-PI3K/Akt signaling pathways. The second part of this review discusses the recent advances and studies highlighting the epigenetic mechanisms mediated by the AhR/CYP1A pathway that control CSC gene expression, self-renewal, and chemoresistance in various human cancers. Furthermore, the review also sheds light on the importance of targeting the epigenetic pathways as a novel therapeutic approach against CSCs.

6-Formylindolo(3,2-b)carbazole Dampens Inflammation and Reduces Endotoxin-Induced Kidney Injury via Nrf2 Activation.

Patients with sepsis are at a high risk of morbidity and mortality due to multiple organ injuries caused by pathological inflammation. Although sepsis is accompanied by multiple organ injuries, acute renal injury is a significant contributor to sepsis morbidity and mortality. Thus, dampening inflammation-induced renal injury may limit severe consequences of sepsis. As several studies have suggested that 6-formylindolo(3,2-b)carbazole (FICZ) is beneficial for treating various inflammatory diseases, we aimed to examine the potential protective effect of FICZ on the acute endotoxin-induced sepsis model of kidney injury. To test this, male C57Bl/6N mice were injected with FICZ (0.2 mg/kg) or vehicle 1 h prior to an injection of either lipopolysaccharides (LPS) (10 mg/kg), to induce sepsis, or phosphate-buffered saline for 24 h. Thereafter, gene expression of kidney injury and pro-inflammatory markers, circulating cytokines and chemokines, and kidney morphology were assessed. Our results show that FICZ reduced LPS-induced acute injury in kidneys from LPS-injected mice. Furthermore, we found that FICZ dampens both renal and systemic inflammation in our sepsis model. Mechanistically, our data indicated that FICZ significantly upregulates NAD(P)H quinone oxidoreductase 1 and heme oxygenase 1 via aryl hydrocarbon receptor (AhR) and nuclear factor erythroid 2-related factor 2 (Nrf2) in the kidneys to lessen inflammation and improve septic acute kidney injury. Overall, the data of our study show that FICZ possesses a beneficial reno-protective effect against sepsis-induced renal injury via dual activation of AhR/Nrf2.

Insight into the physiological and pathological roles of the aryl hydrocarbon receptor pathway in glucose homeostasis, insulin resistance, and diabetes development.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcriptional factor that mediates the toxicities of several environmental pollutants. Decades of research have been carried out to understand the role of AhR as a novel mechanism for disease development. Its involvement in the pathogenesis of cancer, cardiovascular diseases, rheumatoid arthritis, and systemic lupus erythematosus have long been known. One of the current hot research topics is investigating the role of AhR activation by environmental pollutants on glucose homeostasis and insulin secretion, and hence the pathogenesis of diabetes mellitus. To date, epidemiological studies have suggested that persistent exposure to environmental contaminants such as dioxins, with subsequent AhR activation increases the risk of specific comorbidities such as obesity and diabetes. The importance of AhR signaling in various molecular pathways highlights that the role of this receptor is far beyond just xenobiotic metabolism. The present review aims at providing significant insight into the physiological and pathological role of AhR and its regulated enzymes, such as cytochrome P450 1A1 (CYP1A1) and CYP1B1 in both types of diabetes. It also provides a comprehensive summary of the current findings of recent research studies investigating the role of the AhR/CYP1A1 pathway in insulin secretion and glucose hemostasis in the pancreas, liver, and adipose tissues. This review further highlights the molecular mechanisms involved, such as gluconeogenesis, hypoxia-inducible factor (HIF), oxidative stress, and inflammation.

Aryl Hydrocarbon Receptor Promotes Cell Growth, Stemness Like Characteristics, and Metastasis in Human Ovarian Cancer via Activation of PI3K/Akt, ß-Catenin, and Epithelial to Mesenchymal Transition Pathways.

Ovarian cancer (OC) ranks first in cancer-related deaths out of all female reproductive malignancies with high-pitched tumor relapse and chemoresistance. Several reports correlate cancer occurrences with exposure to xenobiotics via induction of a protein receptor named aryl hydrocarbon receptor (AhR). However, the effect of AhR on OC proliferation, expansion, and chemoresistance remains unrevealed. For this purpose, OC cells A2780 and A2780cis cells were treated with AhR activator, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and the effects were determined by Real-Time Cell Analyzer, clonogenic assay, flow cytometry, immunoblotting and wound healing assay. Our results showed that activation of AhR by TCDD in A2780 cells induced the PI3K/AKT pathway followed by induction of anti-apoptotic proteins BCL-2, BCL-xl, and MCL-1. In addition, a significant increase in stemness marker aldehyde dehydrogenase (ALDH1) was observed. This effect was also associated with an accumulation of ß-catenin, a Wnt transcription factor. Moreover, we observed induction of epithelial to mesenchymal transition (EMT) upon AhR activation. In conclusion, the results from the current study confirm that AhR mediates OC progression, stemness characteristics, and metastatic potential via activation of PI3K/Akt, Wnt/ß-catenin, and EMT. This study provides a better insight into the modulatory role of AhR that might help in developing novel therapeutic strategies for OC treatment.

Metformin attenuates V-domain Ig suppressor of T-cell activation through the aryl hydrocarbon receptor pathway in Melanoma: In Vivo and In Vitro Studies.

Melanoma is an aggressive skin cancer with a high rate of metastasis to other organs. Recent studies specified the overexpression of V-domain Ig suppressor of T-cell activation (VISTA) and Aryl Hydrocarbon Receptor (AHR) in melanoma. Metformin shows anti-tumor activities in several cancer types. However, the mechanism is unclear. This study aims to investigate the inhibitory effect of metformin on VISTA via AHR in melanoma cells (CHL-1, B16) and animal models. VISTA and AHR levels were assessed by qPCR, Western blot, immunofluorescence microscope, flow cytometry, and immunohistochemistry. Here, metformin significantly decreased VISTA and AHR levels in vitro and in vivo. Furthermore, metformin inhibited all AHR-regulated genes. VISTA levels were dramatically inhibited by AHR modulations using shRNA and αNF, confirming the central role of AHR in VISTA. Finally, melanoma cells were xenografted in C57BL/6 and nude mice. Metformin significantly reduced the tumor volume and growth rate. Likewise, VISTA and AHR-regulated protein levels were suppressed in both models. These findings demonstrate for the first time that VISTA is suppressed by metformin and identified a new regulatory mechanism through AHR. The data suggest that metformin could be a new potential therapeutic strategy to treat melanoma patients combined with targeted immune checkpoint inhibitors.

Influence of the Aryl Hydrocarbon Receptor Activating Environmental Pollutants on Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is an umbrella term that includes many different disorders that affect the development, communication, and behavior of an individual. Prevalence of ASD has risen exponentially in the past couple of decades. ASD has a complex etiology and traditionally recognized risk factors only account for a small percentage of incidence of the disorder. Recent studies have examined factors beyond the conventional risk factors (e.g., environmental pollution). There has been an increase in air pollution since the beginning of industrialization. Most environmental pollutants cause toxicities through activation of several cellular receptors, such as the aryl hydrocarbon receptor (AhR)/cytochrome P450 (CYPs) pathway. There is little research on the involvement of AhR in contributing to ASD. Although a few reviews have discussed and addressed the link between increased prevalence of ASD and exposure to environmental pollutants, the mechanism governing this effect, specifically the role of AhR in ASD development and the molecular mechanisms involved, have not been discussed or reviewed before. This article reviews the state of knowledge regarding the impact of the AhR/CYP pathway modulation upon exposure to environmental pollutants on ASD risk, incidence, and development. It also explores the molecular mechanisms involved, such as epigenesis and polymorphism. In addition, the review explores possible new AhR-mediated mechanisms of several drugs used for treatment of ASD, such as sulforaphane, resveratrol, haloperidol, and metformin.

EGFR Inhibitor Gefitinib Induces Cardiotoxicity through the Modulation of Cardiac PTEN/Akt/FoxO3a Pathway and Reactive Metabolites Formation: In Vivo and in Vitro Rat Studies.

Gefitinib (GEF) is a selective inhibitor of the epidermal growth factor receptor (EGFR) used to treat non-small cell lung cancer. Yet, few cases of cardiotoxicity have been reported. However, the role of the PTEN/Akt/FoxO3a pathway, which mediates GEF anticancer activity, in GEF cardiotoxicity remains unclear. For this purpose, in vitro H9c2 cells and in vivo rat cardiomyocytes were utilized as study models. Treatment of H9c2 cells and Sprague-Dawley rats with GEF significantly induced the expression of hypertrophic and apoptotic markers at mRNA and protein levels with an increased plasma level of troponin. This was accompanied by induction of autophagy and mitochondrial dysfunction in H9c2 cells. Inhibition of cardiac EGFR activity and Akt cellular content of in vitro and in vivo rat cardiomyocytes by GEF increased PTEN and FoxO3a gene expression and cellular content. Importantly, treatment of H9c2 cells with PI3K/Akt inhibitor increased PTEN and FoxO3a mRNA expression associated with potentiation of GEF cardiotoxicity. In addition, by using LC-MS/MS, we showed that GEF is metabolized in the rat heart microsomes into one cyanide- and two methoxylamine-adduct reactive metabolites, where their formation was entirely blocked by CYP1A1 inhibitor, α-naphthoflavone. The current study concludes that GEF induces cardiotoxicity through modulating the expression and function of the cardiac PTEN/AKT/FoxO3a pathway and the formation of CYP1A1-mediated reactive metabolites.

The role of NF-κB and AhR transcription factors in lead-induced lung toxicity in human lung cancer A549 cells.

Lead (Pb) is recognized as the first heavy metal of the top six toxic air pollutants threatening human health and the second hazardous substance. Pb exposure is associated with lung impairment and high incidences of lung cancer. Nuclear factor kappa B (NF-κB) and aryl hydrocarbon receptor (AhR) signaling pathways are known to be expressed and play an important role in the lung. However, the link between Pb lung toxicity and NF-κB and/or AhR pathways remains unclear. This study was established to explore the role of NF-κB and AhR modulation in Pb-induced lung toxicity in human lung cancer A549 cells. In the current study, treatment of A549 cells with Pb significantly induced cell apoptosis as evidenced by increasing a) the percentage of cells underwent apoptosis determined by flow cytometry and b) p53 mRNA level. Pb treatment induced oxidative stress by a) increasing the formation of reactive oxygen species and b) decreasing GSTA1 mRNA levels. The toxic effects of Pb on the lung was associated with significant increases in NF-κB and AhR levels which was accompanied with increases in downstream targets genes, iNOS and CYP1A1, respectively. Inhibition of NF-κB or AhR either chemically using resveratrol or genetically using small interfering RNA (siRNA) significantly rescued A549 cells from Pb-mediated lung toxicity. The results clearly indicate that Pb-mediated lung toxicities are NF-κB and AhR-dependent mechanism.

Phloretin attenuates STAT-3 activity and overcomes sorafenib resistance targeting SHP-1-mediated inhibition of STAT3 and Akt/VEGFR2 pathway in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common primary liver malignancy. Phloretin (PH) possesses anticancer, antitumor, and hepatoprotective effects, however, the effects and potential mechanisms of phloretin remain elusive. METHODS: Five HCC cells were tested in vitro for sensitivity to PH, Sorafenib (Sor) or both and the apoptosis, signal transduction and phosphatase activity were analyzed. To validate the role of SHP-1, we used PTP inhibitor III and SHP-1 siRNA. Further, we used purified SHP-1 proteins or HCC cells expressing deletion N-SH2 domain or D61A point mutants to study the PH efficacy on SHP-1. The `in vivo studies were conducted using HepG2 and SK-Hep1 and Sor resistant HepG2SR and Huh7SR xenografts. Molecular docking was done with Swiss dock and Auto Dock Vina. RESULTS: PH inhibited cell growth and induced apoptosis in all HCC cells by upregulating SHP-1 expression and downregulating STAT3 expression and further inhibited pAKT/pERK signaling. PH activated SHP-1 by disruption of autoinhibition of SHP-1, leading to reduced p-STAT3Tyr705 level. PH induced apoptosis in two Sor-resistant cell lines and overcome STAT3, AKT, MAPK and VEGFR2 dependent Sor resistance in HCCs. PH potently inhibited tumor growth in both Sor-sensitive and Sor-resistant xenografts in vivo by impairing angiogenesis, cell proliferation and inducing apoptosis via targeting the SHP-1/STAT3 signaling pathway. CONCLUSION: Our data suggest that PH inhibits STAT3 activity in Sor-sensitive and -resistant HCCs via SHP-1-mediated inhibition of STAT3 and AKT/mTOR/JAK2/VEGFR2 pathway. Our results clearly indicate that PH may be a potent reagent for hepatocellular carcinoma and a noveltargeted therapy for further clinical investigations.

Endoplasmic Reticulum Stress: A Critical Molecular Driver of Endothelial Dysfunction and Cardiovascular Disturbances Associated with Diabetes.

Physical inactivity and sedentary lifestyle contribute to the widespread epidemic of obesity among both adults and children leading to rising cases of diabetes. Cardiovascular disease complications associated with obesity and diabetes are closely linked to insulin resistance and its complex implications on vascular cells particularly endothelial cells. Endoplasmic reticulum (ER) stress is activated following disruption in post-translational protein folding and maturation within the ER in metabolic conditions characterized by heavy demand on protein synthesis, such as obesity and diabetes. ER stress has gained much interest as a key bridging and converging molecular link between insulin resistance, oxidative stress, and endothelial cell dysfunction and, hence, represents an interesting drug target for diabetes and its cardiovascular complications. We reviewed here the role of ER stress in endothelial cell dysfunction, the primary step in the onset of atherosclerosis and cardiovascular disease. We specifically focused on the contribution of oxidative stress, insulin resistance, endothelial cell death, and cellular inflammation caused by ER stress in endothelial cell dysfunction and the process of atherogenesis.

Meteorological Factors and Seasonal Stroke Rates: A Four-year Comprehensive Study.

INTRODUCTION: There is a growing body of evidence suggesting that acute cardiovascular events including stroke are not distributed randomly over time but instead depend on months/season of the year. We report the impact of meteorological variables in extremely hot and arid climate on stroke. METHODS: Acute stroke patients admitted from January 2014 to December 2017 were included. The data included demographics, clinical risk factors, temperature, solar radiation, relative humidity, dew point, wind speed, and atmospheric pressure. We calculated stroke rates/100,000/month. RESULTS: There were 3654 cases of stroke (ischemic stroke [IS]: 2956 [80.9%]; and intracerebral hemorrhage [ICH]: 698 [19.1%]) with no difference in hematocrit, creatinine, and blood urea between hot and cold seasons (p > .05). We observed a positive significant correlation of IS with the mean temperature (AOR: 1.023; 95% CI: 1.009-1.036; P = .001) and mean solar radiation (AOR: 1.268; 95% CI: 1.021-1.575; P = .032) showing a 2.3% and 26.8% higher risk relative to ICH respectively, a negative correlation between IS with relative humidity (AOR: 0.99; 95% CI: 0.984-0.997; P = .002), and atmospheric pressure (AOR: 0.977; 95% CI: 0.966-0.989; P < .001) was observed, 1% increase in the relative humidity correlate with 2.4% and 1% lower risk of IS incidence relative to ICH respectively. CONCLUSION: We demonstrated a distinct seasonal pattern in the incidence of stroke with an increase in IS rates relative to ICH during the summer months with higher solar radiations that cannot be explained by physiological measures suggestive of dehydration or hem-concentration.

Genotoxic impact of long-term cigarette and waterpipe smoking on DNA damage and oxidative stress in healthy subjects.

Although a plethora of studies have examined tobacco smoke-cancer disease association, the involvement of cellular genetic toxicity remains unclear. Therefore, the present study provides molecular evidence for a pathway involved in the DNA damage induced by long-term cigarette and waterpipe smoke in human subjects. The study population consisted of 45 subjects who were divided into three groups; healthy nonsmokers group, cigarette smokers group, and waterpipe smokers group. A questionnaire and consent form was distributed and signed by all participants. Total RNA was extracted from the blood using PAXgene Blood RNA Kit and mRNA expression levels of target genes were quantified by RT-PCR. Our results showed that 80% of the participants smoke 20-39 cigarettes/day, whereas 12% smoke more than 40 cigarettes/day. With regard to waterpipe smoke, the majority (46%) smoke more than 5 times/week. Both cigarette and waterpipe smokers showed increased the plasma levels 8-hydroxy-2'-deoxyguanosine (8-OHdG), of DNA damage marker. In addition, the mRNA expression levels of DNA repair genes (OGG1 and XRCC1) were significantly inhibited in both cigarette and waterpipe smokers groups by 30% and 60%, respectively. This was associated with a marked decrease (50%) in the expression of detoxifying genes (NQO1 and GSTA1) with an increase in CYP1A1 mRNA expression, a cancer-activating gene. Both cigarette and waterpipe smokers increased in the plasma concentrations of several toxic heavy metals such as Cd (130%), Pb (47%), and Ni (30%). In conclusion: the present findings clearly explore the genotoxic effect of cigarette and waterpipe smoking on human DNA.

Gold-containing compound BDG-I inhibits the growth of A549 lung cancer cells through the deregulation of miRNA expression.

Gold complex bis(diethyldithiocarbamato-gold(I)) bis(diphenylphosphino) methane (BDG-I) is cytotoxic toward different cancer cell lines. We compared the cytotoxic effect of BDG-I with that of cisplatin in the A549 lung cancer cell line. Additionally, we investigated the molecular mechanism underlying the toxic effect of BDG-I toward the A549 cell line and the identification of cancer-related miRNAs likely to be involved in killing the lung cancer cells. Further, X-ray crystallographic data of the compound were acquired. Using microarray, global miRNA expression profiling in BDG-I-treated A549 cells revealed 64 upregulated and 86 downregulated miRNAs, which targeted 4689 and 2498 genes, respectively. Biological network connectivity of the miRNAs was significantly higher for the upregulated miRNAs than for the downregulated miRNAs. Two of the 10 most upregulated miRNAs (hsa-mir-20a-5p and hsa-mir-15b-5p) were associated with lung cancer. AmiGo2 server and Panther pathway analyses indicated significant enrichment in transcription regulation of miRNA target genes that promote intrinsic kinase-mediated signaling, TGF-ß, and GnRH signaling pathways, as well as oxidative stress responses. BDG-I crystal structure X-ray diffraction studies revealed gold-gold intramolecular interaction [Au…Au = 3.1198 (3) Å] for a single independent molecule, reported to be responsible for its activity against cancer. Our present study sheds light on the development of novel gold complex with favorable anti-cancer therapeutic functionality.

Aryl hydrocarbon receptor/cytochrome P450 1A1 pathway mediates breast cancer stem cells expansion through PTEN inhibition and ß-Catenin and Akt activation.

BACKGROUND: Breast cancer stem cells (CSCs) are small sub-type of the whole cancer cells that drive tumor initiation, progression and metastasis. Recent studies have demonstrated a role for the aryl hydrocarbon receptor (AhR)/cytochrome P4501A1 pathway in CSCs expansion. However, the exact molecular mechanisms remain unclear. METHODS: The current study was designed to a) determine the effect of AhR activation and inhibition on breast CSCs development, maintenance, self-renewal, and chemoresistance at the in vitro and in vivo levels and b) explore the role of ß-Catenin, PI3K/Akt, and PTEN signaling pathways. To test this hypothesis, CSC characteristics of five human breast cancer cells; SKBR-3, MCF-7, and MDA-MB231, HS587T, and T47D treated with AhR activators or inhibitor were determined using Aldefluor assay, side population, and mammosphere formation. The mRNA, protein expression, cellular content and localization of the target genes were determined by RT-PCR, Western blot analysis, and Immunofluorescence, respectively. At the in vivo level, female Balb/c mice were treated with AhR/CYP1A1 inducer and histopathology changes and Immunohistochemistry examination for target proteins were determined. RESULTS: The constitutive mRNA expression and cellular content of CYP1A1 and CYP1B1, AhR-regulated genes, were markedly higher in CSCs more than differentiating non-CSCs of five different human breast cancer cells. Activation of AhR/CYP1A1 in MCF-7 cells by TCDD and DMBA, strong AhR activators, significantly increased CSC-specific markers, mammosphere formation, aldehyde dehydrogenase (ALDH) activity, and percentage of side population (SP) cells, whereas inactivation of AhR/CYP1A1 using chemical inhibitor, α-naphthoflavone (α-NF), or by genetic shRNA knockdown, significantly inhibited the upregulation of ALDH activity and SP cells. Importantly, inactivation of the AhR/CYP1A1 significantly increased sensitization of CSCs to the chemotherapeutic agent doxorubicin. Mechanistically, Induction of AhR/CYP1A1 by TCDD and DMBA was associated with significant increase in ß-Catenin mRNA and protein expression, nuclear translocation and its downstream target Cyclin D1, whereas AhR or CYP1A1 knockdown using shRNA dramatically inhibited ß-Catenin cellular content and nuclear translocation. This was associated with significant inhibition of PTEN and induction of total and phosphorylated Akt protein expressions. Importantly, inhibition of PI3K/Akt pathway by LY294002 completely blocked the TCDD-induced SP cells expansion. In vivo, IHC staining of mammary gland structures of untreated and DMBA (30 mg/kg, IP)- treated mice, showed tremendous inhibition of PTEN expression accompanied with an increase in the expression p-Akt, ß-Catenin and stem cells marker ALDH1. CONCLUSIONS: The present study provides the first evidence that AhR/CYP1A1 signaling pathway is controlling breast CSCs proliferation, development, self-renewal and chemoresistance through inhibition of the PTEN and activation of ß-Catenin and Akt pathways.

Therapeutic potential of carfilzomib, an irreversible proteasome inhibitor, against acetaminophen-induced hepatotoxicity in mice.

Overdose of acetaminophen (APAP) is often associated with hepatotoxicity. Carfilzomib (CFZ) shows multiple pharmacological activities including anti-inflammatory potential. Therefore, this study was undertaken to evaluate the possible therapeutic effects of CFZ against APAP-induced hepatotoxicity. Hepatotoxicity was induced by administration of APAP (350 mg/kg, intraperitoneal). Mice were given CFZ (0.125, 0.25, or 0.5 mg/kg, intraperitoneal) 1.5 h after APAP administration. Animals were sacrificed on 6 h and blood and liver tissue samples were collected for analysis. In CFZ-post-treated group, there was significant and dose-dependent decrease in serum alanine aminotransferase levels. The level of tumor necrosis factor-α (TNF-α), reactive oxygen species, and NO decreased, whereas glutathione increased significantly by CFZ post-treatment. Upregulated mRNA expression of COX-II and iNOS were significantly downregulated by CFZ post-treatment. CFZ may exert its hepatoprotective action by alleviating inflammatory, oxidative, and nitrosative stress via inhibition of TNF-α, COX-II, and iNOS.

Dexamethasone Attenuates LPS-induced Acute Lung Injury through Inhibition of NF-κB, COX-2, and Pro-inflammatory Mediators.

Dexamethasone (DEX) is a synthetic glucocorticoid with potent anti-inflammatory effects that is widely used to treat inflammatory diseases. The aim of the present study was to investigate the possible protective effect of DEX on the lipopolysaccharides (LPS)-induced acute lung injury (ALI) in a mouse model. Animals were pretreated with DEX (5 and 10 mg/kg, i.p.) for seven days and acute lung injury was induced by intranasal (i.n.) administration of LPS on day 7. In the present study, administration of LPS resulted in significant increase in neutrophils and lymphocytes count whereas a substantial reduction in T cell subsets (CD3(+) and CD4(+)) and pro-inflammatory (IL-6 and TNF-α) cytokines occurred, which were reversed by DEX treatment. RT-PCR analysis revealed an increased mRNA expression of IL-6, TNF-α, COX-2, iNOS, and NF-κB p65 and decreased IL-10 in the LPS group, which were reversed by treatment with DEX in lung tissues. Western blot analysis revealed an increased expression of COX-2, iNOS and NF-κB p65 in the LPS-group, which was reduced by treatment with DEX. Compared with the LPS group, the DEX treatment also demonstrated a considerable increase in the protein expression level of IL-10 cytokine. Administration of LPS resulted in marked increase in malondialdehyde (MDA) levels and myeloperoxidase (MPO) activity whereas noticeable decrease in glutathione (GSH) content. These changes were significantly reversed by treatment with DEX. The histological examinations revealed protective effect of DEX while LPS group aggravated lung injury. The present findings demonstrate the potent anti-inflammatory action of the DEX against acute lung injury induced by LPS.

Metformin inhibits 7,12-dimethylbenz[a]anthracene-induced breast carcinogenesis and adduct formation in human breast cells by inhibiting the cytochrome P4501A1/aryl hydrocarbon receptor signaling pathway.

Recent studies have established that metformin (MET), an oral anti-diabetic drug, possesses antioxidant activity and is effective against different types of cancer in several carcinogen-induced animal models and cell lines. However, whether MET can protect against breast cancer has not been reported before. Therefore, the overall objectives of the present study are to elucidate the potential chemopreventive effect of MET in non-cancerous human breast MCF10A cells and explore the underlying mechanism involved, specifically the role of cytochrome P4501A1 (CYP1A1)/aryl hydrocarbon receptor (AhR) pathway. Transformation of the MCF10A cells into initiated breast cancer cells with DNA adduct formation was conducted using 7,12-dimethylbenz[a]anthracene (DMBA), an AhR ligand. The chemopreventive effect of MET against DMBA-induced breast carcinogenesis was evidenced by the capability of MET to restore the induction of the mRNA levels of basic excision repair genes, 8-oxoguanine DNA glycosylase (OGG1) and apurinic/apyrimidinic endonuclease1 (APE1), and the level of 8-hydroxy-2-deoxyguanosine (8-OHdG). Interestingly, the inhibition of DMBA-induced DNA adduct formation was associated with proportional decrease in CYP1A1 and in NAD(P)H: quinone oxidoreductase 1 (NQO1) gene expression. Mechanistically, the involvements of AhR and nuclear factor erythroid 2-related factor-2 (Nrf2) in the MET-mediated inhibition of DMBA-induced CYP1A1 and NQO1 gene expression were evidenced by the ability of MET to inhibit DMBA-induced xenobiotic responsive element and antioxidant responsive element luciferase reporter gene expression which suggests an AhR- and Nrf2-dependent transcriptional control. However, the inability of MET to bind to AhR suggests that MET is not an AhR ligand. In conclusion, the present work shows a strong evidence that MET inhibits the DMBA-mediated carcinogenicity and adduct formation by inhibiting the expression of CYP1A1 through an AhR ligand-independent mechanism.