-
The Science of the Total Environment Jan 2022In this study, we identified major aryl hydrocarbon receptor (AhR) agonists in the sediments from Yeongil Bay (n = 6) using effect-directed analysis. Using the...
In this study, we identified major aryl hydrocarbon receptor (AhR) agonists in the sediments from Yeongil Bay (n = 6) using effect-directed analysis. Using the H4IIE-luc bioassays, great AhR-mediated potencies were found in aromatic fractions (F2) of sediment organic extracts from silica gel column chromatography and sub-fractions (F2.6-F2.8) from reverse phase-HPLC. Full-scan mass spectrometric analysis using GC-QTOFMS was conducted to identify novel AhR agonists in highly potent fractions, such as F2.6-F2.8 of S1 (Gumu Creek). Selection criteria for AhR-active compounds consisted of three steps, including matching factor of NIST library (≥70), aromatic structures, and the number of aromatic rings (≥4). Fifty-nine compounds were selected as tentative AhR agonist candidates, with the AhR-mediated activity being assessed for six compounds for which standard materials were available commercially. Of these compounds, 20-methylcholanthrene, 7-methylbenz[a]anthracene, 10-methylbenz[a]pyrene, and 7,12-dimethylbenz[a]anthracene exhibited significant AhR-mediated potency. Relative potency values of these compounds were determined relative to benzo[a]pyrene to be 3.2, 1.4, 1.2, and 0.2, respectively. EPA positive matrix factorization modeling indicated that the sedimentary AhR-active aromatic compounds primarily originated from coal combustion and vehicle emissions. Potency balance analysis indicated that four novel AhR agonists explained 0.007% to 1.7% of bioassay-derived AhR-mediated potencies in samples.
Topics: Biological Assay; Environmental Monitoring; Geologic Sediments; Polycyclic Aromatic Hydrocarbons; Receptors, Aryl Hydrocarbon
PubMed: 34481160
DOI: 10.1016/j.scitotenv.2021.149969 -
Chemosphere Jan 20223-methylcholanthrene (3 MC) is an environmental compound belonging to the PAHs and is reportedly thought to be a risk factor for the prevalence of hepatic function...
3-methylcholanthrene (3 MC) is an environmental compound belonging to the PAHs and is reportedly thought to be a risk factor for the prevalence of hepatic function disorder. Here, a dose of 0.5 mg/kg of 3 MC was given to 4-week-old male and female mice (F0) in their diet for 6 weeks. After exposure, then the mice were mated between different groups. The first filial (F1) generation offspring of exposed or unexposed parental mice were sacrificed at the age of 5 weeks (F1-5 W), and the potential effects on the F0 and F1 offspring were evaluated. The results showed that the total bile acids (TBAs) in the serum and feces in F0 females and female F1-5 W individuals born from female mice exposed to 3 MC decreased, while the TBAs in the liver increased. The transcriptional levels of major genes participating in synthesis, regulation, transportation and apical uptake was also altered correspondingly. In addition, the transcription of some genes related to inflammation was enhanced in these mice. Further investigation revealed that in addition to distinct changes in the mucus secretion, tight junction proteins and ion transport were induced, and antimicrobial peptides were also disrupted in the intestine of F0 mice and F1-5 W female offspring of maternal mice exposed to 3 MC. Our results suggested that exposure to 3 MC, but not male exposure, had the potential to interfere with BAs metabolism, affecting gut barrier function. Females were more seriously affected than males.
Topics: Animals; Bile Acids and Salts; Enterohepatic Circulation; Female; Liver; Male; Methylcholanthrene; Mice; Reproduction
PubMed: 34346331
DOI: 10.1016/j.chemosphere.2021.131681 -
Biochemical and Biophysical Research... Sep 2021The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that regulates various toxicological and biological functions. We reported previously that...
Activation of the aryl hydrocarbon receptor by 3-methylcholanthrene, but not by indirubin, suppresses mammosphere formation via downregulation of CDC20 expression in breast cancer cells.
The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that regulates various toxicological and biological functions. We reported previously that 3-methylcholanthrene (3MC), an exogenous AhR agonist, inhibited tumorsphere (mammosphere) formation from breast cancer cell lines, while the endogenous AhR agonist, indirubin, very weakly inhibited this process. However, the difference in inhibition mechanism of mammosphere formation by 3MC or indirubin is still unknown. In this study, we established AhR-re-expressing (KOTR-AhR) cells from AhR knockout MCF-7 cells using the tetracycline (Tet)-inducible gene expression systems. To identify any difference in inhibition of mammosphere formation by 3MC or indirubin, RNA-sequencing (RNA-seq) experiments were performed using KOTR-AhR cells. RNA-seq experiments revealed that cell division cycle 20 (CDC20), which regulates the cell cycle and mitosis, was decreased by 3MC, but not by indirubin, in the presence of AhR expression. Furthermore, the mRNA and protein levels of CDC20 were decreased by 3MC in MCF-7 cells via the AhR. In addition, mammosphere formation was suppressed by small interfering RNA-mediated CDC20 knockdown compared to the negative control in MCF-7 cells. These results suggest that AhR activation by 3MC suppresses mammosphere formation via downregulation of CDC20 expression in breast cancer cells. This study provides useful information for the development of AhR-targeted anti-cancer drugs.
Topics: Breast Neoplasms; Cdc20 Proteins; Cell Line, Tumor; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Humans; Indoles; Methylcholanthrene; Receptors, Aryl Hydrocarbon; Spheroids, Cellular; Transcriptome
PubMed: 34280616
DOI: 10.1016/j.bbrc.2021.07.047 -
Journal of Plant Physiology Aug 2021The environmental spread of hydrophobic pollutants has been receiving attention because of specific characteristics of these compounds that make them resistant to...
The environmental spread of hydrophobic pollutants has been receiving attention because of specific characteristics of these compounds that make them resistant to degradation, thus causing various toxic effects on humans as a result of their bioaccumulation. Here, we report the role of zucchini major-latex like proteins (MLPs) on the accumulation of hydrophobic pollutants, as consumption of contaminated crops is one of the main routes for accumulation. Transgenic tobacco plants expressing an aryl hydrocarbon receptor (AhR) gene with a β-glucuronidase (GUS) inducible expression system were transformed with one of the three zucchini MLP genes (PG1, GR1, and GR3). MLP transgenic plants showed a significant increase in the fold induction of GUS activity compared to the parental AhR tobacco plants when one of the most toxic polychlorinated biphenyl (PCB) congeners, 3,3',4,4',5-pentachlorobiphenyl (CB126), was applied. GUS activity was detected in both aerial parts and roots after treatment with the strong carcinogen 3-methylcholanthrene. Phenotypic changes in the MLP tobacco during incubation with CB126 were also observed. The MLP transgenic plant PG1 responded to treatment with 0.32 nM CB126, whereas vector control plants significantly induced GUS activity at 200 nM CB126. Moreover, GUS activities in the MLP plants treated with other PCB congeners were significantly higher than those in the plants given the mock treatment. As GUS activities in the aerial parts of the plants were significantly correlated with the accumulation level of PCBs, these results strongly suggest that zucchini MLPs are related to the translocation of hydrophobic pollutants from the roots to the aerial parts through their binding affinity.
Topics: Biological Transport; Crops, Agricultural; Gene Expression Regulation, Plant; Genes, Plant; Hydrophobic and Hydrophilic Interactions; Plant Roots; Plants, Genetically Modified; Polychlorinated Biphenyls; Soil Pollutants; Nicotiana
PubMed: 34225177
DOI: 10.1016/j.jplph.2021.153464 -
Journal For Immunotherapy of Cancer Jun 2021NY-ESO-1 is a tumor-specific, highly immunogenic, human germ cell antigen of the MAGE-1 family that is a promising vaccine and cell therapy candidate in clinical trial...
BACKGROUND
NY-ESO-1 is a tumor-specific, highly immunogenic, human germ cell antigen of the MAGE-1 family that is a promising vaccine and cell therapy candidate in clinical trial development. The mouse genome does not encode an NY-ESO-1 homolog thereby not subjecting transgenic T-cells to thymic tolerance mechanisms that might impair in-vivo studies. We hypothesized that an NY-ESO-1 T cell receptor (TCR) transgenic mouse would provide the unique opportunity to study avidity of TCR response against NY-ESO-1 for tumor vaccine and cellular therapy development against this clinically relevant and physiological human antigen.
METHODS
To study in vitro and in vivo the requirements for shaping an effective T cell response against the clinically relevant NY-ESO-1, we generated a C57BL/6 HLA-A*0201 background TCR transgenic mouse encoding the 1G4 TCR specific for the human HLA-A2 restricted, NY-ESO-1 SLLMWITQC (9C), initially identified in an NY-ESO-1 positive melanoma patient.
RESULTS
The HLA-A*0201 restricted TCR was positively selected on both CD4 and CD8 cells. Mouse 1G4 T cells were not activated by endogenous autoimmune targets or a large library of non-cognate viral antigens. In contrast, their activation by HLA-A2 NY-ESO-1 complexes was evident by proliferation, CD69 upregulation, interferon-γ production, and interleukin-2 production, and could be tuned using a twofold higher affinity altered peptide ligand, NY-ESO-1. NY-ESO-1 recombinant vaccination of syngeneic mice adoptively transferred with m1G4 CD8 T cells controlled tumor growth in vivo. 1G4 transgenic mice suppressed growth of syngeneic methylcholanthrene (MCA) induced HHD tumor cells expressing the full-length human NY-ESO-1 protein but not MCA HHD tumor cells lacking NY-ESO-1.
CONCLUSIONS
The 1G4 TCR mouse model for the physiological human TCR against the clinically relevant antigen, NY-ESO-1, is a valuable tool with the potential to accelerate clinical development of NY-ESO-1-targeted T-cell and vaccine therapies.
Topics: Animals; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cancer Vaccines; Cell Line, Tumor; Cell Proliferation; Cell Survival; HLA-A2 Antigen; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neoplasm Proteins; Peptide Fragments; Receptors, Antigen, T-Cell; Thymoma; Thymus Neoplasms; Xenograft Model Antitumor Assays
PubMed: 34088742
DOI: 10.1136/jitc-2021-002544 -
BMC Cancer May 2021Despite considerable medical proceedings, cancer is still a leading cause of death. Major problems for tumor therapy are chemoresistance as well as toxic side effects....
BACKGROUND
Despite considerable medical proceedings, cancer is still a leading cause of death. Major problems for tumor therapy are chemoresistance as well as toxic side effects. In recent years, the additional treatment with the antidiabetic drug metformin during chemotherapy showed promising results in some cases. The aim of this study was to develop an in vitro tumor therapy model in order to further investigate the potential of a combined chemotherapy with metformin.
METHODS
Cytotoxic effects of a combined treatment on BALB/c fibroblasts were proven by the resazurin assay. Based on the BALB/c cell transformation assay, the BALB/c tumor therapy model was established successfully with four different and widely used chemotherapeutics from different categories. Namely, Doxorubicin as a type-II isomerase inhibitor, Docetaxel as a spindle toxin, Mitomycin C as an alkylating agent and 5-Fluorouracil as an antimetabolite. Moreover, glucose consumption in the medium supernatant was measured and protein expressions were determined by Western Blotting.
RESULTS
Initial tests for the combined treatment with metformin indicated unexpected results as metformin could partly mitigate the cytotoxic effects of the chemotherapeutic agents. These results were further confirmed as metformin induced resistance to some of the drugs when applied simultaneously in the tumor therapy model. Mechanistically, an increased glucose consumption was observed in non-transformed cells as well as in the mixed population of malignant transformed cell foci and non-transformed monolayer cells, suggesting that metformin could also increase glucose consumption in transformed cells.
CONCLUSION
In conclusion, this study suggests a cautious use of metformin during chemotherapy. Moreover, the BALB/c tumor therapy model offers a potent tool for further mechanistic studies of drug-drug interactions during cancer therapy.
Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; BALB 3T3 Cells; Carcinogens; Cell Survival; Cell Transformation, Neoplastic; Culture Media; Docetaxel; Doxorubicin; Drug Evaluation, Preclinical; Drug Interactions; Fluorouracil; Glucose; Humans; Metformin; Methylcholanthrene; Mice; Mitomycin; Neoplasms
PubMed: 34044797
DOI: 10.1186/s12885-021-08354-x -
Toxicology in Vitro : An International... Aug 2021This study evaluates the impact of physiologically relevant oxygen tensions on the response of HepG2 cells to known inducers and hepatotoxic drugs. We compared...
This study evaluates the impact of physiologically relevant oxygen tensions on the response of HepG2 cells to known inducers and hepatotoxic drugs. We compared transcriptional regulation and CYP1A activity after a 48 h exposure at atmospheric culture conditions (20% O) with representative periportal (8% O) and perivenous (3% O) oxygen tensions. We evaluated cellular responses in 2D and 3D cultures at each oxygen tension in parallel, using monolayers and a paper-based culture platform that supports cells suspended in a collagen-rich environment. Our findings highlight that the toxicity, potency, and mechanism of action of drugs are dependent on both culture format and oxygen tension. HepG2 cells in 3D environments at physiologic oxygen tensions better matched primary human hepatocyte data than HepG2 cells cultured under standard conditions. Despite altered transcriptional regulation with decreasing oxygen tensions, we did not observe the zonation patterns of drug-metabolizing enzymes found in vivo. Our approach demonstrates that oxygen is an important regulator of liver function but it is not the sole regulator. It also highlights the utility of the 3D paper-based culture platform for continued mechanistic studies of microenvironmental influences on cellular responses.
Topics: Acetaminophen; Aflatoxin B1; Arylsulfotransferase; Cell Culture Techniques; Cell Hypoxia; Cell Survival; Cyclophosphamide; Cytochrome P-450 Enzyme System; Gene Expression Regulation, Enzymologic; Glucuronosyltransferase; Hep G2 Cells; Humans; Methylcholanthrene; Oxygen; Polychlorinated Dibenzodioxins
PubMed: 33811995
DOI: 10.1016/j.tiv.2021.105156 -
Biological & Pharmaceutical Bulletin 2021We had previously reported that treatment with the aryl hydrocarbon receptor (AHR) agonist β-naphthoflavone (βNF) suppressed mammosphere formation derived from cancer...
The Differential Selectivity of Aryl Hydrocarbon Receptor (AHR) Agonists towards AHR-Dependent Suppression of Mammosphere Formation and Gene Transcription in Human Breast Cancer Cells.
We had previously reported that treatment with the aryl hydrocarbon receptor (AHR) agonist β-naphthoflavone (βNF) suppressed mammosphere formation derived from cancer stem cells in human breast cancer MCF-7 cells (Cancer Lett., 317, 2012, Zhao et al.). Here, using several AHR agonists, we have investigated the association of this suppression with the classical ability to induce AHR-mediated gene transcription in the xenobiotic response element (XRE). The mammosphere formation assays were performed using wild-type and AHR-knockout MCF-7 cells in the presence of AHR agonists including 3-methylcholanthrene (3MC), benzo[a]pyrene (BaP), 7,12-dimethylbenz[a]anthracene (DMBA), 6-formylindolo[3,2-b]carbazole (FICZ), indirubin, indole-3-carbinol (I3C), indole-3-acetic acid (IAA), and kynurenine (KYN), followed by the XRE-reporter gene assays of the agonists. We showed that treatments with 3MC, BaP, and DMBA strongly suppressed mammosphere formation of the stem cells in an AHR-dependent manner, while other agonists showed weaker suppression. In reporter gene assays, the strength or duration of AHR/XRE-mediated gene transcription was found to be dependent on the agonist. Although strong transcriptional activation was observed with 3MC, FICZ, indirubin, I3C, IAA, or KYN after 6 h of treatment, only weak activation was seen with BaP or DMBA. While transcriptional activation was sustained or increased at 24 h with 3MC, BaP, or DMBA, appreciable reduction was observed with the other agonists. In conclusions, the results demonstrated that the suppressive effects of AHR agonists on mammosphere formation do not necessarily correlate with their abilities to induce AHR-mediated gene transcription. Hence, different AHR functions may be differentially induced in an agonist-dependent manner.
Topics: Basic Helix-Loop-Helix Transcription Factors; Breast Neoplasms; Cell Survival; Gene Knockout Techniques; Genes, Reporter; Humans; Indoles; Kynurenine; MCF-7 Cells; Polycyclic Aromatic Hydrocarbons; Receptors, Aryl Hydrocarbon; Transcription, Genetic
PubMed: 33790107
DOI: 10.1248/bpb.b20-00961 -
Methods in Cell Biology 2021Mouse models of cancer are essential in furthering our understanding both of the mechanisms that drive tumor development and the immune response that develops in...
Mouse models of cancer are essential in furthering our understanding both of the mechanisms that drive tumor development and the immune response that develops in parallel, and also in providing a platform for testing novel anti-cancer therapies. The majority of solid tumor models available rely on the injection of existing cancer cell lines into naïve hosts which, while providing quick and reproducible model systems, typically lack the development of a tumor microenvironment that recapitulates those seen in human cancers. Administration of the carcinogen 3-methylcholanthrene (MCA), allows tumors to develop in situ, forming a tumor microenvironment with an established stroma and vasculature. This article provides a detailed set of protocols for the administration of MCA into mice and the subsequent monitoring of tumors. Protocols are also provided for some of the routinely used downstream applications that can be used for MCA tumors.
Topics: Animals; Disease Models, Animal; Fibrosarcoma; Immunity; Methylcholanthrene; Mice; Tumor Microenvironment
PubMed: 33785169
DOI: 10.1016/bs.mcb.2020.09.007 -
Methods in Cell Biology 2021Lung cancer is one of the deadliest types of cancer and as such requires disease models that are useful for identification of novel pathways for biomarkers as well as to...
Lung cancer is one of the deadliest types of cancer and as such requires disease models that are useful for identification of novel pathways for biomarkers as well as to test therapeutic agents. Adenocarcinoma (ADC), the most prevalent type of lung cancer, is a subtype of non-small cell lung carcinoma (NSCLC) and a disease driven mainly by smoking. However, it is also the most common subtype of lung cancer found in non-smokers with environmental exposures. Chemically driven models of lung cancer, also called primary models of lung cancer, are important because they do not overexpress or delete oncogenes or tumor suppressor genes, respectively, to increase oncogenesis. Instead these models test tumor development without forcing a specific pathway (i.e., Kras). The primary focus of this chapter is to discuss a well-established 2-stage mouse model of lung adenocarcinomas. The initiator (3-methylcholanthrene, MCA) does not elicit many, if any, tumors if not followed by exposure to the tumor promoter (butylated hydroxytoluene, BHT). In sensitive strains, such as A/J, FVB, and BALB, significantly greater numbers of tumors develop following the MCA/BHT protocol compared to MCA alone. BHT does not elicit tumors on its own; it is a non-genotoxic carcinogen and promoter. In these sensitive strains, promotion is also associated with inflammation characterized by infiltrating macrophages, lymphocytes, and neutrophils, and other inflammatory cell types in addition to increases in total protein content reflective of lung hyperpermeability. This 2-stage model is a useful tool to identify unique promotion specific events to then test in future intervention studies.
Topics: Animals; Butylated Hydroxytoluene; Carcinogenesis; Lung; Methylcholanthrene; Mice; Mice, Inbred BALB C
PubMed: 33785163
DOI: 10.1016/bs.mcb.2020.07.003