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1.
J Nucl Cardiol ; 26(1): 275-283, 2019 02.
Article in English | MEDLINE | ID: mdl-28357812

ABSTRACT

BACKGROUND: We evaluated the performance of stress imaging with technetium-99m-labeled tetrofosmin single-photon emission computed tomography (SPECT) and rubidium-82 positron emission tomography (PET) in patients with extreme obesity, defined as body mass index ≥40 kg/m2. METHODS: We identified patients with extreme obesity who underwent angiography in our center and either stress SPECT or PET within the previous six months. Cohorts of patients with extreme obesity and a <5% pretest likelihood of CAD who underwent SPECT (N = 25) or PET (N = 25) were also included. RESULTS: In total, 108 patients who underwent SPECT (N = 57) or PET (N = 51) were identified. Scan interpretation was classified as definitely normal or abnormal in 83.3% of PET and 60.5% of SPECT scans, respectively (P < .01). PET demonstrated higher diagnostic accuracy and normalcy rate. PET was found to have higher specificity for the pooled cohort. Similar findings were observed using stenosis cut-offs of ≥50% and ≥70%. CONCLUSIONS: In patients with extreme obesity, PET enabled more definitive scan interpretation with less artifact compared to SPECT. PET provided higher diagnostic accuracy and specificity in the detection of obstructive coronary artery disease.


Subject(s)
Coronary Artery Disease/diagnostic imaging , Obesity, Morbid/diagnostic imaging , Positron-Emission Tomography , Tomography, Emission-Computed, Single-Photon , Aged , Coronary Angiography , Female , Humans , Image Processing, Computer-Assisted , Male , Middle Aged , Myocardial Perfusion Imaging , Prospective Studies , Registries , Rubidium Radioisotopes , Sensitivity and Specificity , Technetium
2.
Stem Cells Int ; 2016: 4536187, 2016.
Article in English | MEDLINE | ID: mdl-27366154

ABSTRACT

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor belonging to the Per-Arnt-Sim (PAS) family of proteins. The AHR is involved in hematopoietic stem cell (HSC) functions including self-renewal, proliferation, quiescence, and differentiation. We hypothesize that AHR impacts HSC functions by influencing genes that have roles in HSC maintenance and function and that this may occur through regulation of bone marrow (BM) niche cells. We examined BM and niche cells harvested from 8-week-old AHR null-allele (KO) mice in which exon 3 was deleted in the Ahr gene and compared these data to cells from B6 control mice; young and old (10 months) animals were also compared. We report changes in HSCs and peripheral blood cells in mice lacking AHR. Serial transplantation assays revealed a significant increase in long term HSCs. There was a significant increase in mesenchymal stem cells constituting the endosteal BM niche. Gene expression analyses of HSCs revealed an increase in expression of genes involved in proliferation and maintenance of quiescence. Our studies infer that loss of AHR results in increased proliferation and self-renewal of long term HSCs, in part, by influencing the microenvironment in the niche regulating the balance between quiescence and proliferation in HSCs.

3.
Cancer Prev Res (Phila) ; 8(3): 249-57, 2015 Mar.
Article in English | MEDLINE | ID: mdl-25604133

ABSTRACT

(-)-Epigallocatechin gallate (EGCG), a major tea polyphenol, elicits anticancer effects. However, the mechanism of action is not fully understood. Our laboratory previously showed that EGCG inhibits heat shock protein 90 (HSP90). We used nontumorigenic (NT), tumorigenic, and metastatic cancer cells from a novel human prostate cancer progression model to test the hypotheses that certain stages are more or less sensitive to EGCG and that sensitivity is related to HSP90 inhibition. Treatment of cells with EGCG, novobiocin, or 17-AAG resulted in more potent cytotoxic effects on tumorigenic and metastatic cells than NT cells. When tumorigenic or metastatic cells were grown in vivo, mice supplemented with 0.06% EGCG in drinking water developed significantly smaller tumors than untreated mice. Furthermore, EGCG prevented malignant transformation in vivo using the full prostate cancer model. To elucidate the mechanism of EGCG action, we performed binding assays with EGCG-Sepharose, a C-terminal HSP90 antibody, and HSP90 mutants. These experiments revealed that EGCG-Sepharose bound more HSP90 from metastatic cells compared with NT cells and binding occurred through the HSP90 C-terminus. In addition, EGCG bound HSP90 mutants that mimic both complexed and uncomplexed HSP90. Consistent with HSP90 inhibitory activity, EGCG, novobiocin, and 17-AAG induced changes in HSP90-client proteins in NT cells and larger differences in metastatic cells. These data suggest that EGCG may be efficacious for the treatment of prostate cancer because it preferentially targets cancer cells and inhibits a molecular chaperone supportive of the malignant phenotype.


Subject(s)
Antineoplastic Agents/pharmacology , Catechin/analogs & derivatives , Cell Movement/drug effects , Cell Proliferation/drug effects , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Prostatic Hyperplasia/drug therapy , Prostatic Neoplasms/drug therapy , Animals , Apoptosis/drug effects , Blotting, Western , Catechin/pharmacology , Disease Progression , HSP90 Heat-Shock Proteins/genetics , HSP90 Heat-Shock Proteins/metabolism , Humans , Male , Mice , Prostatic Hyperplasia/metabolism , Prostatic Hyperplasia/pathology , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/secondary , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Tumor Cells, Cultured , Wound Healing/drug effects , Xenograft Model Antitumor Assays
4.
Bioorg Med Chem Lett ; 24(10): 2263-6, 2014 May 15.
Article in English | MEDLINE | ID: mdl-24745965

ABSTRACT

(-)-Epigallocatechin gallate (EGCG) is the major flavonoid of green tea and has been widely explored for a range of biological activities including anti-infective, anti-inflammatory, anti-cancer, and neuroprotection. Existing structure-activity data for EGCG has been largely limited to exploration of simple ethers and hydroxyl deletion. EGCG has poor drug-like properties because of multiple phenolic hydroxyl moieties and a metabolically labile ester. This work reports a substantial expansion of structure-activity understanding by exploring a range of semi-synthetic and synthetic derivatives with ester replacements and variously substituted aromatic and alicyclic groups containing more drug-like substituents. Structure-activity relationships for these molecules were obtained for Hsp90 inhibition. The results indicate that amide and sulfonamide linkers are suitable ester replacements. Hydroxylated aromatic rings and the cis-stereochemistry in EGCG are not essential for Hsp90 inhibition. Selected analogs in this series are more potent than EGCG in a luciferase refolding assay for Hsp90 activity.


Subject(s)
Catechin/analogs & derivatives , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Biological Products/chemistry , Biological Products/pharmacology , Catechin/chemistry , Catechin/pharmacology , Drug Discovery , Structure-Activity Relationship
5.
Stem Cells Dev ; 20(2): 313-26, 2011 Feb.
Article in English | MEDLINE | ID: mdl-20486776

ABSTRACT

Neurogenesis involves the proliferation of multipotent neuroepithelial stem cells followed by differentiation into lineage-restricted neural precursor cells (NPCs) during the embryonic period. Interestingly, these progenitor cells express robust levels of the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor that regulates expression of genes important for growth regulation, and xenobiotic metabolism. Upon binding 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a pervasive environmental contaminant and potent AhR ligand, AhR, is activated and disrupts gene expression patterns to produce cellular toxicity. Because of its widespread distribution in the brain during critical proliferative phases of neurogenesis, it is conceivable that AhR participates in NPC expansion. Therefore, this study tested the hypothesis that AhR activation by TCDD disrupts signaling events that regulate NPC proliferation. The C17.2 NPC line served as a model system to (1) assess whether NPCs are targets for TCDD-induced neurotoxicity and (2) characterize the effects of TCDD on NPC proliferation. We demonstrated that C17.2 NPCs express an intact AhR signaling pathway that becomes transcriptionally active after TCDD exposure. (3)H-thymidine and alamar blue reduction assays indicated that TCDD suppresses NPC proliferation in a concentration-dependent manner without the loss of cell viability. Cell cycle distribution analysis by flow cytometry revealed that TCDD-induced growth arrest results from an impaired G1 to S cell cycle transition. Moreover, TCDD exposure altered p27( kip1) and cyclin D1 cell cycle regulatory protein expression levels consistent with a G1 phase arrest. Initial studies in primary NPCs isolated from the ventral forebrain of embryonic mice demonstrated that TCDD reduced cell proliferation through a G1 phase arrest, corroborating our findings in the C17.2 cell line. Together, these observations suggest that the inappropriate or sustained activation of AhR by TCDD during neurogenesis can interfere with signaling pathways that regulate neuroepithelial stem cell/NPC proliferation, which could adversely impact final cell number in the brain and lead to functional impairments.


Subject(s)
Cell Proliferation/drug effects , Neural Stem Cells/drug effects , Neurons/drug effects , Polychlorinated Dibenzodioxins/pharmacology , Receptors, Aryl Hydrocarbon/agonists , Animals , Cell Cycle/drug effects , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Cell Survival/drug effects , Cells, Cultured , DNA Replication/drug effects , Embryo, Mammalian , Female , Gene Expression Regulation, Developmental/drug effects , Male , Mice , Mice, Inbred C57BL , Mitogens/pharmacology , Neural Stem Cells/cytology , Neurogenesis , Neurons/cytology , Pregnancy , Prosencephalon/cytology , Receptors, Aryl Hydrocarbon/metabolism , Tumor Cells, Cultured
6.
Toxicol Sci ; 114(1): 90-100, 2010 Mar.
Article in English | MEDLINE | ID: mdl-19933214

ABSTRACT

The aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor, mediates toxicity of several classes of xenobiotics and also has important physiological roles in differentiation, reproduction, and immunity, although the endogenous ligand(s) mediating these functions is/are as yet unidentified. One candidate endogenous ligand, 2-(1'H-indolo-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE), is a potent AhR agonist in vitro, activates the murine AhR in vivo, but does not induce toxicity. We hypothesized that ITE and the toxic ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), may modify transcription of different sets of genes to account for their different toxicity. To test this hypothesis, primary mouse lung fibroblasts were exposed to 0.5muM ITE, 0.2nM TCDD, or vehicle for 4 h, and total gene expression was evaluated using microarrays. After this short-term and low-dose treatment, several hundred genes were changed significantly, and the response to ITE and TCDD was remarkably similar, both qualitatively and quantitatively. Induced gene sets included the expected battery of AhR-dependent xenobiotic-metabolizing enzymes, as well as several sets that reflect the inflammatory role of lung fibroblasts. Real time quantitative RT-qPCR assay of several selected genes confirmed these microarray data and further suggested that there may be kinetic differences in expression between ligands. These data suggest that ITE and TCDD elicit an analogous change in AhR conformation such that the initial transcription response is the same. Furthermore, if the difference in toxicity between TCDD and ITE is mediated by differences in gene expression, then it is likely that secondary changes enabled by the persistent TCDD, but not by the shorter lived ITE, are responsible.


Subject(s)
Fibroblasts/drug effects , Gene Expression/drug effects , Indoles/toxicity , Lung/drug effects , Polychlorinated Dibenzodioxins/toxicity , Receptors, Aryl Hydrocarbon/metabolism , Thiazoles/toxicity , Animals , Dose-Response Relationship, Drug , Fibroblasts/metabolism , Ligands , Lung/cytology , Lung/metabolism , Mice , Mice, Inbred C57BL , Receptors, Aryl Hydrocarbon/genetics
7.
Biochemistry ; 48(2): 336-45, 2009 Jan 20.
Article in English | MEDLINE | ID: mdl-19113837

ABSTRACT

(-)-Epigallocatechin-3-gallate (EGCG), a major component of green tea, protects against certain types of cancers, although the mechanism has not yet been determined. It was previously demonstrated that EGCG blocks aryl hydrocarbon receptor (AhR)-mediated transcription induced by the potent carcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Unlike other AhR antagonists that directly bind to the AhR, EGCG inhibits AhR-mediated transcription by binding to hsp90. We hypothesize that EGCG exerts anti-AhR and anticancer effects by acting as an hsp90 inhibitor. Using proteolytic footprinting, immunoprecipitation, and an ATP-agarose pull-down assay, EGCG was found to directly modulate the conformation of hsp90 and bind at or near to a C-terminal ATP binding site. Hsp90 chaperone function, as assessed by its ability to mediate refolding of denatured luciferase, was inhibited by EGCG treatment. Hsp90 dimerization, which occurs at the C-terminal end, was also inhibited by EGCG treatment. Coimmunoprecipitation studies showed that EGCG stabilizes an AhR complex that includes hsp90 and XAP2 (hepatitis B virus X-associated protein 2), and decreases the association of aryl hydrocarbon nuclear translocator (Arnt) with ligand-activated AhR. Thus, EGCG, through its ability to bind to hsp90, blocks AhR response element (AhRE) recognition. These studies indicate a novel mechanism whereby EGCG inhibits ligand-induced AhRE binding and AhR-mediated transcriptional activity. In EGCG-treated human ovarian carcinoma SKOV3 cells, decreased levels of several cancer-related hsp90 client proteins, such as ErbB2, Raf-1 and phospho-AKT, were observed. EGCG also modified the association of hsp90 with several cochaperones. Overall, these data indicate that EGCG is a novel hsp90 inhibitor. Further studies are needed to determine if this has a role in the antitumor actions of EGCG.


Subject(s)
Catechin/analogs & derivatives , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Animals , Binding Sites/genetics , Catechin/metabolism , Catechin/pharmacology , Cell Line, Tumor , Chickens , Dimerization , Dose-Response Relationship, Drug , Glutathione Transferase/metabolism , HSP90 Heat-Shock Proteins/genetics , HSP90 Heat-Shock Proteins/isolation & purification , Humans , Ligands , Mice , Models, Biological , Molecular Chaperones/antagonists & inhibitors , Molecular Chaperones/genetics , Peptide Mapping , Plasmids , Protein Binding/genetics , Protein Conformation/drug effects , Receptors, Aryl Hydrocarbon/metabolism , Recombinant Fusion Proteins/metabolism , Response Elements/drug effects , Tea/genetics , Time Factors , Transcription, Genetic/drug effects
8.
Crit Rev Eukaryot Gene Expr ; 18(4): 279-321, 2008.
Article in English | MEDLINE | ID: mdl-18652561

ABSTRACT

Although the aryl hydrocarbon receptor (AhR) has been known as the mediator of the toxicity of particular xenobiotics such as the dioxins, the normal role of this transcription factor in a number of biological processes is just beginning to be recognized. Knowledge of AhR-targeted genes and signaling pathways indicates involvement of AhR in fundamental cell-regulatory pathways. Noted defects in the morphology and functions of certain tissues in the absence of AhR point to critical roles for this protein in developmental processes. Together, the data suggest that the AhR has an important function in controlling the balance among processes involved in cell proliferation, death, and differentiation rather than being essential for them. On the other hand, deregulation of these processes is known to contribute to events such as tumor initiation, promotion, and progression that ultimately lead to malignant tumor formation. Epidemiological and experimental animal data, along with a more detailed understanding of how AhR is involved in regulating particular signaling pathways, provide substantial support for an association between abnormal AhR function and cancer. Here we describe the current understanding of how the AhR may function to regulate both normal and cancerous tissue growth and development.


Subject(s)
Morphogenesis/physiology , Neoplasms/etiology , Receptors, Aryl Hydrocarbon/genetics , Receptors, Aryl Hydrocarbon/physiology , Animals , Cells, Cultured , Evolution, Molecular , Gene Expression Regulation, Developmental , Gene Expression Regulation, Neoplastic , Humans , Ligands , Mice , Mice, Knockout , Models, Biological , Morphogenesis/genetics , Neoplasms/genetics , Neoplasms/physiopathology , Polychlorinated Dibenzodioxins/toxicity , Receptor Cross-Talk , Receptors, Aryl Hydrocarbon/deficiency , Signal Transduction
9.
Mol Pharmacol ; 69(6): 1871-8, 2006 Jun.
Article in English | MEDLINE | ID: mdl-16540597

ABSTRACT

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a widespread environmental pollutant with many toxic effects, including endocrine disruption, reproductive dysfunction, immunotoxicity, liver damage, and cancer. These are mediated by TCDD binding to and activating the aryl hydrocarbon receptor (AhR), a basic helix-loop-helix transcription factor. In this regard, targeting the AhR using novel small molecule inhibitors is an attractive strategy for the development of potential preventive agents. In this study, by screening a chemical library composed of approximately 10,000 compounds, we identified a novel compound, 2-methyl-2H-pyrazole-3-carboxylic acid (2-methyl-4-o-tolylazo-phenyl)-amide (CH-223191), that potently inhibits TCDD-induced AhR-dependent transcription. In addition, CH-223191 blocked the binding of TCDD to AhR and inhibited TCDD-mediated nuclear translocation and DNA binding of AhR. These inhibitory effects of CH-223191 prevented the expression of cytochrome P450 enzymes, target genes of the AhR. Unlike many known antagonists of AhR, CH-223191 did not have detectable AhR agonist-like activity or estrogenic potency, suggesting that CH-223191 is a specific antagonist of AhR. It is noteworthy that CH-223191 potently prevented TCDD-elicited cytochrome P450 induction, liver toxicity, and wasting syndrome in mice. Taken together, these results demonstrate that this novel compound, CH-223191, may be a useful agent for the study of AhR-mediated signal transduction and the prevention of TCDD-associated pathology.


Subject(s)
Antidotes/pharmacology , Azo Compounds/pharmacology , Cytochrome P-450 CYP1A1/antagonists & inhibitors , Dioxins/antagonists & inhibitors , Pyrazoles/pharmacology , Receptors, Aryl Hydrocarbon/antagonists & inhibitors , Animals , Antidotes/chemistry , Azo Compounds/chemistry , Cell Line, Tumor , Cytochrome P-450 CYP1A1/drug effects , Dioxins/metabolism , Dioxins/toxicity , Drug Evaluation, Preclinical , Humans , Liver/drug effects , Liver/pathology , Male , Mice , Mice, Inbred ICR , Protein Transport/drug effects , Pyrazoles/chemistry , Receptors, Aryl Hydrocarbon/metabolism
10.
Mol Pharmacol ; 63(4): 915-24, 2003 Apr.
Article in English | MEDLINE | ID: mdl-12644593

ABSTRACT

To investigate possible species-specificity of aryl hydrocarbon receptor (AhR)-mediated signal transduction pathways, activities of 2,3,7,8-tetrochlorodibenzo-p-dioxin (TCDD) and six synthetic flavonoids were evaluated in mouse hepatoma and guinea pig adenocarcinoma cells transfected with an AhR-responsive luciferase reporter. Rank order potency in these two cell lines was similar for the ability of these flavonoids to antagonize TCDD-induced reporter gene expression. However, in the presence of flavone alone, a species-specific difference in agonist activity was observed. In guinea pig cells, several flavonoids demonstrated agonist activity up to 50% of the maximum TCDD response. In mouse cells, however, no significant agonist activity was observed at the same concentrations based on luciferase enzyme activity, protein expression, and mRNA analysis. Moreover, competitive ligand-binding assays, using [(3)H]TCDD in cytosolic fractions, demonstrated that 3'-methoxy-4'-nitroflavone had a similar IC(50) in both recombinant cell lines, suggesting that the flavone has similar binding affinity to receptors from both species. However, electrophoretic mobility shift assay using the cytosolic fractions demonstrated that this flavone elicited binding to the DRE by guinea pig but not mouse AhR complex. The dependence of the AhR in this differential interaction was further demonstrated using in vitro synthesized guinea pig and mouse Ah receptors and mouse Arnt. Together, these data suggest that the differential agonist/antagonist activity of these flavone derivatives is caused by the efficacy of these flavonoids in eliciting an AhR conformation that recognizes regulatory response elements in a species-specific manner.


Subject(s)
Flavonoids/pharmacology , Polychlorinated Dibenzodioxins/pharmacology , Receptors, Aryl Hydrocarbon/metabolism , Transcription, Genetic/drug effects , Animals , Binding, Competitive , Dose-Response Relationship, Drug , Flavonoids/chemical synthesis , Flavonoids/chemistry , Genes, Reporter , Guinea Pigs , Luciferases/metabolism , Mice , Receptors, Aryl Hydrocarbon/genetics , Species Specificity , Teratogens/pharmacology , Tumor Cells, Cultured
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