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1.
Toxicol Appl Pharmacol ; 431: 115742, 2021 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-34624356

RESUMO

Benzene is a ubiquitous environmental pollutant. Recent population-based studies suggest that benzene exposure is associated with an increased risk for cardiovascular disease. However, it is unclear whether benzene exposure by itself is sufficient to induce cardiovascular toxicity. We examined the effects of benzene inhalation (50 ppm, 6 h/day, 5 days/week, 6 weeks) or HEPA-filtered air exposure on the biomarkers of cardiovascular toxicity in male C57BL/6J mice. Benzene inhalation significantly increased the biomarkers of endothelial activation and injury including endothelial microparticles, activated endothelial microparticles, endothelial progenitor cell microparticles, lung endothelial microparticles, and activated lung and endothelial microparticles while having no effect on circulating levels of endothelial adhesion molecules, endothelial selectins, and biomarkers of angiogenesis. To understand how benzene may induce endothelial injury, we exposed human aortic endothelial cells to benzene metabolites. Of the metabolites tested, trans,trans-mucondialdehyde (10 µM, 18h) was the most toxic. It induced caspases-3, -7 and -9 (intrinsic pathway) activation and enhanced microparticle formation by 2.4-fold. Levels of platelet-leukocyte aggregates, platelet macroparticles, and a proportion of CD4+ and CD8+ T-cells were also significantly elevated in the blood of the benzene-exposed mice. We also found that benzene exposure increased the transcription of genes associated with endothelial cell and platelet activation in the liver; and induced inflammatory genes and suppressed cytochrome P450s in the lungs and the liver. Together, these data suggest that benzene exposure induces endothelial injury, enhances platelet activation and inflammatory processes; and circulatory levels of endothelial cell and platelet-derived microparticles and platelet-leukocyte aggregates are excellent biomarkers of cardiovascular toxicity of benzene.


Assuntos
Benzeno/toxicidade , Doenças Cardiovasculares/induzido quimicamente , Sistema Cardiovascular/efeitos dos fármacos , Animais , Doenças Assintomáticas , Benzeno/administração & dosagem , Biomarcadores/sangue , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Plaquetas/patologia , Cardiotoxicidade , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/patologia , Sistema Cardiovascular/metabolismo , Sistema Cardiovascular/patologia , Micropartículas Derivadas de Células/efeitos dos fármacos , Micropartículas Derivadas de Células/metabolismo , Micropartículas Derivadas de Células/patologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/patologia , Exposição por Inalação , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Leucócitos/patologia , Masculino , Camundongos Endogâmicos C57BL
2.
Toxicol Sci ; 167(2): 426-437, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30346588

RESUMO

Benzene is a ubiquitous pollutant associated with hematotoxicity but its metabolic effects are unknown. We sought to determine if and how exposure to volatile benzene impacted glucose handling. We exposed wild type C57BL/6 mice to volatile benzene (50 ppm × 6 h/day) or HEPA-filtered air for 2 or 6 weeks and measured indices of oxidative stress, inflammation, and insulin signaling. Compared with air controls, we found that mice inhaling benzene demonstrated increased plasma glucose (p = .05), insulin (p = .03), and HOMA-IR (p = .05), establishing a state of insulin and glucose intolerance. Moreover, insulin-stimulated Akt phosphorylation was diminished in the liver (p = .001) and skeletal muscle (p = .001) of benzene-exposed mice, accompanied by increases in oxidative stress and Nf-κb phosphorylation (p = .025). Benzene-exposed mice also demonstrated elevated levels of Mip1-α transcripts and Socs1 (p = .001), but lower levels of Irs-2 tyrosine phosphorylation (p = .0001). Treatment with the superoxide dismutase mimetic, TEMPOL, reversed benzene-induced effects on oxidative stress, Nf-κb phosphorylation, Socs1 expression, Irs-2 tyrosine phosphorylation, and systemic glucose intolerance. These findings suggest that exposure to benzene induces insulin resistance and that this may be a sensitive indicator of inhaled benzene toxicity. Persistent ambient benzene exposure may be a heretofore unrecognized contributor to the global human epidemics of diabetes and cardiovascular disease.


Assuntos
Poluentes Atmosféricos/toxicidade , Benzeno/toxicidade , Exposição por Inalação/efeitos adversos , Resistência à Insulina , Estresse Oxidativo/efeitos dos fármacos , Animais , Contagem de Células Sanguíneas , Glicemia/análise , Insulina/sangue , Camundongos Endogâmicos C57BL
3.
JCI Insight ; 2(9)2017 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-28469073

RESUMO

Mechanisms of atherogenesis have been studied extensively in genetically engineered mice with disturbed cholesterol metabolism such as those lacking either the LDL receptor (Ldlr) or apolipoprotein E (apoe). Few other animal models of atherosclerosis are available. WT rabbits or rats, even on high-fat or high-cholesterol diets, develop sparse atherosclerotic lesions. We examined the effects of Ldlr deletion on lipoprotein metabolism and atherosclerotic lesion formation in Sprague-Dawley rats. Deletion of Ldlr resulted in the loss of the LDLR protein and caused a significant increase in plasma total cholesterol and triglycerides. On normal chow, Ldlr-KO rats gained more weight and were more glucose intolerant than WT rats. Plasma proprotein convertase subtilisin kexin 9 (PCSK9) and leptin levels were higher and adiponectin levels were lower in KO than WT rats. On the Western diet, the KO rats displayed exaggerated obesity and age-dependent increases in glucose intolerance. No appreciable aortic lesions were observed in KO rats fed normal chow for 64 weeks or Western diet for 16 weeks; however, after 34-52 weeks of Western diet, the KO rats developed exuberant atherosclerotic lesions in the aortic arch and throughout the abdominal aorta. The Ldlr-KO rat may be a useful model for studying obesity, insulin resistance, and early-stage atherosclerosis.

4.
J Biol Chem ; 287(14): 11398-409, 2012 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-22228760

RESUMO

Lipid peroxidation products, such as 4-hydroxy-trans-2-nonenal (HNE), cause endothelial activation, and they increase the adhesion of the endothelium to circulating leukocytes. Nevertheless, the mechanisms underlying these effects remain unclear. We observed that in HNE-treated human umbilical vein endothelial cells, some of the protein-HNE adducts colocalize with the endoplasmic reticulum (ER) and that HNE forms covalent adducts with several ER chaperones that assist in protein folding. We also found that at concentrations that did not induce apoptosis or necrosis, HNE activated the unfolded protein response, leading to an increase in XBP-1 splicing, phosphorylation of protein kinase-like ER kinase and eukaryotic translation initiation factor 2α, and the induction of ATF3 and ATF4. This increase in eukaryotic translation initiation factor 2α phosphorylation was prevented by transfection with protein kinase-like ER kinase siRNA. Treatment with HNE increased the expression of the ER chaperones, GRP78 and HERP. Exposure to HNE led to a depletion of reduced glutathione and an increase in the production of reactive oxygen species (ROS); however, glutathione depletion and ROS production by tert-butyl-hydroperoxide did not trigger the unfolded protein response. Pretreatment with a chemical chaperone, phenylbutyric acid, or adenoviral transfection with ATF6 attenuated HNE-induced monocyte adhesion and IL-8 induction. Moreover, phenylbutyric acid and taurine-conjugated ursodeoxycholic acid attenuated HNE-induced leukocyte rolling and their firm adhesion to the endothelium in rat cremaster muscle. These data suggest that endothelial activation by HNE is mediated in part by ER stress, induced by mechanisms independent of ROS production or glutathione depletion. The induction of ER stress may be a significant cause of vascular inflammation induced by products of oxidized lipids.


Assuntos
Aldeídos/metabolismo , Aldeídos/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Peroxidação de Lipídeos , Sequência de Aminoácidos , Animais , Chaperona BiP do Retículo Endoplasmático , Endotélio/citologia , Endotélio/efeitos dos fármacos , Endotélio/metabolismo , Glutationa/metabolismo , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Peroxidação de Lipídeos/efeitos dos fármacos , Dados de Sequência Molecular , Transporte Proteico/efeitos dos fármacos , Proteínas/química , Proteínas/metabolismo , Resposta a Proteínas não Dobradas/efeitos dos fármacos
5.
Mol Cell Endocrinol ; 323(2): 268-76, 2010 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-20302909

RESUMO

Resistance to endocrine therapy is a major clinical problem in breast cancer. The role of ERalpha splice variants in endocrine resistance is largely unknown. We observed reduced protein expression of an N-terminally truncated ERalpha46 in endocrine-resistant LCC2, LCC9, and LY2 compared to MCF-7 breast cancer cells. Transfection of LCC9 and LY2 cells with hERalpha46 partially restored growth inhibition by TAM. Overexpression of hERalpha46 in MCF-7 cells reduced estradiol (E(2))-stimulated endogenous pS2, cyclin D1, nuclear respiratory factor-1 (NRF-1), and progesterone receptor transcription. Expression of oncomiR miR-21 was lower in TAM-resistant LCC9 and LY2 cells compared to MCF-7 cells. Transfection with ERalpha46 altered the pharmacology of E(2) regulation of miR-21 expression from inhibition to stimulation, consistent with the hypothesis that hERalpha46 inhibits ERalpha activity. Established miR-21 targets PTEN and PDCD4 were reduced in ERalpha46-transfected, E(2)-treated MCF-7 cells. In conclusion, ERalpha46 appears to enhance endocrine responses by inhibiting selected ERalpha66 responses.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Proliferação de Células , Resistencia a Medicamentos Antineoplásicos/fisiologia , Receptor alfa de Estrogênio , Regulação Neoplásica da Expressão Gênica , Isoformas de Proteínas , Tamoxifeno/uso terapêutico , Animais , Linhagem Celular Tumoral , Ciclina D1/metabolismo , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Camundongos , Camundongos Nus , MicroRNAs/genética , MicroRNAs/metabolismo , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transcrição Gênica , Transplante Heterólogo
6.
Mol Cancer Ther ; 9(3): 594-605, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20197399

RESUMO

Anacardic acid (AnAc; 2-hydroxy-6-alkylbenzoic acid) is a dietary and medicinal phytochemical with established anticancer activity in cell and animal models. The mechanisms by which AnAc inhibits cancer cell proliferation remain undefined. AnAc 24:1(omega5) was purified from geranium (Pelargonium x hortorum) and shown to inhibit the proliferation of estrogen receptor alpha (ERalpha)-positive MCF-7 and endocrine-resistant LCC9 and LY2 breast cancer cells with greater efficacy than ERalpha-negative primary human breast epithelial cells, MCF-10A normal breast epithelial cells, and MDA-MB-231 basal-like breast cancer cells. AnAc 24:1(omega5) inhibited cell cycle progression and induced apoptosis in a cell-specific manner. AnAc 24:1(omega5) inhibited estradiol (E(2))-induced estrogen response element (ERE) reporter activity and transcription of the endogenous E(2) target genes pS2, cyclin D1, and cathepsin D in MCF-7 cells. AnAc 24:1(omega5) did not compete with E(2) for ERalpha or ERbeta binding, nor did AnAc 24:1(omega5) reduce ERalpha or ERbeta steady-state protein levels in MCF-7 cells; rather, AnAc 24:1(omega5) inhibited ER-ERE binding in vitro. Virtual screening with the molecular docking software Surflex evaluated AnAc 24:1(omega5) interaction with ERalpha ligand binding (LBD) and DNA binding (DBD) domains in conjunction with experimental validation. Molecular modeling revealed AnAc 24:1(omega5) interaction with the ERalpha DBD but not the LBD. Chromatin immunoprecipitation experiments revealed that AnAc 24:1(omega5) inhibited E(2)-ERalpha interaction with the endogenous pS2 gene promoter region containing an ERE. These data indicate that AnAc 24:1(omega5) inhibits cell proliferation, cell cycle progression, and apoptosis in an ER-dependent manner by reducing ER-DNA interaction and inhibiting ER-mediated transcriptional responses.


Assuntos
Ácidos Anacárdicos/farmacologia , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , DNA/metabolismo , Receptor alfa de Estrogênio/antagonistas & inibidores , Transcrição Gênica/efeitos dos fármacos , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , DNA/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genes Reporter/efeitos dos fármacos , Humanos , Ligação Proteica/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacos , Transfecção , Células Tumorais Cultivadas
7.
Nucleic Acids Res ; 37(8): 2584-95, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19264808

RESUMO

Select changes in microRNA (miRNA) expression correlate with estrogen receptor alpha (ER alpha) expression in breast tumors. miR-21 is higher in ER alpha positive than negative tumors, but no one has examined how estradiol (E(2)) regulates miR-21 in breast cancer cells. Here we report that E(2) inhibits miR-21 expression in MCF-7 human breast cancer cells. The E(2)-induced reduction in miR-21 was inhibited by 4-hydroxytamoxifen (4-OHT), ICI 182 780 (Faslodex), and siRNA ER alpha indicating that the suppression is ER alpha-mediated. ER alpha and ER beta agonists PPT and DPN inhibited and 4-OHT increased miR-21 expression. E(2) increased luciferase activity from reporters containing the miR-21 recognition elements from the 3'-UTRs of miR-21 target genes, corroborating that E(2) represses miR-21 expression resulting in a loss of target gene suppression. The E(2)-mediated decrease in miR-21 correlated with increased protein expression of endogenous miR-21-targets Pdcd4, PTEN and Bcl-2. siRNA knockdown of ER alpha blocked the E(2)-induced increase in Pdcd4, PTEN and Bcl-2. Transfection of MCF-7 cells with antisense (AS) to miR-21 mimicked the E(2)-induced increase in Pdcd4, PTEN and Bcl-2. These results are the first to demonstrate that E(2) represses the expression of an oncogenic miRNA, miR-21, by activating estrogen receptor in MCF-7 cells.


Assuntos
Neoplasias da Mama/genética , Estradiol/farmacologia , Regulação Neoplásica da Expressão Gênica , MicroRNAs/metabolismo , Regiões 3' não Traduzidas/química , Proteínas Reguladoras de Apoptose/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Cicloeximida/farmacologia , Dactinomicina/farmacologia , Regulação para Baixo , Estradiol/análogos & derivados , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/agonistas , Receptor beta de Estrogênio/antagonistas & inibidores , Receptor beta de Estrogênio/genética , Feminino , Fulvestranto , Genes Reporter , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Inibidores da Síntese de Ácido Nucleico/farmacologia , Regiões Promotoras Genéticas , Inibidores da Síntese de Proteínas/farmacologia , RNA Antissenso/metabolismo , Proteínas de Ligação a RNA/genética , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia , Proteínas Ativadoras de ras GTPase/genética
8.
FASEB J ; 22(7): 2185-97, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18296501

RESUMO

Epidemiological studies correlate moderate red wine consumption to reduced incidence of cardiovascular disease. Resveratrol is a polyphenolic compound in red wine that has cardioprotective effects in rodents. Although endothelial cell (EC) studies indicate that micromolar resveratrol has diverse biological activities, these concentrations are not physiologically relevant because human oral ingestion provides only brief exposure to nanomolar plasma levels. Previously, we reported that nanomolar resveratrol activated ERK1/2 signaling in bovine aortic ECs (BAECs). The goal of this study was to determine the mechanisms by which nanomolar resveratrol rapidly activates endothelial nitric oxide synthase (eNOS) in human umbilical vein ECs (HUVECs). We report for the first time that resveratrol increased interaction between estrogen receptor alpha (ER alpha), caveolin-1 (Cav-1) and c-Src, and increased phosphorylation of Cav-1, c-Src, and eNOS. Pretreatment with the lipid raft disruptor beta-methyl cyclodextrin or G alpha inhibitor pertussis toxin blocked resveratrol- and E(2)-induced eNOS activation and NO production. Depletion of endogenous ER alpha, not ERbeta, by siRNA attenuated resveratrol- and E(2)-induced ERK1/2, Src, and eNOS phosphorylation. Our data demonstrate that nanomolar resveratrol induces ER alpha-Cav-1-c-SRC interaction, resulting in NO production through a G alpha-protein-coupled mechanism. This study provides important new insights into mechanisms for the beneficial effects of resveratrol in ECs.


Assuntos
Inibidores da Angiogênese/farmacologia , Caveolina 1/fisiologia , Endotélio Vascular/fisiologia , Receptor alfa de Estrogênio/fisiologia , Estilbenos/farmacologia , Veias Umbilicais/fisiologia , Quinases da Família src/fisiologia , Caveolina 1/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Estradiol/farmacologia , Receptor alfa de Estrogênio/efeitos dos fármacos , Etanol/farmacologia , Humanos , Cinética , Fosforilação , Resveratrol , Veias Umbilicais/efeitos dos fármacos , Quinases da Família src/efeitos dos fármacos
9.
Mol Endocrinol ; 22(3): 609-22, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18048642

RESUMO

Estrogen has direct and indirect effects on mitochondrial activity, but the mechanisms mediating these effects remain unclear. Others reported that long-term estradiol (E(2)) treatment increased nuclear respiratory factor-1 (NRF-1) protein in cerebral blood vessels of ovariectomized rats. NRF-1 is a transcription factor that regulates the expression of nuclear-encoded mitochondrial genes, e.g. mitochondrial transcription factor A (TFAM), that control transcription of the mitochondrial genome. Here we tested the hypothesis that E(2) increases NRF-1 transcription resulting in a coordinate increase in the expression of nuclear- and mitochondrial- encoded genes and mitochondrial respiratory activity. We show that E(2) increased NRF-1 mRNA and protein in MCF-7 breast and H1793 lung adenocarcinoma cells in a time-dependent manner. E(2)-induced NRF-1 expression was inhibited by the estrogen receptor (ER) antagonist ICI 182,780 and actinomycin D but not by phosphoinositide-3 kinase and MAPK inhibitors, indicating a genomic mechanism of E(2) regulation of NRF-1 transcription. An estrogen response element (ERE) in the NRF-1 promoter bound ER alpha and ER beta in vitro, and E(2) induced ER alpha and ER beta recruitment to this ERE in chromatin immunoprecipitation assays in MCF-7 cells. The NRF-1 ERE activated reporter gene expression in transfected cells. Small interfering RNA to ER alpha and ER beta revealed that ER alpha mediates E(2)-induced NRF-1 transcription. The E(2)-induced increase in NRF-1 was followed by increased TFAM and the transcription of Tfam-regulated mitochondrial DNA-encoded COI and NDI genes and increased mitochondrial biogenesis. Knockdown of NRF-1 blocked E(2) stimulation of mitochondrial biogenesis and activity, indicating a mechanism by which estrogens regulate mitochondrial function by increasing NRF-1 expression.


Assuntos
Estradiol/farmacologia , Mitocôndrias/efeitos dos fármacos , Fator 1 Nuclear Respiratório/biossíntese , Transcrição Gênica/fisiologia , Western Blotting , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , DNA Mitocondrial/genética , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/genética , Estradiol/análogos & derivados , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/agonistas , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Fulvestranto , Humanos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Proteínas Mitocondriais/biossíntese , Proteínas Mitocondriais/genética , Nitrilas/farmacologia , Fator 1 Nuclear Respiratório/genética , Fenóis , Regiões Promotoras Genéticas/efeitos dos fármacos , Propionatos/farmacologia , Pirazóis/farmacologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética
10.
Cancer Res ; 66(20): 10188-98, 2006 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-17047084

RESUMO

Tamoxifen (TAM) is successfully used for the treatment and prevention of breast cancer. However, many patients that are initially TAM responsive develop tumors that are antiestrogen/TAM resistant (TAM-R). The mechanism behind TAM resistance in estrogen receptor alpha (ERalpha)-positive tumors is not understood. The orphan nuclear receptor chicken ovalbumin upstream promoter transcription factor (COUP-TF)-I interacts directly with 4-hydroxytamoxifen (4-OHT)- and estradiol (E(2))-occupied ERalpha, corepressors NCoR and SMRT, and inhibit E(2)-induced gene transcription in breast cancer cells. Here we tested the hypothesis that reduced COUP-TFI and COUP-TFII correlate with TAM resistance. We report for the first time that COUP-TFII, but not COUP-TFI, is reduced in three antiestrogen/TAM-R cell lines derived from TAM-sensitive (TAM-S) MCF-7 human breast cancer cells and in MDA-MB-231 cells compared with MCF-7. ERalpha and ERbeta protein expression was not different between TAM-S and TAM-R cells, but progesterone receptor (PR) was decreased in TAM-R cells. Further, E(2) increased COUP-TFII transcription in MCF-7, but not TAM-R, cells. Importantly, reexpression of COUP-TFII in TAM-S cells to levels comparable to those in MCF-7 was shown to increase 4-OHT-mediated growth inhibition and increased apoptosis. Conversely, knockdown of COUP-TFII in TAM-S MCF-7 cells blocked growth inhibitory activity and increased 4-OHT agonist activity. 4-OHT increased COUP-TFII-ERalpha interaction approximately 2-fold in MCF-7 cells. COUP-TFII expression in TAM-R cells also inhibited 4-OHT-induced endogenous PR and pS2 mRNA expression. These data indicate that reduced COUP-TFII expression correlates with acquired TAM resistance in human breast cancer cell lines and that COUP-TFII plays a role in regulating the growth inhibitory activity of TAM in breast cancer cells.


Assuntos
Antineoplásicos Hormonais/farmacologia , Neoplasias da Mama/metabolismo , Fator II de Transcrição COUP/biossíntese , Tamoxifeno/farmacologia , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Fator II de Transcrição COUP/genética , Fator II de Transcrição COUP/metabolismo , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/fisiologia , Linhagem Celular Tumoral , Galinhas , Resistencia a Medicamentos Antineoplásicos , Estradiol/análogos & derivados , Estradiol/farmacologia , Receptor alfa de Estrogênio/biossíntese , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/biossíntese , Fulvestranto , Humanos , Receptores de Progesterona/biossíntese , Receptores de Progesterona/genética , Tamoxifeno/análogos & derivados , Transcrição Gênica , Transfecção
11.
Anticancer Res ; 25(4): 2841-9, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16080536

RESUMO

BACKGROUND: The tumor microenvironment is believed to contribute to the malignant properties of tumor cells in heterogeneous tumor tissues. We investigated the impact of hypoxia (1% oxygen) on the expression of cathepsin B and its natural inhibitors cystatin B and C. MATERIALS AND METHODS: Patient-matched oral carcinoma cell lines from primary tumor and lymph node metastasis were used to study the effects of hypoxia on proliferation, protein expression, and proteolytic and inhibitor activities. RESULTS: Hypoxic growth led to elevated cathepsin B expression and activity, and this effect was greater in metastatic than in primary tumor cells. Also, hypoxia led to down-regulation of the inhibitors cystatin C and B, resulting in increased residual activity of cathepsin B. CONCLUSION: These data suggest that the invasive and/or metastatic potential of cells may be enhanced under hypoxia by increasing cathepsin-mediated proteolysis. The results provide strong evidence for the involvement of cathepsin B and its cystatin inhibitors in hypoxia-enhanced tumor progression.


Assuntos
Carcinoma de Células Escamosas/enzimologia , Catepsina B/metabolismo , Cistatinas/metabolismo , Inibidores de Cisteína Proteinase/metabolismo , Neoplasias Bucais/enzimologia , Idoso , Carcinoma de Células Escamosas/patologia , Catepsina B/antagonistas & inibidores , Processos de Crescimento Celular/fisiologia , Hipóxia Celular/fisiologia , Linhagem Celular Tumoral , Cistatina B , Cistatina C , Humanos , Neoplasias Hipofaríngeas/enzimologia , Neoplasias Hipofaríngeas/patologia , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/patologia
12.
Arch Biochem Biophys ; 436(1): 187-95, 2005 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-15752724

RESUMO

We previously demonstrated that overexpression of cathepsin B (CB) protease in oral squamous cell carcinomas correlated positively with advanced tumor stage and poor histologic malignancy grade. Here we examined whether CB contributes to the invasiveness of oral carcinoma cells. For RNA-mediated inhibition, two ribozymes that target CB mRNA were designed and stably expressed in the oral squamous cell carcinoma cell line 1386Tu. Both ribozymes diminished expression of CB mRNA, protein, and activity, without affecting cathepsin D or beta-actin, as determined by quantitative real-time PCR, Western blots, and protease activity assays. Matrigel invasion assays showed that the invasiveness of the cells was significantly reduced by the expressed ribozymes and, surprisingly, the motilities of the ribozyme-transfected cells were also diminished. Our results document a direct role for CB in promoting oral cancer spread and invasion, and open the possibility of controlling oral carcinoma malignancy and metastasis by targeting CB with RNA inhibitor strategies.


Assuntos
Carcinoma de Células Escamosas/patologia , Catepsina B/fisiologia , Movimento Celular/fisiologia , Neoplasias Bucais/patologia , Sequência de Bases , Biomarcadores , Western Blotting , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Dados de Sequência Molecular , Invasividade Neoplásica , Estadiamento de Neoplasias , Peptídeo Hidrolases/metabolismo , Prognóstico , Antígeno Nuclear de Célula em Proliferação/análise , RNA Catalítico/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transfecção , Células Tumorais Cultivadas
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