RESUMO
Millions of tons of all kind of munitions, including mines, bombs and torpedoes have been dumped after World War II in the marine environment and do now pose a new threat to the seas worldwide. Beside the acute risk of unwanted detonation, there is a chronic risk of contamination, because the metal vessels corrode and the toxic and carcinogenic explosives (trinitrotoluene (TNT) and metabolites) leak into the environment. While the mechanism of toxicity and carcinogenicity of TNT and its derivatives occurs through its capability of inducing oxidative stress in the target biota, we had the idea if TNT can induce the gene expression of carbonyl reductase in blue mussels. Carbonyl reductases are members of the short-chain dehydrogenase/reductase (SDR) superfamily. They metabolize xenobiotics bearing carbonyl functions, but also endogenous signal molecules such as steroid hormones, prostaglandins, biogenic amines, as well as sugar and lipid peroxidation derived reactive carbonyls, the latter providing a defence mechanism against oxidative stress and reactive oxygen species (ROS). Here, we identified and cloned the gene coding for carbonyl reductase from the blue mussel Mytilus spp. by a bioinformatics approach. In both laboratory and field studies, we could show that TNT induces a strong and concentration-dependent induction of gene expression of carbonyl reductase in the blue mussel. Carbonyl reductase may thus serve as a biomarker for TNT exposure on a molecular level which is useful to detect TNT contaminations in the environment and to perform a risk assessment both for the ecosphere and the human seafood consumer.
Assuntos
Oxirredutases do Álcool/biossíntese , Bombas (Dispositivos Explosivos) , Monitoramento Ambiental , Substâncias Explosivas/toxicidade , Resíduos Perigosos , Mytilus edulis/efeitos dos fármacos , Trinitrotolueno/toxicidade , Poluentes Químicos da Água/toxicidade , Oxirredutases do Álcool/genética , Animais , Biologia Computacional , Relação Dose-Resposta a Droga , Biomarcadores Ambientais/genética , Indução Enzimática , Mytilus edulis/enzimologia , Mytilus edulis/genética , Oceanos e Mares , Medição de Risco , II Guerra MundialRESUMO
Aberrantly high expression of EVI1 in acute myeloid leukaemia (AML) is associated with poor prognosis. For targeted treatment of EVI1 overexpressing AML a more detailed understanding of aspects of spatiotemporal interaction dynamics of the EVI1 protein is important. EVI1 overexpressing SB1690CB AML cells were used for quantification and protein interaction studies of EVI1 and ΔEVI1. Cells were cell cycle-synchronised by mimosine and nocodazole treatment and expression of EVI1 and related proteins assessed by western blot, immunoprecipitation and immunofluorescence. EVI1 protein levels oscillate through the cell cycle, and EVI1 is degraded partly by the proteasome complex. Both EVI1 and ΔEVI1 interact with the co-repressor CtBP1 but dissociate from CtBP1 complexes during mitosis. Furthermore, a large fraction of EVI1, but not ΔEVI1 or CtBP1, resides in the nuclear matrix. In conclusion, EVI1- protein levels and EVI1-CtBP1 interaction dynamics vary though the cell cycle and differ between EVI1 and ΔEVI1. These data ad to the functional characterisation of the EVI1 protein in AML and will be important for the development of targeted therapeutic approaches for EVI1-driven AML.
Assuntos
Oxirredutases do Álcool/biossíntese , Relógios Biológicos , Ciclo Celular , Proteínas de Ligação a DNA/biossíntese , Regulação Leucêmica da Expressão Gênica , Leucemia Mieloide Aguda/metabolismo , Proteína do Locus do Complexo MDS1 e EVI1/biossíntese , Oxirredutases do Álcool/genética , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Humanos , Leucemia Mieloide Aguda/genética , Proteína do Locus do Complexo MDS1 e EVI1/genéticaRESUMO
Although clear cell renal cell carcinoma (ccRCC) has been shown to result in widespread aberrant cytosine methylation and loss of 5-hydroxymethylcytosine (5hmC), the prognostic impact and therapeutic targeting of this epigenetic aberrancy has not been fully explored. Analysis of 576 primary ccRCC samples demonstrated that loss of 5hmC was strongly associated with aggressive clinicopathologic features and was an independent adverse prognostic factor. Loss of 5hmC also predicted reduced progression-free survival after resection of nonmetastatic disease. The loss of 5hmC in ccRCC was not due to mutational or transcriptional inactivation of ten eleven translocation (TET) enzymes, but to their functional inactivation by l-2-hydroxyglutarate (L2HG), which was overexpressed due to the deletion and underexpression of L2HG dehydrogenase (L2HGDH). Ascorbic acid (AA) reduced methylation and restored genome-wide 5hmC levels via TET activation. Fluorescence quenching of the recombinant TET-2 protein was unaffected by L2HG in the presence of AA. Pharmacologic AA treatment led to reduced growth of ccRCC in vitro and reduced tumor growth in vivo, with increased intratumoral 5hmC. These data demonstrate that reduced 5hmC is associated with reduced survival in ccRCC and provide a preclinical rationale for exploring the therapeutic potential of high-dose AA in ccRCC.
Assuntos
5-Metilcitosina/análogos & derivados , Oxirredutases do Álcool/biossíntese , Ácido Ascórbico/farmacologia , Carcinoma de Células Renais/tratamento farmacológico , Neoplasias Renais/tratamento farmacológico , 5-Metilcitosina/metabolismo , Adulto , Oxirredutases do Álcool/genética , Animais , Carcinoma de Células Renais/enzimologia , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Feminino , Deleção de Genes , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Renais/enzimologia , Neoplasias Renais/genética , Neoplasias Renais/patologia , Masculino , CamundongosRESUMO
Epithelial-mesenchymal transition (EMT) is one of the most important mechanisms in the metastasis of various cancers, including gastric cancer (GC). In this study, we explored the putative significance of miR-644a and its role in EMT-mediated metastasis of GC. We first detected the expression of miR-644a in a cohort of 107 GC tissues using quantitative RT-PCR. The expression of miR-644a was suppressed in GC tissues and was associated with a later clinical stage and tumor metastasis. Restoring the expression of miR-644a could significantly suppress the migration and invasion of HGC-27 and SGC-7901 cells, which might be correlated to its suppressive effect on the EMT process. We also found that carboxyl-terminal-binding protein 1 (CtBP1) was a putative target gene of miR-644a in GC and might be involved in the suppressive effect. Collectively, through targeting CtBP1-mediated suppression of the EMT process, miR-644a might suppress the tumor metastasis of GC cells.
Assuntos
Oxirredutases do Álcool/biossíntese , Proteínas de Ligação a DNA/biossíntese , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica/genética , MicroRNAs/biossíntese , Neoplasias Gástricas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Humanos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismoRESUMO
Effective clearance of all-trans-retinal (atRAL) from retinal pigment epithelial (RPE) cells is important for avoiding its cytotoxicity. However, the metabolism of atRAL in RPE cells is poorly clarified. The present study was designed to analyze metabolic products of atRAL and to compare the cytotoxicity of atRAL versus its derivative all-trans-retinal dimer (atRAL-dimer) in human RPE cells. We found that all-trans-retinol (atROL) and a mixture of atRAL condensation metabolites including atRAL-dimer and A2E were generated after incubating RPE cells with atRAL for 6h, and the amount of atRAL-dimer was significantly higher than that of A2E. In the eyes of Rdh8-/- Abca4-/- mice, a mouse model with defects in retinoid cycle that displays some symbolic characteristics of age-related macular degeneration (AMD), the level of atRAL-dimer was increased compared to wild-type mice, and was even much greater than that of A2E & isomers. The cytotoxicity of atRAL-dimer was reduced compared with its precursor atRAL. The latter could provoke intracellular reactive oxygen species (ROS) overproduction, increase the mRNA expression of several oxidative stress related genes (Nrf2, HO-1, and γ-GCSh), and induce ΔΨm loss in RPE cells. By contrast, the abilities of atRAL-dimer to induce intracellular ROS and oxidative stress were much weaker versus that of concentration-matched atRAL, and atRAL-dimer exhibited no toxic effect on mitochondrial function at higher concentrations. In conclusion, the formation of atRAL-dimer during atRAL metabolic process ameliorates the cytotoxicity of atRAL by reducing oxidative stress.
Assuntos
Sobrevivência Celular/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Epitélio Pigmentado da Retina/efeitos dos fármacos , Retinaldeído/análogos & derivados , Transportadores de Cassetes de Ligação de ATP/biossíntese , Transportadores de Cassetes de Ligação de ATP/genética , Oxirredutases do Álcool/biossíntese , Oxirredutases do Álcool/genética , Animais , Linhagem Celular , DNA Complementar/biossíntese , DNA Complementar/genética , Humanos , Degeneração Macular/genética , Degeneração Macular/prevenção & controle , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Knockout , Mitocôndrias/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Espécies Reativas de Oxigênio/metabolismo , Epitélio Pigmentado da Retina/citologia , Retinaldeído/química , Retinaldeído/toxicidadeRESUMO
Lung cancer has been the most common cancer and the main cause of cancer-related deaths worldwide for several decades. PTGR1 (prostaglandin reductase 1), as a bifunctional enzyme, has been involved in the occurrence and progression of cancer. However, its impact on human lung cancer is rarely reported. In this study, we found that PTGR1 was overexpressed in lung cancer based on the analyses of Oncomine. Moreover, lentivirus-mediated shRNA knockdown of PTGR1 reduced cell viability in human lung carcinoma cells 95D and A549 by MTT and colony formation assay. PTGR1 depletion led to G2/M phase cell cycle arrest and increased the proportion of apoptotic cells in 95D cells by flow cytometry. Furthermore, silencing PTGR1 in 95D cells resulted in decreased levels of cyclin-dependent protein kinase complex (CDK1, CDK2, cyclin A2, and cyclin B1) by western blotting and then PTGR1 is positively correlated with cyclin-dependent protein by using the data mining of the Oncomine database. Therefore, our findings suggest that PTGR1 may play a role in lung carcinogenesis through regulating cell proliferation and is a potential new therapeutic strategy for lung cancer.
Assuntos
Oxirredutases do Álcool/genética , Apoptose/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Proliferação de Células/genética , Oxirredutases do Álcool/biossíntese , Carcinoma Pulmonar de Células não Pequenas/patologia , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , MasculinoRESUMO
PURPOSE: Mice lacking ATP-binding cassette transporter 4 (ABCA4) and retinol dehydrogenase 8 (RDH8) mimic features of human Stargardt disease and age-related macular degeneration. RNA-sequencing of whole eyes was done to study early gene expression changes in Abca4-/-Rdh8-/- mice. METHODS: Abca4-/-Rdh8-/- mice at 4 weeks of age were exposed to intense light. Total RNA was extracted from whole eyes and used to generate RNA libraries that were paired-end sequenced on the Illumina HiSeq 2500 device. Differentially expressed genes were annotated using Gene set enrichment analysis (GSEA). Selected genes in enriched pathways exhibiting differential expression were validated using quantitative qRT-PCR and ELISA. RESULTS: Transcriptome analysis of the whole eye identified 200 genes that were differentially expressed 24 hours after light exposure compared to no light in Abca4-/-Rdh8-/- mice. Expression of several visual cycle and photoreceptor genes were decreased, indicative of photoreceptor/RPE cell death. Gene categories of early stress response genes, inflammatory cytokines, immune factors, and JAK STAT components were upregulated. Lipocalin 2 (Lcn2) was the most upregulated early stress response gene identified. Protein LCN2 was produced by RPE cells and the neural retina after intense light exposure as well as in cultured RPE cells from mice and humans incubated with lipopolysaccharide or photoreceptor outer segments. CONCLUSIONS: Identification of important mediators involved in the crosstalk between the acute stress response and immune activation in RPE cells and the neural retina, such as LCN2, provide novel molecular targets for reducing cellular stress during retinal degeneration.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Oxirredutases do Álcool/genética , Regulação da Expressão Gênica , Luz , Lipocalina-2/genética , Estresse Oxidativo , Degeneração Retiniana/metabolismo , Transportadores de Cassetes de Ligação de ATP/biossíntese , Doença Aguda , Oxirredutases do Álcool/biossíntese , Animais , Morte Celular , Células Cultivadas , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Humanos , Lipocalina-2/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , RNA/genética , Reação em Cadeia da Polimerase em Tempo Real , Degeneração Retiniana/patologia , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/patologia , Epitélio Pigmentado da Retina/efeitos da radiação , Tomografia de Coerência ÓpticaRESUMO
Timely clearance of apoptotic cells is an important step in the resolution of ongoing inflammation and the restoration of tissue integrity and function after acute myocardial infarction. Natural products gallic acid and l-leucine are well-documented for anti-inflammatory and anabolic effects. We synthesized gallic acid-l-leucine (GAL) conjugate via direct coupling gallic acid and l-leucine. The aim of the present study was to investigate the effect of GAL conjugate on the phagocytotic activity of macrophages. By using murine macrophage cell line RAW264.7 as an in vitro model, we evaluated the effect of GAL conjugate on the phagocytic uptake of fluorescently labeled latex beads and apoptotic cardiomyocyte H9c2 cells. We found that GAL conjugate enhanced the phagocytic activity of macrophage RAW264.7 cells in a concentration-dependent manner. Further mechanistic studies revealed that the effect of GAL conjugate on macrophage phagocytosis was positively correlated with the up-regulation of leukotriene B4 12-hydroxydehydrogenase (LTB4DH) expression at both mRNA and protein levels. By ESI-MS based lipidomics profiling, GAL conjugate increased the enzymatic activities of LTB4DH, leading to the formation of lipid metabolites including 12-oxo-LTB4, 13,14-dh-oxo-PGE2 and 13,14-dh-oxo-PGF2α. Interestingly, GAL conjugate failed to increase macrophage phagocytosis upon silencing of LTB4DH by specific siRNA. Moreover, it appeared that GAL conjugate induced LTB4DH expression via activating the Nrf2/HO-1 pathway. After Nrf-2 was silenced by specific siRNA, GAL conjugate no longer induced LTB4DH expression in the Nrf2-siRNA transfected cells. Taken together, our results suggest that GAL enhances macrophage phagocytosis via sequentially activating Nrf2 and up-regulating LTB4DH expression. Thus, GAL conjugate may serve as a lead compound for the development of new anti-inflammatory drugs.
Assuntos
Oxirredutases do Álcool/biossíntese , Ácido Gálico/farmacologia , Leucina/farmacologia , Macrófagos/imunologia , Fagocitose/efeitos dos fármacos , Animais , Anti-Inflamatórios/farmacologia , Western Blotting , Cromatografia Líquida , Macrófagos/metabolismo , Camundongos , Fagocitose/imunologia , Células RAW 264.7 , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Espectrometria de Massas em Tandem , Regulação para CimaRESUMO
The significance of quantitative changes of ALDH1A1 and RDH10 gene expression in 22 non-treated multiple myeloma patients were studied. We found a direct correlation between the expression of ALDH1A1 and RDH10 genes. We showed that ALDHA1 and RDH10 expression were inversely related with expression of a key gene for all-trans-retinoic acid catabolism, CYP26A1, and correlated with expression of RARα and PPARß/ genes. In addition for the first time it was re- vealed that increased expression of ALDH1A1-RDH10-RARα- PPARß/δ pattern could be considered as adverse prognostic factor associated with a higher concentration of paraprotein and worst overall survival of patients with newly diagnosed multiple myeloma.
Assuntos
Oxirredutases do Álcool/biossíntese , Aldeído Desidrogenase/biossíntese , Regulação Neoplásica da Expressão Gênica , Mieloma Múltiplo , Proteínas de Neoplasias/biossíntese , PPAR delta/biossíntese , PPAR beta/biossíntese , Receptor alfa de Ácido Retinoico/biossíntese , Tretinoína/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Oxirredutases do Álcool/genética , Aldeído Desidrogenase/genética , Família Aldeído Desidrogenase 1 , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/genética , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/mortalidade , Mieloma Múltiplo/patologia , Proteínas de Neoplasias/genética , PPAR delta/genética , PPAR beta/genética , Retinal Desidrogenase , Receptor alfa de Ácido Retinoico/genéticaRESUMO
Doxorubicin (DOX) is widely used for the treatment of a wide range of cancers such as breast and lung cancers, and malignant lymphomas, but is generally less efficacious in gastrointestinal cancers. The most accepted explanation for the DOX refractoriness is its resistance development. Here, we established DOX-resistant phenotypes of human gastric MKN45 and colon LoVo cells by continuous exposure to incremental concentrations of the drug. While the parental MKN45 and LoVo cells expressed carbonyl reductase 1 (CBR1) highly and moderately, respectively, the gain of DOX resistance further elevated the CBR1 expression. Additionally, the DOX-elicited cytotoxicity was lowered by overexpression of CBR1 and inversely strengthened by knockdown of the enzyme using small interfering RNA or pretreating with the specific inhibitor quercetin, which also reduced the DOX refractoriness of the two resistant cells. These suggest that CBR1 is a key enzyme responsible for the DOX resistance of gastrointestinal cancer cells and that its inhibitor is useful in the adjuvant therapy. Although CBR1 is known to metabolize DOX to a less toxic anticancer metabolite doxorubicinol, its overexpression in the parental cells hardly show significant reductase activity toward low concentration of DOX. In contrast, the overexpression of CBR1 increased the reductase activity toward an oxidative stress-derived cytotoxic aldehyde 4-oxo-2-nonenal. The sensitivity of the DOX-resistant cells to 4-oxo-2-nonenal was lower than that of the parental cells, and the resistance-elicited hyposensitivity was almost completely ameliorated by addition of the CBR1 inhibitor. Thus, CBR1 may promote development of DOX resistance through detoxification of cytotoxic aldehydes, rather than the drug's metabolism.
Assuntos
Oxirredutases do Álcool/biossíntese , Antibióticos Antineoplásicos/farmacologia , Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos/fisiologia , Oxirredutases do Álcool/antagonistas & inibidores , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Linhagem Celular Tumoral , Neoplasias Gastrointestinais/metabolismo , Humanos , Fator 2 Relacionado a NF-E2/metabolismo , Quercetina/farmacologia , Regulação para CimaRESUMO
Carbonyl reductase 1 (CBR1), an enzyme belonging to the short-chain dehydrogenases/reductases family, has been detected in all human tissues. CBR1 catalyzes the reduction of many xenobiotics, including important drugs (e.g. anthracyclines, nabumetone, bupropion, dolasetron) and harmful carbonyls and quinones. Moreover, it participates in the metabolism of a number of endogenous compounds and it may play a role in certain pathologies. Plant polyphenols are not only present in many human food sources, but are also a component of many popular dietary supplements and herbal medicines. Many studies reviewed herein have demonstrated the potency of certain flavonoids, stilbenes and curcuminoids in the inhibition of the activity of CBR1. Interactions of these polyphenols with transcriptional factors, which regulate CBR1 expression, have also been reported in several studies. As CBR1 plays an important role in drug metabolism as well as in the protection of the organism against potentially harmful carbonyls, the modulation of its expression/activity may have significant pharmacological and/or toxicological consequences. Some polyphenols (e.g. luteolin, apigenin and curcumin) have been shown to be very potent CBR1 inhibitors. The inhibition of CBR1 seems useful regarding the increased efficacy of anthracycline therapy, but it may cause the worse detoxification of reactive carbonyls. Nevertheless, all known information about the interactions of polyphenols with CBR1 have only been based on the results of in vitro studies. With respect to the high importance of CBR1 and the frequent consumption of polyphenols, in vivo studies would be very helpful for the evaluation of risks/benefits of polyphenol interactions with CBR1.
Assuntos
Oxirredutases do Álcool/antagonistas & inibidores , Oxirredutases do Álcool/metabolismo , Polifenóis/farmacologia , Oxirredutases do Álcool/biossíntese , Oxirredutases do Álcool/genética , Animais , Bupropiona/metabolismo , Butanonas/metabolismo , Butirofenonas/metabolismo , Daunorrubicina/metabolismo , Doxorrubicina/metabolismo , Regulação Enzimológica da Expressão Gênica , Haloperidol/metabolismo , Humanos , Indóis/metabolismo , Nabumetona , Neoplasias/enzimologia , Fenilpropionatos/metabolismo , Quinolizinas/metabolismo , Especificidade por Substrato , Xenobióticos/metabolismoRESUMO
BACKGROUND: Neuroblastoma (NB) is an aggressive childhood malignancy in children up to 5 years of age. High-stage tumors frequently relapse even after aggressive multimodal treatment, and then show therapy resistance, typically resulting in patient death. New molecular-targeted compounds that effectively suppress tumor growth and prevent relapse with more efficacy are urgently needed. We and others previously showed that polyamines (PA) like spermidine and spermine are essential for NB tumorigenesis and that DFMO, an inhibitor of the key PA synthesis gene product ODC, is effective both in vitro and in vivo, securing its evaluation in NB clinical trials. To find additional compounds interfering with PA biosynthesis, we tested sulfasalazine (SSZ), an FDA-approved salicylate-based anti-inflammatory and immune-modulatory drug, recently identified to inhibit sepiapterin reductase (SPR). We earlier presented evidence for a physical interaction between ODC and SPR and we showed that RNAi-mediated knockdown of SPR expression significantly reduced native ODC enzyme activity and impeded NB cell proliferation. METHODS: Human NB mRNA expression datasets in the public domain were analyzed using the R2 platform. Cell viability, isobologram, and combination index analyses as a result of SSZ treatment with our without DFMO were carried out in NB cell cultures. Molecular protein-ligand docking was achieved using the GRAMM algorithm. Statistical analyses were performed with the Kruskal-Wallis test, 2log Pearson test, and Student's t test. RESULTS: In this study, we show the clinical relevance of SPR in human NB tumors. We found that high SPR expression is significantly correlated to unfavorable NB characteristics like high age at diagnosis, MYCN amplification, and high INSS stage. SSZ inhibits the growth of NB cells in vitro, presumably due to the inhibition of SPR as predicted by computational docking of SSZ into SPR. Importantly, the combination of SSZ with DFMO produces synergistic antiproliferative effects in vitro. CONCLUSIONS: The results suggest the use of SSZ in combination with DFMO for further experiments, and possible prioritization as a novel therapy for the treatment of NB patients.
Assuntos
Oxirredutases do Álcool/genética , Neuroblastoma/genética , Proteínas/genética , Sulfassalazina/administração & dosagem , Oxirredutases do Álcool/biossíntese , Oxirredutases do Álcool/metabolismo , Carcinogênese/efeitos dos fármacos , Carcinogênese/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Criança , Pré-Escolar , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Lactente , Masculino , Terapia de Alvo Molecular , Neuroblastoma/tratamento farmacológico , Neuroblastoma/patologia , Poliaminas/metabolismo , Proteínas/metabolismoRESUMO
CtBP2, as a transcriptional corepressor of epithelial-specific genes, has been reported to promote tumor due to upregulating epithelial-mesenchymal transition (EMT) in cancer cells. CtBP2 was also demonstrated to contribute to the proliferation of esophageal squamous cell carcinoma (ESCC) cells through a negative transcriptional regulation of p16(INK4A). In this study, for the first time, we reported that CtBP2 expression, along with CCNH/CDK7, was higher in ESCC tissues with lymph node metastases than in those without lymph node metastases. Moreover, both CtBP2 and CCNH/CDK7 were positively correlated with E-cadherin, tumor grade, and tumor metastasis. However, the concrete mechanism of CtBP2's role in enhancing ESCC migration remains incompletely understood. We confirmed that CCNH/CDK7 could directly interact with CtBP2 in ESCC cells in vivo and in vitro. Furthermore, our data demonstrate for the first time that CtBP2 enhanced the migration of ESCC cells in a CCNH/CDK7-dependent manner. Our results indicated that CCNH/CDK7-CtBP2 axis may augment ESCC cell migration, and targeting the interaction of both may provide a novel therapeutic target of ESCC.
Assuntos
Oxirredutases do Álcool/biossíntese , Carcinoma de Células Escamosas/genética , Ciclina H/biossíntese , Quinases Ciclina-Dependentes/biossíntese , Neoplasias Esofágicas/genética , Proteínas do Tecido Nervoso/biossíntese , Idoso , Oxirredutases do Álcool/genética , Carcinoma de Células Escamosas/patologia , Movimento Celular/genética , Proliferação de Células/genética , Proteínas Correpressoras , Ciclina H/genética , Inibidor p16 de Quinase Dependente de Ciclina/biossíntese , Inibidor p16 de Quinase Dependente de Ciclina/genética , Quinases Ciclina-Dependentes/genética , Transição Epitelial-Mesenquimal , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/genética , Quinase Ativadora de Quinase Dependente de CiclinaRESUMO
Retinal disease is the major cause of irreversible blindness in developed countries. Transplantation of photoreceptor precursor cells (PPCs) derived from human embryonic stem cells (hESCs) is a promising and widely applicable approach for the treatment of these blinding conditions. Previously, it has been shown that after transplantation into the degenerating retina, the percentage of PPCs that undergo functional integration is low. The factors that inhibit PPC engraftment remain largely unknown, in part, because so many adverse factors could be at play during in vivo experiments. To advance our knowledge in overcoming potential adverse effects and optimize PPC transplantation, we have developed a novel ex vivo system. Harvested neural retina was placed directly on top of cultured retinal pigment epithelial (RPE) cells from a number of different sources. To mimic PPC transplantation into the subretinal space, hESC-derived PPCs were inserted between the retinal explant and underlying RPE. Explants cocultured with hESC-derived RPE maintained normal gross morphology and viability for up to 2 weeks, whereas the explants cultured on ARPE19 and RPE-J failed by 7 days. Furthermore, the proportion of PPCs expressing ribbon synapse-specific proteins BASSOON and RIBEYE was significantly higher when cocultured with hESC-derived RPE (20% and 10%, respectively), than when cocultured with ARPE19 (only 6% and 2%, respectively). In the presence of the synaptogenic factor thrombospondin-1 (TSP-1), the proportion of BASSOON-positive and RIBEYE-positive PPCs cocultured with hESC-derived RPE increased to â¼30% and 15%, respectively. These data demonstrate the utility of an ex vivo model system to define factors, such as TSP-1, which could influence integration efficiency in future in vivo experiments in models of retinal degeneration.
Assuntos
Células-Tronco Embrionárias Humanas/citologia , Células Fotorreceptoras de Vertebrados/citologia , Retina/citologia , Trombospondina 1/metabolismo , Oxirredutases do Álcool/biossíntese , Oxirredutases do Álcool/genética , Animais , Diferenciação Celular , Linhagem Celular , Transplante de Células , Proteínas Correpressoras , Técnicas de Cocultura , Células Epiteliais/citologia , Humanos , Camundongos , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Técnicas de Cultura de Órgãos , Ratos , Epitélio Pigmentado da Retina/citologiaRESUMO
C-terminal binding protein 2 (CtBP2) is a transcriptional co-repressor that promotes cancer cell migration and invasion by inhibiting multiple tumor suppressor genes that contribute to cell mobility and adhesion. In this investigation, we showed thatCtBP2 expression was increased significantly in HCC tissues when compared to matched normal adjacent liver tissues. We also showed that CtBP2 expression is associated with worse HCC patient prognosis after liver resection. CtBP2 over-expression induced epithelial-mesenchymal transition (EMT) in Huh7 cells and, correspondingly, silencing CtBP2 suppressed EMT in MHCC97H cells. ChIP assays revealed that GLI1 increased CtBP2 transcription by directly binding its promoter. Furthermore, interaction of CtBP2 and Snail Family Zinc Finger 1 (SNAI1), both of which were found to be positively regulated by GLI1, was confirmed by Co-IP assay. SNAI1 knockdown revealed that SNAI1 was essential for CtBP2 induction of the EMT phenotype of HCC cells, and CtBP2 knockdown reversed GLI1-SNAI1 driven EMT in Huh7 cells. Finally, in vivo experiments demonstrated that enhanced CtBP2expression promoted HCC xenograft growth and induced EMT. In conclusion, CtBP2 may serve as a prognostic marker for post liver resection HCC and may play a role during GLI1-driven EMT as a transcriptional co-repressor of SNAI1.
Assuntos
Oxirredutases do Álcool/biossíntese , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/patologia , Proteínas do Tecido Nervoso/biossíntese , Fatores de Transcrição/biossíntese , Oxirredutases do Álcool/genética , Animais , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proteínas Correpressoras , Transição Epitelial-Mesenquimal , Feminino , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/genética , Prognóstico , Fatores de Transcrição da Família Snail , Fatores de Transcrição/genética , Proteína GLI1 em Dedos de ZincoRESUMO
Polycystic ovary syndrome (PCOS) is currently considered a predominantly hyperandrogenic syndrome. In theory, hyperandrogenism can be caused by high level of testosterone (T) as well as by enhanced androgen receptor (AR) activity. C-Terminal binding protein 1 antisense (CTBP1-AS) was a novel long noncoding RNA (lncRNA) to regulate AR activity. In this study, we found that expression level of CTBP1-AS in peripheral blood leukocytes was significantly higher in women with PCOS than that in controls after adjustment for age and body mass index (BMI). Individuals having higher expression of CTBP1-AS had significantly greater disease risk than those having lower expression. We also identified expression of CTBP1-AS as an independent risk factor for PCOS. A positive correlation was observed between the CTBP1-AS expression and the total T (TT) concentration either unadjusted or after adjusting for age, BMI, and homeostatic model assessment insulin resistance. Taken together, our current study presented the first evidence that the lncRNA CTBP1-AS, a novel AR modulator, is associated with PCOS in Chinese population and established the possibility that abnormal CTBP1-AS expression is a risk factor for PCOS and it is a predictor of variability in serum TT level in Chinese women with PCOS.
Assuntos
Oxirredutases do Álcool/biossíntese , Oxirredutases do Álcool/genética , Povo Asiático/genética , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/genética , Síndrome do Ovário Policístico/genética , Síndrome do Ovário Policístico/metabolismo , Receptores Androgênicos/fisiologia , Adulto , Feminino , HumanosRESUMO
PURPOSE: Ulcerative colitis and colitis-associated colorectal cancer (CAC) is a serious health issue, but etiopathological factors remain unclear. Aldo-keto reductase 1B10 (AKR1B10) is specifically expressed in the colonic epithelium, but downregulated in colorectal cancer. This study was aimed to investigate the etiopathogenic role of AKR1B10 in ulcerative colitis and CAC. EXPERIMENTAL DESIGN: Ulcerative colitis and CAC biopsies (paraffin-embedded sections) and frozen tissues were collected to examine AKR1B10 expression. Aldo-keto reductase 1B8 (the ortholog of human AKR1B10) knockout (AKR1B8(-/-)) mice were produced to estimate its role in the susceptibility and severity of chronic colitis and associated dysplastic lesions, induced by dextran sulfate sodium (DSS) at a low dose (2%). Genome-wide exome sequencing was used to profile DNA damage in DSS-induced colitis and tumors. RESULTS: AKR1B10 expression was markedly diminished in over 90% of ulcerative colitis and CAC tissues. AKR1B8 deficiency led to reduced lipid synthesis from butyrate and diminished proliferation of colonic epithelial cells. The DSS-treated AKR1B8(-/-) mice demonstrated impaired injury repair of colonic epithelium and more severe bleeding, inflammation, and ulceration. These AKR1B8(-/-) mice had more severe oxidative stress and DNA damage, and dysplasias were more frequent and at a higher grade in the AKR1B8(-/-) mice than in wild-type mice. Palpable masses were seen in the AKR1B8(-/-) mice only, not in wild-type. CONCLUSIONS: AKR1B8 is a critical protein in the proliferation and injury repair of the colonic epithelium and in the pathogenesis of ulcerative colitis and CAC, being a new etiopathogenic factor of these diseases.
Assuntos
Oxirredutases do Álcool/genética , Colite Ulcerativa/patologia , Colo/patologia , Mucosa Intestinal/patologia , Oxirredutases Atuantes sobre Doadores de Grupos Aldeído ou Oxo/metabolismo , Oxirredutases do Álcool/biossíntese , Oxirredutases do Álcool/metabolismo , Aldo-Ceto Redutases , Animais , Sequência de Bases , Proliferação de Células , Transformação Celular Neoplásica/genética , Colite Ulcerativa/induzido quimicamente , Neoplasias Colorretais/patologia , Dano ao DNA/genética , Sulfato de Dextrana , Modelos Animais de Doenças , Células Epiteliais/metabolismo , Humanos , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Estresse Oxidativo/genética , Oxirredutases Atuantes sobre Doadores de Grupos Aldeído ou Oxo/biossíntese , Oxirredutases Atuantes sobre Doadores de Grupos Aldeído ou Oxo/genética , Espécies Reativas de Oxigênio/metabolismo , Análise de Sequência de DNARESUMO
BACKGROUND: The hypermethylation of Alcohol dehydrogenase iron containing 1 (ADHFE1) was recently reported to be associated with colorectal cancer (CRC) differentiation. However, the effect of alcohol on ADHFE1 hypermethylation in CRC is still unclear. METHODS: The methylation status and expression levels of ADHFE1 were investigated in primary tumor tissues and adjacent normal tissues of 73 patients with CRC, one normal colon cell line, and 4 CRC cell lines (HT-29, SW480, DLD-1, and LoVo) by quantitative methylation-specific polymerase chain reaction (QMSP) and real-time reverse transcription polymerase chain reaction (real time PCR), respectively. The effect of alcohol on the methylation status of ADHFE1 was analyzed in HT-29, SW480, DLD-1, and CCD18Co cells using QMSP, real-time PCR, immunoblot, and cell proliferation assay. RESULTS: ADHFE1 was hypermethylated in 69 of 73 CRC tissues (95%) compared to adjacent normal tissues (p<0.05). The mRNA expression of ADHFE1 was significantly reduced in CRC compared to adjacent normal tissues (p<0.05) and its expression was decreased in the alcohol consumption group (p<0.05). ADHFE1 was hypermethylated and its expression was decreased in 4 CRC cell lines compared with normal colon cell line. Alcohol induced hypermethylation of ADHFE1, decreased its expression, and stimulated cell proliferation of HT-29, SW480, and DLD-1cells. CONCLUSION: These results demonstrate that the promoter hypermethylation of ADHFE1 is frequently present in CRC and alcohol induces methylation-mediated down expression of ADHFE1 and proliferation of CRC cells.
Assuntos
Oxirredutases do Álcool/genética , Álcoois/toxicidade , Neoplasias Colorretais/genética , Metilação de DNA/efeitos dos fármacos , Proteínas Mitocondriais/genética , Oxirredutases do Álcool/biossíntese , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/induzido quimicamente , Neoplasias Colorretais/patologia , Regulação Neoplásica da Expressão Gênica , Células HT29 , Humanos , Proteínas Mitocondriais/biossíntese , Regiões Promotoras Genéticas/efeitos dos fármacosRESUMO
To identify novel tumor-associated proteins, we analyzed the protein expression patterns from experimental hepatocellular carcinoma (HCC) that were induced using hepatocarcinogenesis models in rats. Rats were subjected to two previously described protocols of hepatocarcinogenesis using diethylnitrosamine as a carcinogen: the alternative Solt-Farber (aS&F) protocol, which induces HCC within 9 months, and Schiffer's model, which induces cirrhosis and multifocal HCC within 18 weeks. The patterns of protein expression from tumors and normal liver tissue were examined by SDS-PAGE and the bands identified at 33-34 kDa were analyzed by mass spectrometry. The prostaglandin reductase 1 (PTGR1) showed the highest number of peptides, with a confidence of level >99%. The increased expression of PTGR1 in tumors was confirmed in these two models by Western blotting and by increase in alkenal/one oxidoreductase activity (25-fold higher than normal liver). In addition, the gene expression level of Ptgr1, as measured by qRT-PCR, was increased during cancer development in a time-dependent manner (200-fold higher than normal liver). Furthermore, PTGR1 was detected in the cytoplasm of neoplastic cells in rat tumors and in 12 human HCC cases by immunohistochemistry. These analyses were performed by comparing the expression of PTGR1 to that of two well-known markers of hepatocarcinoma, Glutathione S-transferase pi 1 (GSTP1) in rats and glypican-3 in humans. The increased expression and activity of PTGR1 in liver carcinogenesis encourage further research aimed at understanding the metabolic role of PTGR1 in HCC and its potential application for human cancer diagnosis and treatment.
Assuntos
Oxirredutases do Álcool/biossíntese , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Proteínas de Neoplasias/biossíntese , Oxirredutases do Álcool/genética , Animais , Carcinoma Hepatocelular/patologia , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas Experimentais/genética , Neoplasias Hepáticas Experimentais/patologia , RatosRESUMO
High-throughput techniques have identified numerous antisense (AS) transcripts and long non-coding RNAs (ncRNAs). However, their significance in cancer biology remains largely unknown. Here, we report an androgen-responsive long ncRNA, CTBP1-AS, located in the AS region of C-terminal binding protein 1 (CTBP1), which is a corepressor for androgen receptor. CTBP1-AS is predominantly localized in the nucleus and its expression is generally upregulated in prostate cancer. CTBP1-AS promotes both hormone-dependent and castration-resistant tumour growth. Mechanistically, CTBP1-AS directly represses CTBP1 expression by recruiting the RNA-binding transcriptional repressor PSF together with histone deacetylases. CTBP1-AS also exhibits global androgen-dependent functions by inhibiting tumour-suppressor genes via the PSF-dependent mechanism thus promoting cell cycle progression. Our findings provide new insights into the functions of ncRNAs that directly contribute to prostate cancer progression.