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1.
Phytother Res ; 30(6): 971-80, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26989859

RESUMO

Phytoestrogens are selective estrogen receptor modulators (SERMs) with potential for use in hormone replacement therapy (HRT) to relieve peri/postmenopausal symptoms. This study was aimed at elucidating the molecular mechanisms underlying the SERM properties of the extract of Korean-grown Opuntia ficus-indica (KOFI). The KOFI extract induced estrogen response element (ERE)-driven transcription in breast and endometrial cancer cell lines and the expression of endogenous estrogen-responsive genes in breast cancer cells. The flavonoid content of different KOFI preparations affected ERE-luciferase activities, implying that the flavonoid composition likely mediated the estrogenic activities in cells. Oral administration of KOFI decreased the weight gain and levels of both serum glucose and triglyceride in ovariectomized (OVX) rats. Finally, KOFI had an inhibitory effect on the 17ß-estradiol-induced proliferation of the endometrial epithelium in OVX rats. Our data demonstrate that KOFI exhibited SERM activity with no uterotrophic side effects. Therefore, KOFI alone or in combination with other botanical supplements, vitamins, or minerals may be an effective and safe alternative active ingredient to HRTs, for the management of postmenopausal symptoms. Copyright © 2016 John Wiley & Sons, Ltd.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Opuntia/química , Receptores de Estrogênio/química , Animais , Feminino , Humanos , Extratos Vegetais/farmacologia , Ratos , Ratos Sprague-Dawley , Transfecção
3.
Cytotherapy ; 17(10): 1421-33, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26212610

RESUMO

BACKGROUND AIMS: It is important to improve the migratory ability of dendritic cells (DCs) and to increase DC potency for successful DC-based cancer immunotherapy. The intracellular Ca(2+) signaling pathway has an important role on the regulation of DC migration. Our preliminary studies revealed that sarco/endoplasmic reticulum Ca(2+) transport ATPase 2 (SERCA2) expression was inversely related to DC migratory capacity, and the expression level of p-cofilin and SERCA2 on mature DCs showed a counter-trend. METHODS: We selected the appropriate six maturation cocktails on the basis of the expression levels of SERCA2 and p-cofilin and investigated the functional characteristics and migratory capacity of mature DCs. Among the these six maturation cocktails, DCIFN-γ/IL-1ß/Poly-I:C showed potent type 1 immune response with interleukin (IL)-12p70 production and strong Th1-polarization, and this DC elicited strong antigen-specific cytotoxic T-lymphocyte responses. RESULTS: Interestingly, DCIFN-γ/IL-1ß/Poly-I:C showed lower expression of SERCA2 and higher expression of p-cofilin compared with those matured with the use of other cocktails. In vitro migration assay showed that DCs matured with the use of this maturation cocktail had significantly increased migratory ability compared with αDC1s and other DCs. CONCLUSIONS: Interferon-γ, IL-1ß and Poly-I:C maturation cocktail may be used in the field of cancer immunotherapy to generate potent immune-stimulatory DCs with improved type 1 immune response and migration capacity.


Assuntos
Fatores de Despolimerização de Actina/metabolismo , Movimento Celular/imunologia , Células Dendríticas/imunologia , Imunoterapia/métodos , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Linfócitos T Citotóxicos/imunologia , Adenosina Trifosfatases , Sinalização do Cálcio/imunologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células , Células Cultivadas , Células Dendríticas/transplante , Retículo Endoplasmático/metabolismo , Humanos , Interferon gama/farmacologia , Interleucina-10/metabolismo , Interleucina-12/metabolismo , Interleucina-1beta/farmacologia , Subunidade p19 da Interleucina-23/metabolismo , Neoplasias/terapia , Poli I-C/farmacologia
4.
Bioorg Med Chem Lett ; 24(7): 1730-3, 2014 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-24630560

RESUMO

Rhus verniciflua Stokes has been used as a traditional herbal medicine in Asia. In this study, the effect of R. verniciflua extract on human aromatase (cytochrome P450 19, CYP19) activity was investigated to elucidate the mechanism for the effect of R. verniciflua extract on androgen hormone levels. Androstenedione was used as a substrate and incubated with R. verniciflua extract in cDNA-expressed CYP19 supersomes in the presence of NADPH, and estrone formation was measured using liquid chromatography-tandem mass spectrometry. R. verniciflua extract was assessed at concentrations of 10-1000 µg/mL. The resulting data showed that R. verniciflua extract inhibited CYP19-mediated estrone formation in a concentration-dependent manner with an IC50 value of 136 µg/mL. Subsequently, polyphenolic compounds from R. verniciflua extract were tested to identify the ingredients responsible for the aromatase inhibitory effects by R. verniciflua extract. As a result, butin showed aromatase inhibitory effect in a concentration-dependent manner with an IC50 value of 9.6 µM, whereas the inhibition by other compounds was negligible. These results suggest that R. verniciflua extract could modulate androgen hormone levels via the inhibition of CYP19 activity and butin is a major ingredient responsible for this activity.


Assuntos
Inibidores da Aromatase/farmacologia , Aromatase/metabolismo , Benzopiranos/farmacologia , Extratos Vegetais/farmacologia , Estruturas Vegetais/química , Rhus/química , Inibidores da Aromatase/química , Inibidores da Aromatase/isolamento & purificação , Benzopiranos/química , Benzopiranos/isolamento & purificação , Relação Dose-Resposta a Droga , Humanos , Medicina Tradicional , Estrutura Molecular , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Relação Estrutura-Atividade
5.
Nature ; 452(7183): 98-102, 2008 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-18288109

RESUMO

Cytokines affect a variety of cellular functions, including regulation of cell numbers by suppression of programmed cell death. Suppression of apoptosis requires receptor signalling through the activation of Janus kinases and the subsequent regulation of members of the B-cell lymphoma 2 (Bcl-2) family. Here we demonstrate that a Bcl-2-family-related protein, Hax1, is required to suppress apoptosis in lymphocytes and neurons. Suppression requires the interaction of Hax1 with the mitochondrial proteases Parl (presenilin-associated, rhomboid-like) and HtrA2 (high-temperature-regulated A2, also known as Omi). These interactions allow Hax1 to present HtrA2 to Parl, and thereby facilitates the processing of HtrA2 to the active protease localized in the mitochondrial intermembrane space. In mouse lymphocytes, the presence of processed HtrA2 prevents the accumulation of mitochondrial-outer-membrane-associated activated Bax, an event that initiates apoptosis. Together, the results identify a previously unknown sequence of interactions involving a Bcl-2-family-related protein and mitochondrial proteases in the ability to resist the induction of apoptosis when cytokines are limiting.


Assuntos
Metaloproteases/metabolismo , Proteínas Mitocondriais/metabolismo , Processamento de Proteína Pós-Traducional , Proteínas/metabolismo , Serina Endopeptidases/metabolismo , Animais , Apoptose , Sobrevivência Celular , Genes Letais , Serina Peptidase 2 de Requerimento de Alta Temperatura A , Peptídeos e Proteínas de Sinalização Intracelular , Linfócitos/citologia , Linfócitos/metabolismo , Metaloproteases/deficiência , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Mitocondriais/química , Proteínas Mitocondriais/deficiência , Neurônios/citologia , Neurônios/metabolismo , Ligação Proteica , Proteínas/genética , Serina Endopeptidases/química , Proteína X Associada a bcl-2/metabolismo
6.
Ann Hematol ; 90(12): 1419-26, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21465188

RESUMO

For wide application of a dendritic cell (DC) vaccination in myeloma patients, easily available tumor antigens should be developed. We investigated the feasibility of cellular immunotherapy using autologous alpha-type 1-polarized dendritic cells (αDC1s) loaded with apoptotic allogeneic myeloma cells, which could generate myeloma-specific cytotoxic T lymphocytes (CTLs) against autologous myeloma cells in myeloma patients. Monocyte-derived DCs were matured by adding the αDC1-polarizing cocktail (TNFα/IL-1ß/IFN-α/IFN-γ/poly-I:C) and loaded with apoptotic allogeneic CD138(+) myeloma cells from other patients with matched monoclonal immunoglobulins as a tumor antigen. There were no differences in the phenotypic expression between αDC1s loaded with apoptotic autologous and allogeneic myeloma cells. Autologous αDC1s effectively took up apoptotic allogeneic myeloma cells from other patients with matched subtype. Myeloma-specific CTLs against autologous target cells were successfully induced by αDC1s loaded with allogeneic tumor antigen. The cross-presentation of apoptotic allogeneic myeloma cells to αDC1s could generate CTL responses between myeloma patients with individual matched monoclonal immunoglobulins. There was no difference in CTL responses between αDC1s loaded with autologous tumor antigen and allogeneic tumor antigen against targeting patient's myeloma cells. Our data indicate that autologous DCs loaded with allogeneic myeloma cells with matched immunoglobulin can generate potent myeloma-specific CTL responses against autologous myeloma cells and can be a highly feasible and effective method for cellular immunotherapy in myeloma patients.


Assuntos
Apresentação de Antígeno/imunologia , Antígenos de Neoplasias/imunologia , Apresentação Cruzada/imunologia , Células Dendríticas/imunologia , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/patologia , Vacinas Anticâncer/imunologia , Células Dendríticas/citologia , Citometria de Fluxo , Humanos , Fenótipo , Linfócitos T Citotóxicos/imunologia , Transplante Autólogo/imunologia
7.
Ann Hematol ; 89(8): 795-801, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20238114

RESUMO

To induce a potent cytotoxic T lymphocyte (CTL) response, various tumor antigens should be loaded onto dendritic cells (DCs). In multiple myeloma (MM), it is difficult to obtain a sufficient number of autologous tumor cells as a source of tumor antigens in the clinical setting. We investigated the feasibility of immunotherapy in patients with MM, using myeloma-specific CTLs generated in vitro by alpha-type 1-polarized DCs (alphaDC1s) loaded with the ultraviolet B-irradiated allogeneic myeloma cell line, ARH77. alphaDC1s significantly increased the expression of several costimulatory molecules without differences in loading with tumor antigens. alphaDC1s showed a high production of interleukin-12 during maturation and after subsequent stimulation with CD40L but were not significantly affected by loading tumor antigens. Myeloma-specific CTLs against autologous myeloma cells from MM patients were induced by alphaDC1s pulsed with apoptotic ARH77 cells. Our data indicate that autologous DCs loaded with an allogeneic myeloma cell line can generate potent myeloma-specific CTL responses against autologous myeloma cells and might provide a practical method for cellular immunotherapy in patients with MM.


Assuntos
Apoptose/imunologia , Linhagem Celular Tumoral/imunologia , Células Dendríticas/imunologia , Mieloma Múltiplo , Transplante de Neoplasias , Linfócitos T Citotóxicos/imunologia , Transplante Homólogo , Apresentação de Antígeno/imunologia , Antígenos de Neoplasias/imunologia , Humanos , Imunoterapia/métodos , Ativação Linfocitária/imunologia , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/patologia , Linfócitos T Citotóxicos/citologia
8.
J Steroid Biochem Mol Biol ; 112(1-3): 117-21, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18832037

RESUMO

We previously demonstrated that the expression of Mullerian inhibiting substance (MIS) in Sertoli cells is downregulated by tumor necrosis factor alpha (TNF-alpha), which is secreted by meiotic germ cells, in mouse testes. Several studies have reported that MIS that is secreted by Sertoli cells inhibits steroidogenesis and, thus, the synthesis of testosterone in testicular Leydig cells. Here, we demonstrate that in TNF-alpha knockout testes, which show high levels of MIS, steroidogenesis is decreased compared to that in wild-type testes. The levels of testosterone and the mRNA levels of steroidogenesis-related genes were significantly lower after puberty in TNF-alpha knockout testes than in wild-type testes. Furthermore, the number of sperm was reduced in TNF-alpha knockout mice. Histological analysis revealed that spermatogenesis is also delayed in TNF-alpha knockout testes. In conclusion, TNF-alpha knockout mice show reduced testicular steroidogenesis, which is likely due to the high level of testicular MIS compared to that seen in wild-type mice.


Assuntos
Células de Sertoli/fisiologia , Testículo/fisiologia , Testosterona/metabolismo , Fator de Necrose Tumoral alfa/fisiologia , Animais , Hormônio Antimülleriano/biossíntese , Masculino , Camundongos , Camundongos Knockout , Espermatogênese , Testículo/citologia , Fator de Necrose Tumoral alfa/genética
9.
Mol Cell Biol ; 25(12): 4841-52, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15923603

RESUMO

The SWI3-related gene product (SRG3), a component of the mouse SWI/SNF complex, has been suggested to have an alternative function. Here, we demonstrate that in the prostate transactivation of the androgen receptor (AR) is modulated by SRG3 in multiple ways. The expression of SRG3, which is developmentally regulated in the prostate, is induced by androgen through AR. SRG3 in turn enhances the transactivation of AR, providing a positive feedback regulatory loop. The SRG3 coactivation of AR transactivation is achieved through the recruitment of coactivator SRC-1, the protein level of which is upregulated by SRG3, providing another pathway of positive regulation. Interestingly, SRG3 coactivation of AR transactivation is fully functional in BRG1/BRM-deficient C33A cells and the AR/SRG3/SRC-1 complex formed in vivo contains neither BRG1 nor BRM protein, suggesting the possibility of an SRG3 function independent of the SWI/SNF complex. Importantly, the AR/SRG3/SRC-1 complex occupies androgen response elements on the endogenous SRG3 and PSA promoter in an androgen-dependent manner in mouse prostate and LNCaP cells, respectively, inducing gene expression. These results suggest that the multiple positive regulatory mechanisms of AR transactivation by SRG3 may be important for the rapid proliferation of prostate cells during prostate development and regeneration.


Assuntos
Receptores Androgênicos/metabolismo , Fatores de Transcrição/metabolismo , Ativação Transcricional , Acetiltransferases/genética , Acetiltransferases/metabolismo , Animais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Histona Acetiltransferases , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Coativador 1 de Receptor Nuclear , Coativadores de Receptor Nuclear , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Regiões Promotoras Genéticas , Próstata/citologia , Próstata/fisiologia , Ratos , Ratos Sprague-Dawley , Receptores Androgênicos/genética , Fatores de Transcrição/genética , Técnicas do Sistema de Duplo-Híbrido , Fatores de Transcrição de p300-CBP
10.
Mol Cell Biol ; 23(17): 6000-12, 2003 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-12917325

RESUMO

The expression of Mullerian inhibiting substance (MIS), a key molecule in sex differentiation and reproduction, is tightly regulated. It has been suggested that meiotic germ cells repress MIS expression in testicular Sertoli cells, although the substance responsible for this cell-cell communication remains unknown. Here, we present the cytokine tumor necrosis factor alpha (TNF-alpha) as a strong candidate for such a substance and its downstream molecular events. TNF-alpha inhibited MIS expression in testis organ cultures, and TNF-alpha(-/-) testes showed high and prolonged MIS expression. Furthermore, in transient-transfection assays TNF-alpha suppressed the MIS promoter that was activated by steroidogenic factor 1 (SF-1), one of the major transcription factors that regulate MIS expression. The modulation of SF-1 transactivation by TNF-alpha is through the activation of NF-kappa B, which subsequently interacts with SF-1 and represses its transactivation. The physical association of NF-kappa B with SF-1 was shown by yeast two-hybrid protein interaction, glutathione S-transferase pull-down, and coimmunoprecipitation (ChIP) analyses. ChIP assays also revealed that endogenous NF-kappa B, as well as SF-1, is recruited to the MIS promoter upon TNF-alpha signaling. SF-1-bound NF-kappa B subsequently recruits histone deacetylases to inhibit the SF-1-activated gene expression. These results may identify, for the first time, the responsible substance and its action mechanism underlying the repression of MIS expression by meiotic germ cells in the testis.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Regulação para Baixo/fisiologia , Glicoproteínas , Inibidores do Crescimento/metabolismo , NF-kappa B/genética , Hormônios Testiculares/metabolismo , Testículo/fisiologia , Fatores de Transcrição/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Hormônio Antimülleriano , Células Cultivadas , Proteínas de Ligação a DNA/genética , Inibidores Enzimáticos/farmacologia , Fatores de Transcrição Fushi Tarazu , Inibidores do Crescimento/genética , Células HeLa/efeitos dos fármacos , Inibidores de Histona Desacetilases , Histona Desacetilases/metabolismo , Proteínas de Homeodomínio , Humanos , Ácidos Hidroxâmicos/farmacologia , Técnicas In Vitro , Masculino , Camundongos , Camundongos Endogâmicos ICR , Camundongos Knockout , NF-kappa B/metabolismo , Regiões Promotoras Genéticas , Ratos , Ratos Sprague-Dawley , Receptores Citoplasmáticos e Nucleares , Células de Sertoli/metabolismo , Fator Esteroidogênico 1 , Hormônios Testiculares/genética , Fatores de Transcrição/genética , Fator de Necrose Tumoral alfa/genética
11.
Mol Cell Biol ; 24(7): 2593-604, 2004 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15024051

RESUMO

Tumor necrosis factor alpha (TNF-alpha) has been demonstrated to inhibit steroidogenesis in Leydig cells at the transcriptional level of steroidogenic enzymes. However, the molecular mechanism of this observed gene repression is not well understood. We now demonstrate that nuclear factor kappaB (NF-kappaB) activated by TNF-alpha inhibits the transactivation of orphan nuclear receptors, which regulate the expression of steroidogenic-enzyme genes. TNF-alpha treatment suppressed the luteinizing-hormone-induced or Nur77/SF-1-stimulated promoter activity of steroidogenic-enzyme genes in Leydig cells. The TNF-alpha-mediated gene suppression was blocked by treatment with an inhibitor of NF-kappaB. In addition, overexpression of the p65 (RelA) subunit of NF-kappaB showed the same effect as TNF-alpha and inhibited Nur77 transactivation, suggesting the involvement of NF-kappaB activation in the observed gene repression. Physical association of Nur77 with p65 was revealed by mammalian two-hybrid, GST pull-down, and coimmunoprecipitation analyses. The NF-kappaB inhibition of Nur77 transactivation was likely due to the competition of p65 for Nur77 binding with coactivators. Finally, chromatin immunoprecipitation assays revealed that TNF-alpha treatment caused the recruitment of NF-kappaB to the promoter of the steroidogenic-enzyme p450c17 gene, supporting the hypothesis that the TNF-alpha-mediated gene repression involves NF-kappaB inhibition of the transcriptional activity of Nur77 and other orphan nuclear receptors. These findings provide a molecular mechanism underlying the inhibition of testicular steroidogenesis by proinflammatory cytokines.


Assuntos
Regulação Enzimológica da Expressão Gênica , Células Intersticiais do Testículo/metabolismo , NF-kappa B/metabolismo , Esteroides/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Linhagem Celular , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Histona Acetiltransferases , Histona Desacetilases/metabolismo , Humanos , Células Intersticiais do Testículo/citologia , Masculino , Camundongos , Coativador 1 de Receptor Nuclear , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Progesterona/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , Ratos , Receptores Citoplasmáticos e Nucleares , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Testosterona/metabolismo , Fator de Transcrição RelA , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
12.
Mol Endocrinol ; 20(5): 984-95, 2006 May.
Artigo em Inglês | MEDLINE | ID: mdl-16455820

RESUMO

The basic leucine zipper transcription factor, CCAAT enhancer-binding protein-alpha (C/EBPalpha), negatively regulates cell proliferation and induces terminal differentiation of various cell types. C/EBPalpha is expressed in the prostate, but its potential role in the tissue is unknown. Herein, we show that C/EBPalpha is highly expressed at the stage of growth arrest during prostate development. Furthermore, overexpression of C/EBPalpha decreases the rate of DNA synthesis in LNCaP prostate cancer cells. Investigation of the potential cross-talk between C/EBPalpha and androgen receptor (AR) that is responsible for androgen-dependent prostate proliferation demonstrates that androgen-dependent transactivation of AR is strongly repressed by C/EBPalpha. C/EBPalpha directly binds AR in vitro and forms a complex with AR in vivo. C/EBPalpha neither prevents the nuclear translocation of AR nor disrupts the N/C-terminal interaction of AR, which are both necessary for its proper transactivation activity upon ligand binding. To modulate AR transactivation, however, C/EBPalpha does compete with AR coactivators for AR binding. Additionally, C/EBPalpha is recruited onto AR-target promoters with AR and is further able to inhibit the expression of endogenous prostate-specific antigen in prostate cancer cells. Our results suggest C/EBPalpha as a potent AR corepressor and provide insight into the role of C/EBPalpha in prostate development and cancer.


Assuntos
Antagonistas de Receptores de Andrógenos , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Proteínas Repressoras/metabolismo , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/análise , Linhagem Celular Tumoral , Proliferação de Células , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Regiões Promotoras Genéticas , Próstata/química , Próstata/crescimento & desenvolvimento , Próstata/metabolismo , Antígeno Prostático Específico/genética , Neoplasias da Próstata/química , Neoplasias da Próstata/genética , Ratos , Receptores Androgênicos/genética , Proteínas Repressoras/análise , Ativação Transcricional
13.
Mol Endocrinol ; 19(9): 2245-57, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15919722

RESUMO

Androgen receptor (AR) is important in male sexual differentiation and testicular function. Here, we demonstrate the regulation of AR expression and its transactivation by the basic helix-loop-helix (bHLH) transcription factor Pod-1, the expression of which in postnatal testis reciprocally coincides with the expression of AR. Pod-1 represses the promoter activity of AR, possibly through its E-box. An AR promoter region of 169 bp, which harbors one canonical E-box, is sufficient for the Pod-1-repression and bound by purified Pod-1 proteins. Pod-1 also suppresses the transactivation of AR. Transient transfection analyses of mammalian cells show that Pod-1 represses AR transactivation in a dose-dependent manner. Furthermore, yeast two-hybrid, glutathione-S-transferase-pull-down, and co-immunoprecipitation analyses reveal that Pod-1 directly associates with AR through its N-terminal region and through the DNA binding-hinge domain of AR. Interestingly, Pod-1 recruits histone deacetylase (HDAC)-1 to inhibit both promoter activity and transactivation of AR. Overexpression of HDAC1 further inhibits the Pod-1-mediated repressions and Pod-1 directly interacts with HDAC1. Furthermore, chromatin immunoprecipitation assay reveals that HDAC1 is recruited with Pod-1 to the endogenous AR promoter and the androgen-regulated Pem promoter. Taken together, these results suggest that Pod-1, which controls AR transcription and function, may play an important role in the development and function of the testis.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação para Baixo , Regulação da Expressão Gênica , Histona Desacetilases/metabolismo , Receptores Androgênicos/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Células Cultivadas , Imunoprecipitação da Cromatina , Elementos E-Box , Histona Desacetilase 1 , Proteínas de Homeodomínio/genética , Humanos , Masculino , Camundongos , Regiões Promotoras Genéticas , Receptores Androgênicos/genética , Deleção de Sequência , Testículo/crescimento & desenvolvimento , Fatores de Transcrição/genética , Ativação Transcricional , Técnicas do Sistema de Duplo-Híbrido
14.
Exp Mol Med ; 48(8): e253, 2016 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-27538371

RESUMO

The migration of dendritic cells (DCs) to secondary lymphoid organs depends on chemoattraction through the interaction of the chemokine receptors with chemokines. However, the mechanism of how lymphoid chemokines attract DCs to lymphoid organs remains unclear. Here, we demonstrate the mechanism of DC migration in response to the lymphoid chemokine CCL21. CCL21-mediated DC migration is controlled by the regulation of sarcoplasmic reticulum Ca(2+) ATPase 2 (SERCA2) expression rather than through the activation of mitogen-activated protein kinases CCL21-exposed mature DCs (mDCs) exhibited decreased SERCA2 expression but not decreased phospholamban (PLB) or Hax-1 expression, which are known to be SERCA2-interacting proteins. In addition, CCL21 did not affect the mRNA levels of SERCA2 or its interacting protein Hax-1. Interestingly, SERCA2 expression was inversely related to DC migration in response to chemokine stimulation. The migratory capacity of CCL21-treated mDCs was decreased by the phospholipase C inhibitor U73122 and by the protein kinase C inhibitor BAPTA-AM. The migratory capacities of mDCs were increased in response to SERCA2 siRNA expression but were decreased by SERCA2 overexpression. In addition, DCs treated with a SERCA2-specific inhibitor (cyclopiazonic acid) had significantly increased migratory capacities as mDCs regardless of SERCA2 expression. Moreover, SERCA2 expression was dependent on DC maturation induced by cytokines or Toll-like receptor agonists. Therefore, the migratory capacities differed in differentially matured DCs. Taken together, these results suggest that SERCA2 contributes to the migration of CCL21-activated DCs as an important feature of the adaptive immune response and provide novel insights regarding the role of SERCA2 in DC functions.


Assuntos
Quimiocina CCL21/imunologia , Quimiotaxia , Células Dendríticas/imunologia , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/imunologia , Imunidade Adaptativa , Células Cultivadas , Células Dendríticas/citologia , Humanos , Monócitos/citologia , Monócitos/imunologia
15.
Mol Endocrinol ; 18(1): 13-25, 2004 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-14576337

RESUMO

Androgen receptor (AR) that mediates androgen action is a crucial factor in male reproductive functions. Here, we report a novel AR corepressor ARR19 (androgen receptor corepressor-19 kDa), which has been isolated as a putative androgen-induced gene from murine testis. ARR19 encoding a leucine-rich protein is expressed only in male reproductive organs such as testis and prostate. ARR19 expression in the testis is developmentally regulated. Functional analysis conducted by the transient transfection of mammalian cells shows that ARR19 represses AR transactivation in a dose-dependent manner. Furthermore, yeast two-hybrid and glutathione S-transferase pull-down analyses reveal that ARR19 directly associates with AR through the N-terminal and leucine zipper-containing regions of ARR19 and the DNA binding-hinge domain of AR. Interestingly, ARR19 localized in the cytoplasmic compartment cotranslocates into the nucleus with AR upon androgen exposure. The ARR19 repression of AR transactivation is through the recruitment of histone deacetylase 4 (HDAC4) by ARR19. Overexpression of HDAC4 further inhibits the ARR19-repressed AR transactivation. In addition, ARR19 directly interacts with HDAC4 in vitro. Furthermore, DNA-protein complex immunoprecipitation assays reveal that HDAC4 is recruited to an androgen-regulated promoter through ARR19. Taken together, the results suggest that ARR19 may act as an AR corepressor in vivo and play an important role in male reproductive functions.


Assuntos
Histona Desacetilases/metabolismo , Receptores Androgênicos/genética , Proteínas Repressoras/fisiologia , Antagonistas de Receptores de Andrógenos , Animais , Linhagem Celular , Linhagem Celular Tumoral , Clonagem Molecular , DNA Complementar , Biblioteca Gênica , Células HeLa , Humanos , Proteínas com Domínio MARVEL , Masculino , Proteínas de Membrana , Camundongos , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/metabolismo , Testículo/fisiologia , Ativação Transcricional , Transfecção
16.
Mol Endocrinol ; 18(8): 1929-40, 2004 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-15155786

RESUMO

DAX-1 (dosage-sensitive sex reversal adrenal hypoplasia congenital critical region on the X chromosome, gene 1) (NROB1) is an atypical member of the nuclear receptor family, which lacks the classical zinc finger DNA binding domain and acts as a coregulator of a number of nuclear receptors. In this study, we have found that DAX-1 is a novel coregulator of the orphan nuclear receptor Nur77 (NR4A1). We demonstrate that DAX-1 represses the Nur77 transactivation by transient transfection assays. Specific interaction between Nur77 and DAX-1 was detected by coimmunoprecipitation, yeast two-hybrid, and glutathione-S-transferase pull-down assays. The ligand binding domain of DAX-1 and the activation function-2 domain of Nur77 were determined as the direct interaction domains between DAX-1 and Nur77. In vitro competition binding assay showed that DAX-1 repressed Nur77 transactivation through the competition with steroid receptor coactivator-1 for the binding of Nur77. Moreover, DAX-1 repressed Nur77- and LH-dependent increase of cytochrome P450 protein 17 promoter activity in transient transfection assays. Furthermore, Nur77-mediated transactivation was significantly increased by down-regulation of DAX-1 expression with DAX-1 small interfering RNA in testicular Leydig cell line, K28. LH treatment induced a transient increase in Nur77 mRNA, whereas LH repressed DAX-1 expression in a time- and dose-dependent manner in K28 cells. In addition, immunohistochemical analysis showed the expression of Nur77 in mouse testicular Leydig cells. These results suggest that DAX-1 acts as a novel coregulator of the orphan nuclear receptor Nur77, and that the DAX-1 may play a key role in the regulation of Nur77-mediated steroidogenesis in testicular Leydig cells.


Assuntos
Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/metabolismo , Receptores do Ácido Retinoico/metabolismo , Proteínas Repressoras/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Ativação Transcricional , Animais , Ligação Competitiva , Linhagem Celular , Receptor Nuclear Órfão DAX-1 , Proteínas de Ligação a DNA/genética , Expressão Gênica/efeitos dos fármacos , Histona Acetiltransferases , Células Intersticiais do Testículo/citologia , Células Intersticiais do Testículo/efeitos dos fármacos , Células Intersticiais do Testículo/metabolismo , Hormônio Luteinizante/farmacologia , Masculino , Camundongos , Coativador 1 de Receptor Nuclear , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares , Regiões Promotoras Genéticas/genética , Ligação Proteica , Receptores Citoplasmáticos e Nucleares , Receptores do Ácido Retinoico/genética , Receptores de Esteroides , Proteínas Repressoras/genética , Esteroide 17-alfa-Hidroxilase/genética , Fatores de Transcrição/genética , Ativação Transcricional/genética
17.
J Med Food ; 18(1): 21-30, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25372471

RESUMO

Rhus verniciflua stokes (RVS) is known to promote blood circulation by preventing blood stasis, although the active ingredients and the underlying mechanism are unclear. Platelets are the primary cells that regulate circulation and contribute to the development of diverse cardiovascular diseases by aggregation and thrombosis. The study assessed the antiplatelet activity of RVS and sought to identify the active constituents. Pretreatment of washed platelets with RVS heartwood extract blunted the aggregatory response of platelets to collagen. In the subfractions, fisetin, butein, and sulfuretin were identified as effective inhibitors of platelet aggregation by collagen, thrombin, and adenosine-5'-diphosphate. Antiplatelet activities of all three compounds were concentration dependent, and fisetin had longer in vitro duration of action compared with butein or sulfuretin. Extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase activation by collagen was prevented by fisetin, whereas butein and sulfuretin failed to inhibit ERK and p38 activation was not affected by any of the compounds. Rats orally administered 100 mg/(kg·day(-1)) fisetin for 7 days were resistant to arterial thrombosis, although total extract of RVS heartwood exhibited little effect at a dose of 1000 mg/(kg·day(-1)). RVS heartwood may have cardiovascular protective activity by inhibiting platelet aggregation. The active constituents are fisetin, butein, and sulfuretin, and fisetin is orally effective against thrombosis.


Assuntos
Benzofuranos/farmacologia , Plaquetas/efeitos dos fármacos , Chalconas/farmacologia , Flavonoides/farmacologia , Inibidores da Agregação Plaquetária/farmacologia , Agregação Plaquetária/efeitos dos fármacos , Rhus/química , Animais , Benzofuranos/uso terapêutico , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/prevenção & controle , Chalconas/uso terapêutico , Relação Dose-Resposta a Droga , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Flavonoides/uso terapêutico , Flavonóis , Masculino , Fitoterapia , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Inibidores da Agregação Plaquetária/uso terapêutico , Ratos Sprague-Dawley , Trombose/metabolismo , Trombose/prevenção & controle , Madeira/química , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
18.
Biomol Ther (Seoul) ; 23(3): 283-9, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25995828

RESUMO

The half-dried leaves of Stewartia. pseudocamellia were extracted with hot water (SPE) and partitioned with n-hexane (SPEH), dichloromethane (SPED), and ethyl acetate (SPEE) successively. SPE and SPEE showed significant inhibitory effects against melanogenesis and tyrosinase activities. By bioassay-guided isolation, ten phenolic compounds were isolated by column chromatography from SPEE. The whitening effect of the isolated compounds from SPEE were tested for the inhibitory activities against melanogenesis using B16 melanoma cells, in vitro inhibition of tyrosinase, and L-3,4-dihydorxy-indole-2-carboxylic acid (L-DOPA) auto-oxidation assay. A cytotoxic activity assay was done to examine the cellular toxicity in Raw 264.7 macrophage cells. Of the compounds isolated, gallic acid and quercetin revealed significant inhibitory activities against melanogenesis compared to arbutin. In particular, quercetin exhibited similar inhibitory activities against tyrosinase and L-DOPA oxidation without cytotoxicity. These results suggested that SPE could be used as a potential source of natural skin-whitening material in cosmetics as well as in food products.

19.
Arch Pharm Res ; 38(11): 2049-58, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25784057

RESUMO

Rhus verniciflua stokes (RVS) is a popular medicinal plant in oriental medicines which is commonly used to resolve extravasated blood. To elucidate the molecular mechanism of the role of RVS extracts on the regulation of lipid and cholesterol biosynthesis, we investigated whether RVS extract protect the hyperlipidemia in western diet-induced C57BL6/J mice. Mice fed a western diet and additionally RVS extracts was administered orally at a dose of 0.1 or 1 g/kg/day for 2 weeks respectively. Group with higher dose of RVS extract showed a significantly decreased body weight compared with western diet fed mice groups. And total cholesterol, LDL-cholesterol levels and fatty liver formation were also improved especially in group of mice fed western diet supplemented high dose RVS extracts. Next, synthesis of hepatic bile acids were significantly increased in RVS extract fed groups. Furthermore, RVS extracts significantly increase promoter activity of Cyp7a1 via up-regulate the transcriptional expression level of LXRα. Our data suggest that RVS extracts could be a potent therapeutic ingredient for prevent a hyperlipidemia via increase of bile acids biosynthesis.


Assuntos
Ácidos e Sais Biliares/biossíntese , Hiperlipidemias/prevenção & controle , Extratos Vegetais/farmacologia , Rhus/química , Animais , Colesterol/biossíntese , Colesterol 7-alfa-Hidroxilase/metabolismo , Dieta Ocidental/efeitos adversos , Relação Dose-Resposta a Droga , Lipídeos/biossíntese , Receptores X do Fígado , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores Nucleares Órfãos/genética , Extratos Vegetais/administração & dosagem
20.
J Immunother ; 37(1): 16-25, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24316552

RESUMO

Flagellin, the cognate ligand for toll-like receptor 5, has potent adjuvant activity in various vaccines. However, its efficacy in generating dendritic cells (DCs) remains contentious. This study assessed how efficaciously Vibrio vulnificus FlaB (v-FlaB) could be used in generating a potent DC to induce antigen-specific cytotoxic T lymphocytes (CTLs). Mature DCs (mDCs) induced by the combination of v-FlaB/TNFα/IFNα were significantly more potent in inducing specific anticancer immune responses compared with the standard DCs that were maturated by the conventional cytokine cocktail of TNFα/IL-1ß/IL-6/PGE(2). The potent mDCs produced a higher level of interleukin (IL)-12p70 and polarized naive CD4(+) T cells more towards Th1-type cells, markedly increased antigen-specific CD8(+) T-cell number and significantly enhanced the induction of lytic enzymes in antigen-specific CD8(+) CTLs and sensitized CD3(+) T cells to produce higher number of interferon (IFN)γ-secreting cells. As a result, the mDCs produced more potent antigen-specific CTLs against the MART-1 and expressed higher levels of homing receptors CCR5 and CXCR3. More importantly, the v-FlaB/TNFα/IFNα-DCs generated from melanoma patients produced strong autologous CTLs with efficient cytotoxic activities. In conclusion, v-FlaB combined with tumor necrosis factor (TNF)α and IFNα can generate potent DCs which produce functionally active CTLs and that may have potential as a potent cancer vaccine.


Assuntos
Citocinas/farmacologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Flagelina/imunologia , Mediadores da Inflamação/farmacologia , Neoplasias/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Movimento Celular/imunologia , Citotoxicidade Imunológica/efeitos dos fármacos , Células Dendríticas/metabolismo , Humanos , Interleucina-12/biossíntese , Ligantes , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Receptores CCR7/metabolismo , Células Th1/efeitos dos fármacos , Células Th1/imunologia
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