RESUMEN
Leydig cells produce androgens that are essential for male sex differentiation and reproductive function. Leydig cell function is regulated by several hormones and signaling molecules, including growth hormone (GH). Although GH is known to upregulate Star gene expression in Leydig cells, its molecular mechanism of action remains unknown. The STAT5B transcription factor is a downstream effector of GH signaling in other systems. While STAT5B is present in both primary and Leydig cell lines, its function in these cells has yet to be ascertained. Here we report that treatment of MA-10 Leydig cells with GH or overexpression of STAT5B induces Star messenger RNA levels and increases steroid hormone output. The mouse Star promoter contains a consensus STAT5B element (TTCnnnGAA) at -756 bp to which STAT5B binds in vitro (electrophoretic mobility shift assay and supershift) and in vivo (chromatin immunoprecipitation) in a GH-induced manner. In functional promoter assays, STAT5B was found to activate a -980 bp mouse Star reporter. Mutating the -756 bp element prevented STAT5B binding but did not abrogate STAT5B-responsiveness. STAT5B was found to functionally cooperate with DNA-bound cJUN. The STAT5B/cJUN cooperation was only observed in Leydig cells and not in Sertoli or fibroblast cells, indicating that additional Leydig cell-enriched transcription factors are required. The STAT5B/cJUN cooperation was lost only when both STAT5B and cJUN elements were mutated. In addition to identifying the Star gene as a novel target for STAT5B in Leydig cells, our data provide important new insights into the mechanism of GH and STAT5B action in the regulation of Leydig cell function.
Asunto(s)
Hormona del Crecimiento/farmacología , Células Intersticiales del Testículo/metabolismo , Fosfoproteínas/genética , Proteínas Proto-Oncogénicas c-jun/fisiología , Factor de Transcripción STAT5/genética , Animales , Secuencia de Bases , Sitios de Unión , Línea Celular , ADN/química , ADN/metabolismo , Expresión Génica/efectos de los fármacos , Células Intersticiales del Testículo/clasificación , Masculino , Ratones , Fosfoproteínas/análisis , Fosfoproteínas/fisiología , Regiones Promotoras Genéticas , ARN Mensajero/análisis , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/fisiología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Secretory carcinoma of the salivary glands is a relatively new disease concept, and is characterized by "morphological resemblance to mammary secretory carcinoma and ETV6-NTRK3 gene fusion." Herein we describe a confusing case and briefly discuss practical diagnostic problems. The patient was a 71-year-old Japanese man who had a tumor consistent with secretory carcinoma at the microscopic and immunohistochemical levels. Immunohistochemically, EMA and S100 protein were noted to be positive along with various cytokeratins as well as mammaglobin and pSTAT5. Moreover, vimentin was focally positive. Smooth muscle actin, p63, p40, and androgen receptor were negative. However, a search using fluorescence in situ hybridization did not reveal a definite split signal for the ETV6 gene. It is presumed that confirming the diagnosis of secretory carcinoma without genetic retrieval will be accepted as a diagnostic method, and we hope that worldwide general recognition may earlier reach "gradual acceptance."
Asunto(s)
Biomarcadores de Tumor/análisis , Carcinoma Secretor Análogo al Mamario/diagnóstico , Neoplasias de la Parótida/diagnóstico , Proteínas Proto-Oncogénicas c-ets/análisis , Proteínas Represoras/análisis , Anciano , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunohistoquímica , Queratinas/análisis , Queratinas/genética , Masculino , Carcinoma Secretor Análogo al Mamario/metabolismo , Carcinoma Secretor Análogo al Mamario/patología , Neoplasias de la Parótida/metabolismo , Neoplasias de la Parótida/patología , Proteínas S100/análisis , Proteínas S100/genética , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/genética , Proteína ETS de Variante de Translocación 6RESUMEN
A growth hormone (GH) injection is able to induce the phosphorylated form of the signal transducer and activator of transcription 5 (pSTAT5) in a large number of cells throughout the mouse brain. The present study had the objective to map the distribution of GH-responsive cells in the brain of rats that received an intracerebroventricular injection of GH and compare it to the pattern found in mice. We observed that rats and mice exhibited a similar distribution of GH-induced pSTAT5 in the majority of areas of the telencephalon, hypothalamus and brainstem. However, rats exhibited a higher density of GH-responsive cells than mice in the horizontal limb of the diagonal band of Broca (HDB), supraoptic and suprachiasmatic nuclei, whereas mice displayed more GH-responsive cells than rats in the hippocampus, lateral hypothalamic area and dorsal motor nucleus of the vagus (DMX). Since both HDB and DMX contain acetylcholine-producing neurons, pSTAT5 was co-localized with choline acetyltransferase in GH-injected animals. We found that 50.0 ± 4.5% of cholinergic neurons in the rat HDB coexpressed GH-induced pSTAT5, whereas very few co-localizations were observed in the mouse HDB. In contrast, rats displayed fewer cholinergic neurons responsive to GH in the DMX at the level of the area postrema. In summary, pSTAT5 can be used as a marker of GH-responsive cells in the rat brain. Although rats and mice exhibit a relatively similar distribution of GH-responsive neurons, some species-specific differences exist, as exemplified for the responsiveness to GH in distinct populations of cholinergic neurons.
Asunto(s)
Mapeo Encefálico/métodos , Receptores de Somatotropina/análisis , Factor de Transcripción STAT5/análisis , Acetilcolina , Animales , Encéfalo/metabolismo , Tronco Encefálico/metabolismo , Colina O-Acetiltransferasa/metabolismo , Neuronas Colinérgicas/metabolismo , Hormona del Crecimiento/metabolismo , Hormona del Crecimiento/farmacología , Hipocampo/metabolismo , Hipotálamo/metabolismo , Infusiones Intraventriculares , Masculino , Bulbo Raquídeo/metabolismo , Ratones , Ratones Endogámicos C57BL , Fosforilación , Ratas , Ratas Long-Evans , Receptores de Somatotropina/metabolismo , Factor de Transcripción STAT5/metabolismoRESUMEN
Secretory carcinoma (SC) of the salivary gland is a relatively newly described disease, separate from acinic cell carcinoma (ACC), which frequently displays ETV6-NTRK3 gene fusion. However, the differences between SC and ACC remain unclear. Here, histological reevaluation of 12 formerly diagnosed ACC cases was performed, which yielded a new diagnosis of SC in four cases due to a lack of obvious acinar-like cells. Immunohistochemically, phosphorylated signal transducer and activator of transcription 5 (p-STAT5) was expressed in SC but not in ACC, whereas discovered on GIST-1 (DOG1) was expressed in ACC but not in SC. Molecular analysis was possible in three SC cases, of which two showed the ETV6-NTRK3 fusion transcript on reverse-transcription polymerase chain reaction, as well as breaks in the ETV6 gene on fluorescence in situ hybridization. However, the remaining SC cases did not show this fusion transcript. Recently, several reports have suggested that SC might not be adequately diagnosed if the focus is placed solely on the ETV6-NTRK3 fusion gene due to genetic diversity. In this regard, immunohistochemistry of p-STAT5 and DOG1 is expected to be a useful alternative diagnostic tool to discriminate SC from ACC.
Asunto(s)
Biomarcadores de Tumor/análisis , Carcinoma de Células Acinares/diagnóstico , Inmunohistoquímica , Proteínas de Fusión Oncogénica/genética , Neoplasias de la Parótida/diagnóstico , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Anoctamina-1/análisis , Anoctamina-1/metabolismo , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Carcinoma de Células Acinares/genética , Carcinoma de Células Acinares/patología , Errores Diagnósticos , Femenino , Heterogeneidad Genética , Humanos , Hibridación Fluorescente in Situ , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/análisis , Proteínas de Neoplasias/metabolismo , Glándula Parótida/patología , Neoplasias de la Parótida/genética , Neoplasias de la Parótida/patología , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/metabolismo , Adulto JovenRESUMEN
BACKGROUND: This study was conducted to investigate the treatment efficacies and immunological mechanisms of action of dioscin in mice with chicken collagen type II-induced arthritis (CIA). METHODS: The CIA mice was randomly divided into the model group (M), dioscin group (D), and tripterygium group (T); a normal control group (C) was also included. Each group was orally administered with related drugs or an equal volume of solvent (group C) starting on the 21st day of primary immunity, after which the levels of T helper 17 cells (Th17), regulatory T cells (Tregs), and their related factors were detected on the 35th day. RESULTS: Compared to group C, group M exhibited significantly increased levels of interleukin 17 (IL-17) and IL-6 and decreased IL-27 (p < 0.05). Group D exhibited significantly decreased levels of IL-17 and IL-6 compared with group M (p < 0.05). Group M showed a significantly increased ratio of Th17 cells (p < 0.05), while dioscin significantly reduced this ratio (p < 0.05). Groups M and C showed no significant difference in the ratio of Tregs (p > 0.05) but dioscin significantly increased this ratio (p < 0.05). Group M significantly increased signal transducer and activator of transcription 3 (STAT3) and STAT5 compared with that in group C (p < 0.05), while the T and D groups showed significantly reduced levels of STAT3 and STAT5 (p < 0.05). CONCLUSION: Dioscin may affect the differentiation of Th17 and Tregs and secretion of related factors by regulating CD4 T cell subset-related signal transduction and the expression of transcription-activating factor STAT3 and STAT5, thus exerting useful immunoregulatory roles in CIA mice.
Asunto(s)
Artritis Experimental/tratamiento farmacológico , Diosgenina/análogos & derivados , Linfocitos T Reguladores/efectos de los fármacos , Células Th17/efectos de los fármacos , Animales , Artritis Experimental/inmunología , Pollos , Colágeno Tipo II , Diosgenina/farmacología , Interleucina-17/sangre , Interleucina-6/sangre , Interleucinas/fisiología , Masculino , Ratones , Ratones Endogámicos DBA , Factor de Transcripción STAT3/análisis , Factor de Transcripción STAT5/análisis , Linfocitos T Reguladores/inmunología , Células Th17/inmunologíaRESUMEN
OBJECTIVE: The purpose of this study was to reduce the time to tumor onset in a diethylnitrosamine (DEN)-induced hepatocellular carcinoma (HCC) swine model via partial liver embolization (PLE) and to characterize the model for use in translational research. METHODS: Eight Yucatan miniature pigs were injected intraperitoneally with either saline (n = 2) or DEN (n = 6) solution weekly for 12 weeks. Three of the DEN-treated pigs underwent PLE. The animals underwent periodic radiological evaluation, liver biopsy, and blood sampling, and full necropsy was performed at study termination (â¼29 months). RESULTS: All DEN-treated pigs developed hepatic adenoma and HCC. PLE accelerated the time to adenoma development but not to HCC development. Biomarker analysis results showed that IGF1 levels decreased in all DEN-treated pigs as functional liver capacity decreased with progression of HCC. VEGF and IL-6 levels were positively correlated with disease progression. Immunohistochemical probing of HCC tissues demonstrated the expression of several important survival-promoting proteins. CONCLUSION: To our knowledge, we are the first to demonstrate an accelerated development of hepatic neoplasia in Yucatan miniature pigs. Our HCC swine model closely mimics the human condition (i.e., progressive disease stages and expression of relevant molecular markers) and is a viable translational model.
Asunto(s)
Adenoma/sangre , Adenoma/patología , Carcinoma Hepatocelular/sangre , Modelos Animales de Enfermedad , Neoplasias Hepáticas Experimentales/sangre , Neoplasias Hepáticas Experimentales/patología , Adenoma/inducido químicamente , Animales , Carcinoma Hepatocelular/inducido químicamente , Carcinoma Hepatocelular/patología , Dietilnitrosamina , Embolia/inducido químicamente , Femenino , Hormona del Crecimiento/sangre , Factor I del Crecimiento Similar a la Insulina/metabolismo , Interleucina-6/sangre , Janus Quinasa 2/análisis , Neoplasias Hepáticas Experimentales/inducido químicamente , Vena Porta , Receptores de Somatomedina/análisis , Factor de Transcripción STAT3/análisis , Factor de Transcripción STAT5/análisis , Porcinos , Porcinos Enanos , Factores de Tiempo , Factor A de Crecimiento Endotelial Vascular/sangre , alfa-Fetoproteínas/metabolismoRESUMEN
BACKGROUND: Regulatory T cells (Tregs) are key players in immune tolerance. They express the transcription factor FOXP3 and are dependent of the STAT5 signaling for their homeostasis. So far, the study of phosphorylated epitopes by flow cytometry required treating the cells with methanol, which is harmful for several epitopes. METHODS: Here we assessed whether the PerFix EXPOSE reagent kit (PFE)(Beckman Coulter) allowed monitoring the phosphorylation level of STAT5 in Treg subpopulations together with complex immunophenotyping. Results observed with the PFE kit were compared to those observed without cell permeabilization for surface markers, with paraformaldehyde permeabilization for non-phosphorylated intracellular epitopes, and with methanol-based permeabilization for phosphoSTAT5 staining. RESULTS: In human PBMCs, the PFE kit allowed the detection of surface antigens, FOXP3, KI67 and phosphoSTAT5 in similar proportions to what was observed without permeabilization (for surface antigens), or with PFA or methanol permeabilizations for FOXP3/KI67 and phosphoSTAT5, respectively. Comparable observations were made with murine splenocytes. Further, the PFE kit allowed determining the response of different human and murine Treg subsets to IL-2. It also allowed demonstrating that human Treg subsets with the highest levels of phosphoSTAT5 had also the highest suppressive activity in vitro, and that anti-thymocyte glogulin (ATG) induced Treg independently of the STAT5 pathway, both in vitro and in vivo. CONCLUSIONS: We have validated a multicolor staining method that allows monitoring phosphoSTAT5 levels in Treg subsets. This staining could be useful to monitor responses of various Treg subsets to IL-2 therapy.
Asunto(s)
Factor de Transcripción STAT5/análisis , Coloración y Etiquetado/métodos , Subgrupos de Linfocitos T/inmunología , Linfocitos T Reguladores/inmunología , Adulto , Animales , Femenino , Citometría de Flujo/métodos , Xenoinjertos , Humanos , Inmunofenotipificación/métodos , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Fosforilación , Adulto JovenRESUMEN
Few studies have comprehensively analysed histopathological findings of bone marrow clots for diagnosis of haematopoietic cell dysplasia. In particular, a limited number of studies have assessed the use of haematoxylin and eosin (H&E) staining, which is generally considered less informative than May-Giemsa staining. In the current study, the utility of bone marrow clot specimens for diagnosis was examined using H&E staining and immunohistochemistry. Patients with myelodysplastic syndromes (MDS) and myelodysplastic/myeloproliferative neoplasm (MDS/MPN), including chronic myelomonocytic leukaemia (CMML), atypical chronic myeloid leukaemia (aCML) lacking Philadelphia chromosome, and juvenile myelomonocytic leukaemia (JMML), were selected for histological evaluation. H&E stained specimens were advantageous for observation of atypical basophilic staining of the cytoplasm and nucleus related to dysplasia. This finding was significantly supported for both MDS and MDS/MPN (pâ<â0.05 versus May-Giemsa staining); therefore, we concluded that H&E staining could be used for identification of dysplastic cells. In addition, despite the loss of tissue structure, phosphorylated Stat5 immunostaining was sufficiently useful for the observation of myelodysplastic blasts. Thus, clot specimens are useful for diagnosis of haematopoietic dysplasia by pathologists.
Asunto(s)
Citodiagnóstico/métodos , Enfermedades Mielodisplásicas-Mieloproliferativas/diagnóstico , Factor de Transcripción STAT5/biosíntesis , Coloración y Etiquetado/métodos , Biopsia con Aguja , Médula Ósea/patología , Eosina Amarillenta-(YS) , Hematoxilina , Humanos , Inmunohistoquímica , Fosforilación , Factor de Transcripción STAT5/análisisRESUMEN
It has been previously shown that the long-term inhibition of milking-induced prolactin (PRL) release by quinagolide (QN), a dopamine agonist, reduces milk yield in dairy cows. To further demonstrate that PRL is galactopoietic in cows, we performed a short-term experiment that used PRL injections to restore the release of PRL at milking in QN-treated cows. Nine Holstein cows were assigned to treatments during three 5-d periods in a 3×3 Latin square design: 1) QN: twice-daily i.m. injections of 1mg of QN; 2) QN-PRL: twice-daily i.m. injections of 1mg of QN and twice-daily (at milking time) i.v. injections of PRL (2µg/kg body weight); and 3) control: twice-daily injections of the vehicles. Mammary epithelial cells (MEC) were purified from milk so that their viability could be assessed, and mammary biopsies were harvested for immunohistological analyses of cell proliferation using PCNA and STAT5 staining. In both milk-purified MEC and mammary tissue, the mRNA levels of milk proteins and BAX were determined using real-time reverse-transcription PCR. Daily QN injections reduced milking-induced PRL release. The area under the PRL curve was similar in the control and PRL injection treatments, but the shape was different. The QN treatment decreased milk, lactose, protein, and casein production. Injections of PRL did not restore milk yield but tended to increase milk protein yield. In mammary tissue, the percentage of STAT5-positive cells was reduced during QN but not during QN-PRL in comparison with the control treatment. The percentage of PCNA-positive cells was greater during QN-PRL injections than during the control or QN treatment and tended to be lower during QN than during the control treatment. In milk-purified MEC, κ-casein and α-lactalbumin mRNA levels were lower during QN than during the control treatment, but during QN-PRL, they were not different from the control treatment. In mammary tissue, the BAX mRNA level was lower during QN-PRL than during QN. The number of MEC exfoliated into milk was increased by QN injections but tended to be decreased by PRL injections. Injections of PRL also increased the viability of MEC harvested from milk. Although PRL injections at milking could not reverse the effect of QN treatment on milk production, their effects on cell survival and exfoliation and on gene expression suggest that the effect of QN treatment on the mammary gland is due to QN's inhibition of PRL secretion.
Asunto(s)
Aminoquinolinas/administración & dosificación , Bovinos/metabolismo , Lactancia/efectos de los fármacos , Glándulas Mamarias Animales/efectos de los fármacos , Prolactina/administración & dosificación , Prolactina/antagonistas & inhibidores , Animales , Caseínas/metabolismo , Proliferación Celular/efectos de los fármacos , Suplementos Dietéticos , Agonistas de Dopamina/farmacología , Células Epiteliales/química , Células Epiteliales/citología , Femenino , Lactalbúmina/metabolismo , Lactosa/análisis , Glándulas Mamarias Animales/química , Glándulas Mamarias Animales/citología , Leche/citología , Proteínas de la Leche/genética , Antígeno Nuclear de Célula en Proliferación/análisis , ARN Mensajero/análisis , Factor de Transcripción STAT5/análisisRESUMEN
Messenger RNA (mRNA), a single-strand ribonucleic acid with functional gene information is usually abnormally expressed in cancer cells and has become a promising biomarker for the study of tumor progress. Hairpin DNA-coated gold nanoparticle (hDAuNP) beacon containing a bare gold nanoparticle (AuNP) as fluorescence quencher and thiol-terminated fluorescently labeled stem-loop-stem oligonucleotide sequences attached by Au-S bond is currently a new nanoscale biodiagnostic platform capable of mRNA detection, in which the design of the loop region sequence is crucial for hybridizing with the target mRNA. Hence, in this study, to improve the sensitivity and selectivity of hDAuNP beacon simultaneously, the loop region of hairpin DNA was screened by bioinformatics strategy. Here, signal transducer and activator of transcription 5b (STAT5b) mRNA was selected and used as a practical example. The results from the combined characterizations using optical techniques, flow cytometry assay, and cell microscopic imaging showed that after optimization, the as-prepared hDAuNP beacon had higher selectivity and sensitivity for the detection of STAT5b mRNA in living cells, as compared with our previous beacon. Thus, the bioinformatics method may be a promising new strategy for assisting in the designing of the hDAuNP beacon, extending its application in the detection of mRNA expression and the resultant mRNA-based biological processes and disease pathogenesis.
Asunto(s)
Biología Computacional/métodos , Oro/química , Nanopartículas del Metal/química , ARN Mensajero , Factor de Transcripción STAT5 , Células Hep G2 , Humanos , ARN Mensajero/análisis , ARN Mensajero/química , ARN Mensajero/genética , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/genética , Factor de Transcripción STAT5/metabolismoRESUMEN
Prolactin (PRL) is important in the regulation of milk synthesis in mammary epithelial cells (MEC). In cattle, circulating levels of PRL are not limiting, suggesting the possible involvement of other factors that may control the response to PRL at the cellular level. The effects of milking frequency (MF) on milk synthesis are controlled locally within mammary glands and involve PRL signaling. To further investigate this relationship between MF and PRL signaling, udder halves of 17 dairy cows were milked either 4 times a day (4×) or once a day (1×) for 14 d in early lactation. Mammary biopsies were obtained 3 to 5h following milking from both udder halves of 10 cows, and changes in PRL and associated pathways were measured. The abundance of STAT5A mRNA was higher after 4× milking, whereas that of the PRL receptor (PRLR) and STAT3 were lower relative to that after 1× milking. In 4× mammary tissues, the protein levels of STAT5, activated STAT5, and ß1-integrin were higher, whereas the those of the long isoform of PRL receptor and activated STAT3 were lower than 1× tissues. The activation of STAT5 correlated strongly with major milk protein mRNA abundance (r=0.86 to 0.94) and ß1-integrin protein levels (r=0.91). These results confirm that major milk protein gene expression is associated with STAT5 activation and suggests that the STAT5 and ß1-integrin signaling pathways are linked. Modulation of ß1-integrin abundance in response to changes in MF may be a mechanism that controls the MEC ability to respond to PRL and therefore its secretory activity.
Asunto(s)
Industria Lechera/métodos , Integrina beta1/metabolismo , Lactancia/fisiología , Glándulas Mamarias Animales/metabolismo , Factor de Transcripción STAT5/metabolismo , Animales , Bovinos , Células Epiteliales , Femenino , Expresión Génica , Humanos , Integrina beta1/análisis , Glándulas Mamarias Animales/química , Leche , Proteínas de la Leche/genética , Prolactina/sangre , ARN Mensajero/análisis , ARN Mensajero/metabolismo , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/genética , Transducción de Señal/fisiologíaRESUMEN
Xbp1, a key mediator of the unfolded protein response (UPR), is activated by IRE1α-mediated splicing, which results in a frameshift to encode a protein with transcriptional activity. However, the direct function of Xbp1 in epithelial cells during mammary gland development is unknown. Here we report that the loss of Xbp1 in the mammary epithelium through targeted deletion leads to poor branching morphogenesis, impaired terminal end bud formation, and spontaneous stromal fibrosis during the adult virgin period. Additionally, epithelial Xbp1 deletion induces endoplasmic reticulum (ER) stress in the epithelium and dramatically inhibits epithelial proliferation and differentiation during lactation. The synthesis of milk and its major components, α/ß-casein and whey acidic protein (WAP), is significantly reduced due to decreased prolactin receptor (Prlr) and ErbB4 expression in Xbp1-deficient mammary epithelium. Reduction of Prlr and ErbB4 expression and their diminished availability at the cell surface lead to reduced phosphorylated Stat5, an essential regulator of cell proliferation and differentiation during lactation. As a result, lactating mammary glands in these mice produce less milk protein, leading to poor pup growth and postnatal death. These findings suggest that the loss of Xbp1 induces a terminal UPR which blocks proliferation and differentiation during mammary gland development.
Asunto(s)
Proliferación Celular , Proteínas de Unión al ADN/metabolismo , Lactancia , Glándulas Mamarias Animales/citología , Glándulas Mamarias Animales/crecimiento & desarrollo , Factores de Transcripción/metabolismo , Animales , Diferenciación Celular , Células Cultivadas , Proteínas de Unión al ADN/análisis , Proteínas de Unión al ADN/genética , Células Epiteliales/citología , Células Epiteliales/metabolismo , Femenino , Eliminación de Gen , Glándulas Mamarias Animales/ultraestructura , Ratones , Ratones Endogámicos C57BL , Prolactina/análisis , Prolactina/metabolismo , Receptor ErbB-4/análisis , Receptor ErbB-4/metabolismo , Factores de Transcripción del Factor Regulador X , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/metabolismo , Factores de Transcripción/análisis , Factores de Transcripción/genética , Respuesta de Proteína Desplegada , Proteína 1 de Unión a la X-BoxRESUMEN
T helper (TH)-cell subsets, such as TH1 and TH17, mediate inflammation in both peripheral tissues and central nervous system. Here we show that STAT5 is required for T helper-cell pathogenicity in autoimmune neuroinflammation but not in experimental colitis. Although STAT5 promotes regulatory T cell generation and immune suppression, loss of STAT5 in CD4+ T cells resulted in diminished development of experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis. Our results showed that loss of encephalitogenic activity of STAT5-deficient autoreactive CD4+ T cells was independent of IFN-γ or interleukin 17 (IL-17) production, but was due to the impaired expression of granulocyte-macrophage colony-stimulating factor (GM-CSF), a crucial mediator of T-cell pathogenicity. We further showed that IL-7-activated STAT5 promotes the generation of GM-CSF-producing CD4+ T cells, which were preferentially able to induce more severe EAE than TH17 or TH1 cells. Consistent with GM-CSF-producing cells being a distinct subset of TH cells, the differentiation program of these cells was distinct from that of TH17 or TH1 cells. We further found that IL-3 was secreted in a similar pattern as GM-CSF in this subset of TH cells. In conclusion, the IL-7-STAT5 axis promotes the generation of GM-CSF/IL-3-producing TH cells. These cells display a distinct transcriptional profile and may represent a novel subset of T helper cells which we designate as TH-GM.
Asunto(s)
Encefalomielitis Autoinmune Experimental/inmunología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/inmunología , Factor de Transcripción STAT5/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Animales , Colitis/genética , Colitis/inmunología , Colitis/patología , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/patología , Eliminación de Gen , Factor Estimulante de Colonias de Granulocitos y Macrófagos/análisis , Interleucina-7/análisis , Interleucina-7/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/genética , Linfocitos T Colaboradores-Inductores/metabolismo , Linfocitos T Colaboradores-Inductores/patologíaRESUMEN
BACKGROUND: Genetic variations associated with interstitial lung diseases (ILD) have not been extensively studied in Japanese infants. METHODS: Forty-three infants with unexplained lung dysfunction were studied. All 43, 22, and 17 infants underwent analyses of surfactant protein (SP)-C gene (SFTPC) and ATP-binding cassette A3 gene (ABCA3), SP-B gene (SFTPB), and SP-B western blotting, respectively. Two and four underwent assessment of granulocyte macrophage colony-stimulating factor-stimulating phosphorylation of signal transducer and activator of transcription-5 (pSTAT-5) and analyses of FOXF1 gene (FOXF1), respectively. RESULTS: ILD were diagnosed clinically in nine infants: four, three, and two had interstitial pneumonitis, hereditary pulmonary alveolar proteinosis (hPAP), and alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV), respectively. Genetic variations considered responsible were detected in six (67%) of the nine infants with ILD: three with hPAP (SFTPC p.Leu45Arg and p.Gln145fs, and ABCA3 p.Arg1583Trp/p.Val1495CysfsX21), two with interstitial pneumonitis (SFTPC p.Lys63Glu and p.Ser72Asn/p.Gly100Ala), and one with ACD/MPV (FOXF1 p.Leu300ArgfsX79). None showed SFTPB mutations or defects in pSTAT-5. The 17 bronchoalveolar lavage or tracheal aspirates contained enough SP-B protein. CONCLUSION: The SP-C abnormality was most prevalent, and SP-B deficiency was rare in Japanese infants with hereditary ILD.
Asunto(s)
Variación Genética , Enfermedades Pulmonares Intersticiales/genética , Transportadoras de Casetes de Unión a ATP/genética , Edad de Inicio , Pueblo Asiatico/genética , Femenino , Factores de Transcripción Forkhead/genética , Marcadores Genéticos , Predisposición Genética a la Enfermedad , Herencia , Humanos , Lactante , Recién Nacido , Japón/epidemiología , Leucocitos Mononucleares/química , Enfermedades Pulmonares Intersticiales/diagnóstico , Enfermedades Pulmonares Intersticiales/etnología , Enfermedades Pulmonares Intersticiales/fisiopatología , Enfermedades Pulmonares Intersticiales/terapia , Masculino , Fenotipo , Fosforilación , Proteinosis Alveolar Pulmonar/congénito , Proteinosis Alveolar Pulmonar/etnología , Proteinosis Alveolar Pulmonar/genética , Proteína B Asociada a Surfactante Pulmonar/análisis , Proteína B Asociada a Surfactante Pulmonar/deficiencia , Proteína B Asociada a Surfactante Pulmonar/genética , Proteína C Asociada a Surfactante Pulmonar/genética , Sistema de Registros , Factor de Transcripción STAT5/análisisRESUMEN
OBJECTIVE: To examine the relationship of defects in interleukin (IL)-7-induced naive CD4 T-cell homeostasis with residual immune activation and CD4 T-cell senescence in HIV patients receiving antiretroviral therapy (ART) who exhibit persistent CD4 T-cell deficiency. DESIGN: IL-7 induced proliferation of, and IL-7 receptor signalling in, total and naive CD4 T cells of HIV patients who had low (<350 cells/µl) or normal (>500 cells/µl) CD4 T-cell counts on ART was examined and related to markers of CD4 T-cell activation and senescence and innate immune activation. METHODS: Total, naive (CD45RA CD27) and CD31 naive CD4 T cells from aviremic HIV patients (n=39) with nadir CD4 T-cell counts less than 100 cells/µl, who had received ART for a median time of 7 (range 1-11) years, were assessed for CD127 expression, proliferation (Ki67), signal transducer and activator of transcription 5 phosphorylation (pSTAT5) and CD127 modulation following IL-7 stimulation. Changes were related to proportions of CD4 T cells expressing HLA-DR or CD57 and plasma levels of sCD14, CXCL9 and CXCL10. RESULTS: Patients with CD4 T-cell deficiency exhibited lower expression of CD127 on total, naive and CD31 naive CD4 T cells. Downregulation of CD127 after culture with IL-7 correlated inversely with CD4 T-cell counts and directly with Ki67 expression. Induction of pSTAT5 in CD4 T-cell subsets was greater in patients with normal CD4 T-cell counts. CD127 expression correlated inversely with proportions of CD4CD57 T cells, and pSTAT5 induction correlated inversely with CD4 T-cell expression of HLA-DR and CD57. CONCLUSION: Defects of IL-7 signalling in HIV patients with persistent CD4 T-cell deficiency receiving ART are associated with CD4 T-cell activation and senescence.
Asunto(s)
Antirretrovirales/uso terapéutico , Linfocitos T CD4-Positivos/inmunología , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/inmunología , Interleucina-7/inmunología , Activación de Linfocitos , Transducción de Señal , Adulto , Anciano , Antígenos CD/análisis , Proliferación Celular , Femenino , Humanos , Inmunofenotipificación , Antígeno Ki-67/análisis , Masculino , Persona de Mediana Edad , Factor de Transcripción STAT5/análisisAsunto(s)
Cromosomas Humanos Par 8/ultraestructura , Cromosomas Humanos Par 9/ultraestructura , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Sistema de Señalización de MAP Quinasas , Proteína Quinasa 1 Activada por Mitógenos/fisiología , Proteína Quinasa 3 Activada por Mitógenos/fisiología , Proteínas de Fusión Oncogénica/fisiología , Translocación Genética , Adulto , Cromosomas Humanos Par 8/genética , Cromosomas Humanos Par 9/genética , Eritropoyesis , Células Madre Hematopoyéticas/patología , Humanos , Cariotipificación , Leucemia Mielógena Crónica BCR-ABL Positiva/sangre , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Masculino , Células Neoplásicas Circulantes , Células Madre Neoplásicas/patología , Proteínas de Fusión Oncogénica/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Transcripción STAT3/análisis , Factor de Transcripción STAT5/análisisRESUMEN
Nuclear localization of signal transducer and activator of transcription (Stat) 5 marks good prognosis in estrogen receptor/progesterone receptor-positive breast tumors. This positive characteristic is counteracted by studies in laboratory animals demonstrating that deregulated Stat5 activity may convert proper mammary development into a latent oncogenic process. Tumorigenesis is initiated during the parity cycles, most probably during pregnancy, when the activated Stat5 antagonizes or manipulates parity's protective mechanisms. For example, it can alter the differentiation/proliferation balance, induce growth hormone signaling, cause specific alteration in chromatin structure, inhibit tumor-suppressor activity and induce DNA damage that counteracts the enhanced DNA-damage response exerted by parity. Palpable tumors develop after a latent period from individual cells. This happens in the estropausal period in transgenic mice maintaining deregulated Stat5 activity in the mammary gland, or during involution, months after transplantation of transfected cells with constitutively active Stat5. Candidate vulnerable cells are those which maintain high nuclear Stat5 activity. Due to the hazardous outcome of deregulated Stat5 activity in these cells, such as induced DNA damage or high cyclin D1 activity, the gland is prone to transformation. The developing tumors are mostly adenocarcinomas or their subtypes. They are estrogen receptor-positive and maintain a specific Stat5 gene signature that allows tracking their inducer. From a clinical point of view, deregulated Stat5 activity represents a genuine risk factor for breast cancer. Monitoring Stat5 activity during vulnerable periods and developing specific tools for its suppression in breast epithelial cells could potentially limit new incidence of the disease.
Asunto(s)
Neoplasias de la Mama/etiología , Factor de Transcripción STAT5/fisiología , Animales , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Diferenciación Celular , Transformación Celular Neoplásica , Femenino , Humanos , Paridad , Embarazo , Pronóstico , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/antagonistas & inhibidoresRESUMEN
The objective of this study was to characterize the circulating concentrations of insulin-like growth factor-I (IGF-I) and the hepatic expression of key genes regulating the somatotropic axis in cows divergent in genetic merit for fertility traits but with similar genetic merit for milk production traits. A total of 11 cows with good genetic merit for fertility (Fert+) and 12 cows with poor genetic merit for fertility (Fert-) underwent liver biopsy by percutaneous punch technique on d 20 (±6.7 d) prepartum and on d 2 (±1.5 d), d 58 (±3.7 d), d 145 (±13 d), and d 245 (±17.1 d) postpartum. Total RNA was isolated and the mRNA expression of growth hormone receptor (GHR 1A and GHRtot), IGF-I, janus tyrosine kinase 2 (JAK2), signal transducer and activator of transcription 5B (STAT5B), suppressor of cytokine signaling 3 (SOCS-3), acid-labile subunit (ALS), and IGF-binding proteins (IGFBP1 to IGFBP6) were measured by real-time quantitative PCR. During lactation, the circulating concentrations of IGF-I were 34% greater in Fert+ cows. The Fert+ cows had increased mean expression of IGF-I mRNA during the study; however, the difference in IGF-I mRNA abundance between Fert+ and Fert- cows was most pronounced at d 145 and 245. The expression of IGFBP3 and ALS transcript was similar in Fert+ and Fert- cows for the duration of the study. The Fert- cows, however, had greater expression of IGFBP2, IGFBP4, IGFBP5, and IGFBP6. Genotype had no effect on mRNA abundance of GHR 1A, STAT5B, JAK2, or SOCS-3. Genetic merit for fertility traits affects hepatic expression of key genes of the somatotropic axis regulating the synthesis, bioavailability, and stability of circulating IGF-I.
Asunto(s)
Bovinos/genética , Fertilidad/genética , Lactancia/genética , Hígado/metabolismo , Preñez/genética , Carácter Cuantitativo Heredable , Animales , Proteínas Portadoras/análisis , Proteínas Portadoras/genética , Bovinos/fisiología , Femenino , Fertilidad/fisiología , Genes/genética , Genes/fisiología , Glicoproteínas/análisis , Glicoproteínas/genética , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/análisis , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/análisis , Janus Quinasa 2/análisis , Janus Quinasa 2/genética , Lactancia/fisiología , Hígado/química , Embarazo , Preñez/fisiología , Receptores de Somatotropina/análisis , Receptores de Somatotropina/genética , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/genética , Proteínas Supresoras de la Señalización de Citocinas/análisis , Proteínas Supresoras de la Señalización de Citocinas/genéticaRESUMEN
Hyperphagia during pregnancy, despite rising concentrations of the satiety hormone leptin, suggests that a state of leptin resistance develops. This study investigated the satiety response and hypothalamic responses to leptin during pregnancy in the mouse. Pregnant (day 13) and nonpregnant mice received an i.p. injection of either leptin or vehicle and then 24-h food intake was measured. Further groups of pregnant and nonpregnant mice were perfused 2 h after leptin or vehicle injections and brains were processed for pSTAT3 and pSTAT5 immunohistochemistry. Leptin treatment significantly decreased food intake in nonpregnant mice. In pregnant mice, however, leptin treatment did not suppress food intake, indicating a state of leptin resistance. In the arcuate nucleus, leptin treatment increased the number of cells positive for pSTAT3, a marker of leptin activity, to a similar degree in both nonpregnant and pregnant mice. In the ventromedial nucleus (VMN), the leptin-induced increase in pSTAT3-positive cell number was significantly reduced in pregnant mice compared to that in nonpregnant mice. In nonpregnant mice, leptin treatment had no effect on the number of pSTAT5-positive cells, suggesting that in this animal model, leptin does not act through STAT5. In pregnant mice, basal levels of pSTAT5 were higher than in nonpregnant mice, and leptin treatment led to a decrease in the number of pSTAT5-positive cells in the hypothalamus. Overall, these results demonstrate that during pregnancy in the mouse, a state of leptin resistance develops, and this is associated with a reduced sensitivity of the VMN to leptin.
Asunto(s)
Ingestión de Alimentos/efectos de los fármacos , Hipotálamo/efectos de los fármacos , Leptina/farmacología , Preñez/fisiología , Factor de Transcripción STAT5/metabolismo , Transducción de Señal/efectos de los fármacos , Animales , Núcleo Arqueado del Hipotálamo/química , Núcleo Arqueado del Hipotálamo/efectos de los fármacos , Resistencia a Medicamentos , Femenino , Hipotálamo/química , Hipotálamo/fisiología , Ratones , Ratones Endogámicos C57BL , Fosforilación/efectos de los fármacos , Embarazo , Factor de Transcripción STAT3/análisis , Factor de Transcripción STAT3/efectos de los fármacos , Factor de Transcripción STAT3/metabolismo , Factor de Transcripción STAT5/análisis , Factor de Transcripción STAT5/efectos de los fármacos , Saciedad/efectos de los fármacos , Núcleo Hipotalámico Ventromedial/química , Núcleo Hipotalámico Ventromedial/efectos de los fármacosRESUMEN
BACKGROUND: Because cell signaling and cell metabolic pathways are executed through proteins, protein signatures in primary tumors are useful for identifying key nodes in signaling networks whose alteration is associated with malignancy and/or clinical outcomes. This study aimed to determine protein signatures in primary lung cancer tissues. METHODOLOGY/ PRINCIPAL FINDINGS: We analyzed 126 proteins and/or protein phosphorylation sites in case-matched normal and tumor samples from 101 lung cancer patients with reverse-phase protein array (RPPA) assay. The results showed that 18 molecules were significantly different (p<0.05) by at least 30% between normal and tumor tissues. Most of those molecules play roles in cell proliferation, DNA repair, signal transduction and lipid metabolism, or function as cell surface/matrix proteins. We also validated RPPA results by Western blot and/or immunohistochemical analyses for some of those molecules. Statistical analyses showed that Ku80 levels were significantly higher in tumors of nonsmokers than in those of smokers. Cyclin B1 levels were significantly overexpressed in poorly differentiated tumors while Cox2 levels were significantly overexpressed in neuroendocrinal tumors. A high level of Stat5 is associated with favorable survival outcome for patients treated with surgery. CONCLUSIONS/ SIGNIFICANCE: Our results revealed that some molecules involved in DNA damage/repair, signal transductions, lipid metabolism, and cell proliferation were drastically aberrant in lung cancer tissues, and Stat5 may serve a molecular marker for prognosis of lung cancers.