RESUMO
Some endocrine organs are frequent targets of autoimmune attack. Here, we addressed the origin of autoimmune disease from the viewpoint of feedback control. Endocrine tissues maintain mass through feedback loops that balance cell proliferation and removal according to hormone-driven regulatory signals. We hypothesized the existence of a dedicated mechanism that detects and removes mutant cells that missense the signal and therefore hyperproliferate and hypersecrete with potential to disrupt organismal homeostasis. In this mechanism, hypersecreting cells are preferentially eliminated by autoreactive T cells at the cost of a fragility to autoimmune disease. The "autoimmune surveillance of hypersecreting mutants" (ASHM) hypothesis predicts the presence of autoreactive T cells in healthy individuals and the nature of self-antigens as peptides from hormone secretion pathway. It explains why some tissues get prevalent autoimmune disease, whereas others do not and instead show prevalent mutant-expansion disease (e.g., hyperparathyroidism). The ASHM hypothesis is testable, and we discuss experimental follow-up.
Assuntos
Doenças Autoimunes/imunologia , Diabetes Mellitus Tipo 1/imunologia , Glândulas Endócrinas/imunologia , Sistema Endócrino/imunologia , Vigilância Imunológica/imunologia , Animais , Doenças Autoimunes/genética , Doenças Autoimunes/patologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/patologia , Glândulas Endócrinas/citologia , Glândulas Endócrinas/metabolismo , Sistema Endócrino/citologia , Sistema Endócrino/metabolismo , Feminino , Humanos , Vigilância Imunológica/genética , Masculino , Mutação , Linfócitos T/citologia , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Peribiliary glands (PBGs) are accessory glands with mucinous and serous acini in the biliary tree. The PBG is composed of a heterogeneous cell population, such as mucus- and pancreatic enzyme-producing epithelial cells, whereas it constitutes niches for multipotential stem/progenitor cells in the human extrahepatic bile duct (EHBD). By contrast, the nature of PBGs in the mouse EHBD remains unclear. Our aim was to establish a method for isolating and characterizing PBG-constituting cells in the mouse EHBD. We found that trophoblast cell surface protein 2 (Trop2) was expressed in the luminal epithelium of mouse EHBD exclusively, but not in the PBG. On the basis of the differential expression profile of Trop2, lumen-forming biliary epithelial cells (LBECs) and PBG-constituting epithelial cells (PBECs) were separately isolated for further characterization. Gene expression analysis revealed that the isolated mouse PBECs expressed several marker genes related to human PBGs. In the colony formation assay, PBECs showed significantly higher colony formation capacity than LBECs. In the organoid formation assay, PBECs formed cystic organoid with LBEC-like phenotype. Interestingly, PBECs proliferated, accompanied by reexpression of Trop2 in vivo after bile duct ligation. Furthermore, the unique expression profile of Trop2 was conserved in human EHBD. Our findings indicate that Trop2 is a useful marker in investigating the pathophysiological roles and characteristics of mouse and human PBGs in biliary diseases.
Assuntos
Antígenos de Neoplasias/metabolismo , Ductos Biliares Extra-Hepáticos/citologia , Moléculas de Adesão Celular/metabolismo , Glândulas Endócrinas/citologia , Células-Tronco/citologia , Animais , Ductos Biliares Extra-Hepáticos/metabolismo , Neoplasias do Sistema Biliar/metabolismo , Neoplasias do Sistema Biliar/patologia , Proliferação de Células , Células Cultivadas , Glândulas Endócrinas/metabolismo , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Fenótipo , Células-Tronco/metabolismoAssuntos
Células-Tronco Adultas/citologia , Glândulas Endócrinas/citologia , Glândulas Endócrinas/fisiologia , Doenças do Sistema Endócrino/terapia , Animais , Biomarcadores/metabolismo , Diferenciação Celular , Transplante de Células , Reprogramação Celular , Feminino , Hormônios/metabolismo , Humanos , Masculino , Neoplasias/terapia , Cultura Primária de Células , Fatores SexuaisRESUMO
The Esophageal Cancer Related Gene 4 (ECRG4) is a highly conserved tumour suppressor gene encoding various peptides (augurin, CΔ16 augurin, ecilin, argilin, CΔ16 argilin) which can be processed and secreted. In the present work, we examined ECRG4 expression and location in a wide range of rat organs and reviewed the available literature. ECRG4 mRNA was identified in all examined tissues by quantitative PCR (qPCR). ECRG4 immunoreaction was mainly cytoplasmic, and was detected in heart and skeletal muscles, smooth muscle cells showing only weak reactions. In the digestive system, ECRG4 immunostaining was stronger in the esophageal epithelium, bases of gastric glands, hepatocytes and pancreatic acinar epithelium. In the lymphatic system, immunoreactive cells were detectable in the thymus cortex, lymph node medulla and splenic red pulp. In the central and peripheral nervous systems, different neuronal groups showed different reaction intensities. In the endocrine system, ECRG4 immunoreaction was detected in the hypothalamic paraventricular and supraoptic nuclei, hypophysis, thyroid and parathyroid glands, adrenal zona glomerularis and medulla and Leydig cells, as well as in follicular and luteal cells of the ovary. In the literature, ECRG4 has been reported to inhibit cell proliferation and increase apoptosis in various cell types. It is down-regulated, frequently due to hypermethylation, in esophageal, prostate, breast and colon cancers, together with glioma (oncosuppressor function), although it is up-regulated in papillary thyroid cancer (oncogenic role). ECRG4 expression is also higher in non-proliferating cells of the lymphatic system. In conclusion, our identification of ECRG4 in many structures suggests the involvement of ECRG4 in the tumorigenesis of other organs and also the need for further research. In addition, on the basis of the location of ECRG4 in neurons and endocrine cells and the fact that it can be secreted, its role as a neurotransmitter/neuromodulator and endocrine factor must be examined in depth in the future.
Assuntos
Regulação Neoplásica da Expressão Gênica/genética , Proteínas Supressoras de Tumor/biossíntese , Proteínas Supressoras de Tumor/genética , Sequência de Aminoácidos , Animais , Glândulas Endócrinas/citologia , Glândulas Endócrinas/metabolismo , Feminino , Imuno-Histoquímica , Masculino , Dados de Sequência Molecular , Neurônios/metabolismo , Reação em Cadeia da Polimerase , RNA Mensageiro/biossíntese , Ratos , Ratos Wistar , Distribuição TecidualRESUMO
Cancer stem-like cells are a subpopulation of self-renewing cells that are more resistant to chemotherapy and radiation therapy than the other surrounding cancer cells. The cancer stem cell model predicts that only a subset of cancer cells possess the ability to self-renew and produce progenitor cells that can reconstitute and sustain tumor growth. Evidence supporting the existence of cancer stem-like cells in the thyroid, pituitary, and in other endocrine tissues is rapidly accumulating. These cells have been studied using specific biomarkers including: CD133, CD44, Nestin, Nanog, and aldehyde dehydrogenase enzyme. Putative cancer stem-like cells can be studied in vitro using serum-free media supplemented with basic fibroblast growth factor and epidermal growth factor grown in low attachment plates or in extracellular matrix leading to sphere formation in vitro. Cancer stem-like cells can also be separated by fluorescent cell sorting and used for in vitro or in vivo studies. Injection of enriched populations of cancer stem-like cells (also referred to as tumor initiating cells) into immunodeficient mice results in growth of xenografts which express cancer stem-like biomarkers. Human cancer stem-like cells have been identified in thyroid cancer cell lines, in primary thyroid cancers, in normal pituitary, and in pituitary tumors. Other recent studies suggest the existence of stem cells and cancer stem-like cells in endocrine tumors of the gastrointestinal tract, pancreas, lungs, adrenal, parathyroid, and skin. New discoveries in this field may lead to more effective therapies for highly aggressive and lethal endocrine cancers.
Assuntos
Glândulas Endócrinas/citologia , Células-Tronco Neoplásicas/fisiologia , Células-Tronco/fisiologia , Animais , Trato Gastrointestinal/citologia , Humanos , Células Neuroendócrinas/fisiologia , Pâncreas/citologia , Hipófise/citologia , Glândula Tireoide/citologiaRESUMO
BACKGROUND & AIMS: Although regeneration of intrahepatic bile ducts has been extensively studied and intrahepatic progenitor cells have been identified, few studies have focussed on the extrahepatic bile duct (EHBD). We hypothesized that local progenitor cells are present within the EHBD of humans. Human EHBD specimens (n = 17) were included in this study. METHODS: Specimens of normal EHBD tissue were obtained from healthy donor livers (n = 6), mildly injured EHBD from patients with cholangitis (n = 6) and severely injured EHBD from patients with ischaemic type biliary lesions (n = 5). Double immunostaining for K19 and the proliferation marker Ki-67 was performed to identify and localize proliferating cells. In addition, immunofluorescent doublestaining using antibodies against K19 and c-Kit was performed to identify and localize cholangiocytes co-expressing putative progenitor cell markers. RESULTS: In normal EHBD, few Ki-67(+) cells were detected, whereas large numbers of Ki-67(+) were found in the diseased EHBD. In EHBD affected by cholangitis, Ki-67(+) cells were mainly located in the basal layer of the lumen. EHBD specimens from patients with ischaemic type biliary lesions displayed histological signs of epithelial cell loss and large numbers of Ki-67(+) cells were observed in the peribiliary glands. C-Kit expression was localized throughout the EHBD wall and immunofluorescent doublestaining identified a few K19(+) /c-Kit(+) cells in the luminal epithelium of the EHBD as well as in the peribiliary glands. CONCLUSIONS: These findings support the hypothesis that progenitor cells exist in the EHBD and that the peribiliary glands can be considered a local progenitor cell niche in the human EHBD.
Assuntos
Ductos Biliares Extra-Hepáticos/citologia , Glândulas Endócrinas/citologia , Epitélio/fisiologia , Regeneração/fisiologia , Células-Tronco/citologia , Ductos Biliares Extra-Hepáticos/patologia , Imunofluorescência , Humanos , Imuno-Histoquímica , Queratina-7 , Antígeno Ki-67 , Proteínas Proto-Oncogênicas c-kit/metabolismoRESUMO
FGF21 is a novel metabolic regulator involved in the control of glucose homeostasis, insulin sensitivity, and ketogenesis. The liver has been considered the main site of production and release of FGF21 into the blood. Here, we show that, after thermogenic activation, brown adipose tissue becomes a source of systemic FGF21. This is due to a powerful cAMP-mediated pathway of regulation of FGF21 gene transcription. Norepinephrine, acting via ß-adrenergic, cAMP-mediated, mechanisms and subsequent activation of protein kinase A and p38 MAPK, induces FGF21 gene transcription and also FGF21 release in brown adipocytes. ATF2 binding to the FGF21 gene promoter mediates cAMP-dependent induction of FGF21 gene transcription. FGF21 release by brown fat in vivo was assessed directly by analyzing arteriovenous differences in FGF21 concentration across interscapular brown fat, in combination with blood flow to brown adipose tissue and assessment of FGF21 half-life. This analysis demonstrates that exposure of rats to cold induced a marked release of FGF21 by brown fat in vivo, in association with a reduction in systemic FGF21 half-life. The present findings lead to the recognition of a novel pathway of regulation the FGF21 gene and an endocrine role of brown fat, as a source of FGF21 that may be especially relevant in conditions of activation of thermogenic activity.
Assuntos
Tecido Adiposo Marrom/metabolismo , Glândulas Endócrinas/metabolismo , Fatores de Crescimento de Fibroblastos/biossíntese , Regulação da Expressão Gênica/fisiologia , Termogênese/fisiologia , Tecido Adiposo Marrom/citologia , Animais , Células Cultivadas , Temperatura Baixa , AMP Cíclico/genética , AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/genética , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Glândulas Endócrinas/citologia , Ativação Enzimática/fisiologia , Fatores de Crescimento de Fibroblastos/genética , Masculino , Camundongos , Camundongos Mutantes , Ratos , Ratos Wistar , Transcrição Gênica/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
We report the first demonstration of the embryonal patch patterns of endocrine organs and the polyclonality of hormone-producing cell populations using chimeric mice produced by aggregation of C57BL/6-Tg(CAG-EGFP)C14-Y01-FM131Osb transgenic mice and BALB/C mice. Confocal laser scanning microscopy (CLSM) analysis for enhanced green fluorescent protein (EGFP) and immunohistochemistry with anti-EGFP antibody revealed that all endocrine organs of chimeric mice had a mosaic appearance of EGFP-positive patches and EGFP-negative patches. The patches composed of EGFP-positive cells were distinctive in their size and shape. The pituitary patches were large and irregular, representing a geographical pattern. In contrast, parathyroid, pancreatic islet, and adrenal medulla patches were small and demarcated, representing an island-like pattern. Thyroid follicles and adrenal cortex cords showed a mixture of monophenotypia and polyphenotypia, indicating polyclonal embryonic origin. Furthermore, we studied the tissue clonality of hormone-producing cell populations in the pituitary, thyroid, and pancreatic islets using a combination method of CLSM for EGFP and immunohistochemistry for hormones. All the pituitary cell populations of GH, prolactin, TSH, FSH, LH, and ACTH, the calcitonin-producing cell population in the thyroid, and the insulin- and glucagon-producing cell populations in pancreatic islets had mosaic patterns in EGFP expression in the chimeric mice, suggesting polyclonal embryonic origin. In conclusion, the different patch patterns of the endocrine organs could contribute to the understanding of embryonic development and organization of endocrine organs. Furthermore, we clearly demonstrate that all hormone-producing cell populations are of polyclonal embryonic origin, derived from more than two progenitor cells.
Assuntos
Quimera/genética , Glândulas Endócrinas/embriologia , Glândulas Endócrinas/metabolismo , Proteínas de Fluorescência Verde/genética , Animais , Quimera/metabolismo , Glândulas Endócrinas/citologia , Feminino , Proteínas de Fluorescência Verde/metabolismo , Imuno-Histoquímica , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/embriologia , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microscopia Confocal , Modelos Biológicos , Hipófise/citologia , Hipófise/embriologia , Hipófise/metabolismo , Gravidez , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Glândula Tireoide/citologia , Glândula Tireoide/embriologia , Glândula Tireoide/metabolismoRESUMO
Detection of differentiation in general and adipogenesis specifically is conventionally practised by taking only the few cells into account which are visible in the field of view provided by optical microscopy using high-resolution objectives. Other methods of quantification of adipogenic differentiation such as real time PCR, measurement of glycerophosphate dehydrogenase activity or adipogenesis assays only provide integral information lacking spatial resolution and information on the fraction of differentiated cells. Here we used high-resolution scanning and automated image processing to automatically analyze and quantify cell numbers in the range of 20,000. For optimisation of the approach, human gland stem cells (GSC) were differentiated to the adipogenic phenotype comprising inclusion of lipid vesicles. Oil red O and 4',6'-diamidino-2-phenylindole (DAPI) staining made it possible to derive the number of differentiated cells in relation to the total number of cells. For evaluation of the image processing software we verified our results using adipogenesis assay and phase contrast based cell counting. We developed a method of determining differentiation efficiencies covering the range from 10% down to 100ppm with the same image processing and an identical set of parameters, matching the results of the adipogenesis assay. Our approach is based on a statistically significant number of cells and shows high sensitivity taking into account the heterogeneous differentiation pattern of adipogenesis in GSC and other stem cells.
Assuntos
Adipócitos/citologia , Adipogenia/fisiologia , Diferenciação Celular/fisiologia , Glândulas Endócrinas/citologia , Células-Tronco/citologia , Células 3T3/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/ultraestrutura , Adulto , Animais , Automação , Diferenciação Celular/efeitos dos fármacos , Dexametasona/farmacologia , Glândulas Endócrinas/efeitos dos fármacos , Glândulas Endócrinas/ultraestrutura , Humanos , Insulina/farmacologia , Masculino , Camundongos , Pessoa de Meia-Idade , Transplante de Células-Tronco , Células-Tronco/efeitos dos fármacos , Células-Tronco/ultraestrutura , Glândula Submandibular/citologiaRESUMO
Aquaporins (AQPs), a family of water channels expressed in epithelial cells, function to transport water in a bidirectional manner to facilitate transepithelial fluid absorption and secretion. Additionally, AQP1 and AQP5 are found in pancreatic zymogen granules and synaptic vesicles and are involved in vesicle swelling and exocytosis in exocrine cells and neurons. Here, we show AQP1 is in dense-core secretory granule (DCSG) membranes of endocrine tissue: pituitary and adrenal medulla. The need for AQP1 in endocrine cell function was examined by stable transfection of AQP1 antisense RNA into AtT20 cells, a pituitary cell line, to down-regulate AQP1 expression. These AQP1-deficient cells showed more than 60% depletion of DCSGs and significantly decreased DCSG protein levels, including proopiomelanocotin/pro-ATCH and prohormone convertase 1/3, but not non-DCSG proteins. Pulse-chase studies revealed that whereas DCSG protein synthesis was unaffected, approximately 50% of the newly synthesized proopiomelanocortin was degraded within 1 h. Low levels of ACTH were released upon stimulation, indicating that the small number of DCSGs that were made in the presence of the residual AQP1 were functionally competent for exocytosis. Analysis of anterior pituitaries from AQP1 knockout mice showed reduced prohormone convertase 1/3, carboxypeptidase E, and ACTH levels compared to wild-type mice demonstrating that our results observed in AtT20 cells can be extended to the animal model. Thus, AQP1 is important for maintaining DCSG biogenesis and normal levels of hormone secretion in pituitary endocrine cells.
Assuntos
Aquaporina 1/metabolismo , Glândulas Endócrinas/citologia , Vesículas Secretórias/metabolismo , Hormônio Adrenocorticotrópico/metabolismo , Animais , Aquaporina 1/genética , Linhagem Celular , Células Clonais , Regulação para Baixo , Glândulas Endócrinas/metabolismo , Glândulas Endócrinas/ultraestrutura , Camundongos , Camundongos Knockout , Adeno-Hipófise/metabolismo , Pró-Opiomelanocortina/biossíntese , Pró-Opiomelanocortina/genética , Biossíntese de Proteínas , Processamento de Proteína Pós-Traducional , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Vesículas Secretórias/ultraestrutura , Transfecção , Regulação para CimaRESUMO
We previously presented evidence that proliferative human islet precursor cells may be derived in vitro from adult islets by epithelial-to-mesenchymal transition (EMT) and show here that similar fibroblast-like cells can be derived from mouse islets. These mouse cell populations exhibited changes in gene expression consistent with EMT. Both C-peptide and insulin mRNAs were undetectable in expanded cultures of mouse islet-derived precursor cells (mIPCs). After expansion, mIPCs could be induced to migrate into clusters and differentiate into hormone-expressing islet-like aggregates. Although early morphological changes suggesting EMT were observed by time-lapse microscopy when green fluorescent protein-labeled beta cells were placed in culture, the expanded precursor cell population was not fluorescent. Using two mouse models in which beta cells were permanently made either to express alkaline phosphatase or to have a deleted M(3) muscarinic receptor, we provide evidence that mIPCs in long term culture are not derived from beta cells.
Assuntos
Glândulas Endócrinas/citologia , Células Epiteliais/citologia , Células Secretoras de Insulina/citologia , Mesoderma/citologia , Células-Tronco/citologia , Animais , Diferenciação Celular , DNA/metabolismo , Regulação da Expressão Gênica , Genoma , Camundongos , Camundongos Knockout , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor Muscarínico M3/deficiência , Receptor Muscarínico M3/genéticaRESUMO
To identify cell populations directly responsive to prolactin (PRL), GH, erythropoietin, or granulocyte-colony stimulating factor within the physiological setting of an intact mammal, we combined in situ detection of hormone-activated signal transducer and activator of transcription (Stat)-5 in rats with high-throughput tissue array analysis using cutting-edge matrix assembly (CEMA). Inducible activation of Stat5a/b, as judged by levels of nuclear-localized, phosphoTyr694/699-Stat5a/b, served as an immediate and sensitive in situ marker of receptor signaling in rat tissues after injection into male and female rats of a single, receptor-saturating dose of hormone for maximal receptor activation. CEMA tissue arrays facilitated analysis of most tissues, including architecturally complex, thin-walled, and stratified tissues such as gut and skin. In 40 tissues analyzed, 35 PRL-responsive and 32 GH-responsive cell types were detected, of which 22 cell types were responsive to both hormones. Interestingly, PRL but not GH activated Stat5 in nearly all of the endocrine glands. In mammary glands, PRL activated Stat5 in a majority of luminal epithelial cells but not myoepithelial cells, stromal fibroblasts, or adipocytes, whereas GH activated Stat5 in a significant fraction of myoepithelial cells, fibroblasts, and adipocytes but only in a minority of luminal cells. Finally, the organism-wide screening revealed a yet-to-be identified erythropoietin-responsive cell type in connective tissue. CEMA tissue arrays provide cost-effective in situ analysis of large numbers of tissues. Biomarker-based identification of cell populations responsive to individual hormones may shed new light on endocrine disease as well as improve understanding of effects and side effects of hormones and drugs.
Assuntos
Eritropoetina/farmacologia , Expressão Gênica/efeitos dos fármacos , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Hormônio do Crescimento/farmacologia , Prolactina/farmacologia , Análise Serial de Tecidos , Animais , Glândulas Endócrinas/citologia , Glândulas Endócrinas/efeitos dos fármacos , Glândulas Endócrinas/metabolismo , Feminino , Gônadas/citologia , Gônadas/efeitos dos fármacos , Gônadas/metabolismo , Hibridização In Situ , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Masculino , Especificidade de Órgãos/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Neuropeptides B and W (NPB and NPW) have been identified as endogenous ligands of the G protein-coupled receptors (GPR) 7 and 8, which in humans are expressed in the hypothalamus and probably involved in the regulation of energy homeostasis and feeding behavior. GPR8 is absent in the rat, where the GPR8-like receptor (GPR8-LR) has been described. Reverse transcription-polymerase chain reaction detected the expression of NPB, NPW, GPR7 and GPR8-LR mRNAs in the hypothalamus, anterior pituitary, thyroid and parathyroid glands, pancreatic islets, adrenal glands, ovary and testis of the rat. Immunocytochemistry demonstrated the presence of NPB and NPW immunoreactivities in these same glands. Radioimmune assay showed that the bolus intraperitoneal injection of 2 nmol/100 g NPB or NPW raised the plasma levels of parathyroid hormone, corticosterone and testosterone. NPB also increased the blood concentration of thyroxine, and NPW that of ACTH and estradiol. Taken together, these findings allow us to suggest that NPB and NPW play a role in the autocrine-paracrine functional regulation of the endocrine system in the rat.
Assuntos
Glândulas Endócrinas/metabolismo , Neuropeptídeos/metabolismo , Receptores de Neuropeptídeos/metabolismo , Animais , Glândulas Endócrinas/citologia , Imuno-Histoquímica , Masculino , RNA Mensageiro/metabolismo , Radioimunoensaio , Ratos , Ratos WistarRESUMO
Exfoliated endometrial cells are a common finding on Papanicolaou smears obtained during the menstrual period. They have a characteristic morphologic appearance, but whether they are of endometrial glandular, stromal, or mixed origin is still debated. Cervical ThinPrep (Cytyc, Marlboro, MA) slides obtained during menses from women younger than 40 years were immunostained for a stromal (CD10) and an epithelial (keratin proteins) marker. Endometrial cell clusters were scored as purely stromal (composed entirely of CD10+ cells), purely glandular (entirely keratin+ cells), or mixed stromal and glandular. Of the 59 endometrial cell clusters scored, 22 (37%) were purely stromal, 17 (29%) purely glandular, and 20 (34%) mixed stromal and glandular. Exfoliated endometrial cell clusters often are composed exclusively of stromal cells. Mixed stromal-glandular groups and entirely glandular groups also are common. The distinction between stromal and glandular cells is not always possible by conventional Papanicolaou staining but can be established by immunocytochemical study.
Assuntos
Endométrio/citologia , Teste de Papanicolaou , Células Estromais/citologia , Esfregaço Vaginal/métodos , Adulto , Glândulas Endócrinas/citologia , Glândulas Endócrinas/patologia , Endométrio/patologia , Feminino , Humanos , Queratinas/análise , Menstruação , Neprilisina/análise , Células Estromais/patologiaRESUMO
Synaptotagmins (syt) form a large family of transmembrane proteins and some of its isoforms are known to regulate calcium-induced membrane fusion during vesicular traffic. In view of the reported implication of the isoform syt8 in exocytosis we investigated the expression, localisation and calcium-sensitivity of syt8 in secretory cells. An immunopurified antipeptide antibody was generated which is directed against a C-terminal sequence and devoid of crossreactivity towards syt1 to 12. Subcellular fractionation and immunocytochemistry revealed two forms of synaptotagmin 8 (50 and 40 kDa). Whereas the 40-kDa was present in the cytosol in brain, in PC12 and in clonal beta-cells, the 50-kDa form was localised in very typical clusters and partially colocalised with the SNARE protein Vti1a. Moreover, in primary hippocampal neurons syt8 was only found within the soma. Amplification of syt8 by RT-PCR indicated that the observed protein variants were not generated by alternative splicing of the 6th exon and are most likely linked to variations in the N-terminal region. In contrast to the established calcium sensor syt2, endogenous cytosolic syt8 and transiently expressed syt8-C2AB-eGFP did not translocate upon a raise in cytosolic calcium in living cells. Syt8 is therefore not a calcium sensor in exocytotic membrane fusion in endocrine cells.
Assuntos
Cálcio/farmacologia , Membrana Celular/metabolismo , Glândulas Endócrinas/citologia , Regulação da Expressão Gênica , Neurônios/metabolismo , Sistemas Neurossecretores/citologia , Sinaptotagminas/metabolismo , Animais , Anticorpos/metabolismo , Especificidade de Anticorpos , Cálcio/metabolismo , Citosol/metabolismo , Glândulas Endócrinas/metabolismo , Éxons/genética , Neurônios/citologia , Sistemas Neurossecretores/metabolismo , Células PC12 , Ligação Proteica , Transporte Proteico , Proteínas Qb-SNARE/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Solubilidade , Sinaptotagminas/genéticaRESUMO
Tanycytes are bipolar cells bridging the cerebrospinal fluid (CSF) to the portal capillaries and may link the CSF to neuroendocrine events. During the perinatal period a subpopulation of radial glial cells differentiates into tanycytes, a cell lineage sharing some properties with astrocytes and the radial glia, but displaying unique and distinct morphological, molecular, and functional characteristics. Four populations of tanycytes, alpha(1,2) and beta(1,2), can be distinguished. These subtypes express differentially important functional molecules, such as glucose and glutamate transporters; a series of receptors for neuropeptide and peripheral hormones; secretory molecules such as transforming growth factors, prostaglandin E(2), and the specific protein P85; and proteins of the endocytic pathways. This results in functional differences between the four subtypes of tanycytes. Thus, alpha(1,2) tanycytes do not have barrier properties, whereas beta(1,2) tanycytes do. Different types of tanycytes use different mechanisms to internalize and transport cargo molecules; compounds internalized via a clathrin-dependent endocytosis would only enter tanycytes from the CSF. There are also differences in the neuron-tanycyte relationships; beta(1,2) tanycytes are innervated by peptidergic and aminergic neurons, but alpha(1,2) tanycytes are not. Important aspects of the neuron-beta(1) tanycyte relationships have been elucidated. Tanycytes can participate in the release of gonadotropin-releasing hormone (GnRH) to the portal blood by expressing estrogen receptors, absorbing molecules from the CSF, and providing signal(s) to the GnRH neurons. Removal of tanycytes prevents the pulse of GnRH release into the portal blood, the peak of luteinizing hormone, and ovulation. The discovery in tanycytes of new functional molecules is opening a new field of research. Thus, thyroxine deiodinase type II, an enzyme generating triiodothyronine (T(3)) from thyroxine, appears to be exclusively expressed by tanycytes, suggesting that these cells are the main source of brain T(3). Glucose transporter-2 (GLUT-2), a low-affinity transporter of glucose and fructose, and ATP-sensitive K(+) channels are expressed by tanycytes, suggesting that they may sense CSF glucose concentrations.
Assuntos
Hipotálamo Médio/citologia , Hipotálamo Médio/fisiologia , Sistemas Neurossecretores/fisiologia , Animais , Astrócitos/citologia , Astrócitos/metabolismo , Astrócitos/fisiologia , Barreira Hematoencefálica/citologia , Barreira Hematoencefálica/fisiologia , Encéfalo/citologia , Encéfalo/fisiologia , Líquido Cefalorraquidiano/fisiologia , Glândulas Endócrinas/citologia , Glândulas Endócrinas/fisiologia , Endocitose/fisiologia , Epêndima/química , Epêndima/citologia , Feminino , Hormônio Liberador de Gonadotropina/sangue , Hormônio Liberador de Gonadotropina/líquido cefalorraquidiano , Hipotálamo Médio/metabolismo , Masculino , Neuroglia/citologia , Neuroglia/metabolismo , Neuroglia/fisiologia , Neurônios/fisiologia , Sistemas Neurossecretores/citologia , Ratos , Células-Tronco/citologia , Células-Tronco/fisiologiaRESUMO
Clusterin is shown to contain putative amphipathic alpha-helices that mediate hydrophobic interactions with numerous types of molecules and may be involved in clearance of cellular debris caused by cell injury or death. To assess this function in vivo, we have cloned the full-length cDNA encoding guinea fowl (Numida meleagris) clusterin and studied its synthesis and expression pattern in specific cell types in pituitary. Quantity of clusterin mRNA expressed in pituitary and endocrine tissues was quantified by real-time PCR. Highest levels were detected in gonads. In situ hybridization showed clusterin mRNA in endocrine cells and folliculostellate cells. Clusterin protein detected by immunohistochemistry was observed in endocrine cells, folliculostellate cells and in colloid. The expression pattern suggests that clusterin is produced by endocrine cells for cytoprotection. Degenerating endocrine cells are phagocytosed by folliculostellate cells and digested by their lysosomal enzymes. In folliculostellate cells clusterin interacts and aggregates with by-products of digestion that subsequently become stored in colloid.
Assuntos
Perfilação da Expressão Gênica , Glicoproteínas/metabolismo , Chaperonas Moleculares/metabolismo , Hipófise/metabolismo , Aves Domésticas/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Clusterina , Coloides/química , DNA Complementar/genética , Glândulas Endócrinas/citologia , Glândulas Endócrinas/metabolismo , Glicoproteínas/genética , Técnicas Imunoenzimáticas , Hibridização In Situ , Chaperonas Moleculares/genética , Dados de Sequência Molecular , Fagocitose , Hipófise/citologia , Reação em Cadeia da Polimerase , Sondas RNA , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de AminoácidosRESUMO
Pdx-1 plays a key role in the development of the pancreas and the control of islet gene transcription and has also been proposed as a dominant regulator of the alpha- vs. beta-cell phenotype via extinction of proglucagon expression. To ascertain the relationship between Pdx-1 and proglucagon gene expression, we examined the effect of enhanced pdx-1 expression on proglucagon gene expression in murine islet alphaTC-1 and GLUTag enteroendocrine cells. Although adenoviral transduction increased the levels of pdx-1 mRNA transcripts and nuclear Pdx-1 protein, overexpression of pdx-1 did not repress endogenous proglucagon gene expression in alphaTC-1 or GLUTag cells or murine islets. Immunohistochemical analysis of cells transduced with Ad-pdx-1 demonstrated multiple individual islet or enteroendocrine cells exhibiting both nuclear Pdx-1 and cytoplasmic glucagon-like peptide-1 immunopositivity. The failure of pdx-1 to inhibit endogenous proglucagon gene expression was not attributable to defects in Pdx-1 nuclear translocation or DNA binding as demonstrated using Western blotting and EMSA analyses. Furthermore, Ad-pdx-1 transduction did not repress proglucagon promoter activity in alphaTC-1 or GLUTag cells. Taken together, these findings demonstrate that pdx-1 alone is not sufficient for specification of the hormonal phenotype or extinction of proglucagon gene expression in islet or enteroendocrine cells.
Assuntos
Glândulas Endócrinas/metabolismo , Glucagon/antagonistas & inibidores , Glucagon/genética , Proteínas de Homeodomínio/fisiologia , Mucosa Intestinal/metabolismo , Ilhotas Pancreáticas/metabolismo , Precursores de Proteínas/antagonistas & inibidores , Precursores de Proteínas/genética , Transativadores/fisiologia , Transcrição Gênica/fisiologia , Animais , Linhagem Celular , Glândulas Endócrinas/citologia , Glucagon/metabolismo , Peptídeo 1 Semelhante ao Glucagon , Proteínas de Homeodomínio/metabolismo , Intestinos/citologia , Ilhotas Pancreáticas/citologia , Camundongos , Fragmentos de Peptídeos/metabolismo , Proglucagon , Precursores de Proteínas/metabolismo , Transativadores/metabolismo , Transdução GenéticaRESUMO
Antagonistic activities of glucagon and insulin control metabolism in mammals, and disruption of this balance underlies diabetes pathogenesis. Insulin-producing cells (IPCs) in the brain of insects such as Drosophila also regulate serum glucose, but it remains unclear whether insulin is the sole hormonal regulator of glucose homeostasis and whether mechanisms of glucose-sensing and response in IPCs resemble those in pancreatic islets. Here we show, by targeted cell ablation, that Drosophila corpora cardiaca (CC) cells of the ring gland are also essential for larval glucose homeostasis. Unlike IPCs, CC cells express Drosophila cognates of sulphonylurea receptor (Sur) and potassium channel (Ir), proteins that comprise ATP-sensitive potassium channels regulating hormone secretion by islets and other mammalian glucose-sensing cells. They also produce adipokinetic hormone, a polypeptide with glucagon-like functions. Glucose regulation by CC cells is impaired by exposure to sulphonylureas, drugs that target the Sur subunit. Furthermore, ubiquitous expression of an akh transgene reverses the effect of CC ablation on serum glucose. Thus, Drosophila CC cells are crucial regulators of glucose homeostasis and they use glucose-sensing and response mechanisms similar to islet cells.
Assuntos
Drosophila/citologia , Drosophila/metabolismo , Glândulas Endócrinas/citologia , Glândulas Endócrinas/fisiologia , Glucose/metabolismo , Homeostase , Canais de Potássio Corretores do Fluxo de Internalização , Transportadores de Cassetes de Ligação de ATP/agonistas , Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Drosophila/genética , Drosophila/crescimento & desenvolvimento , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Genes de Insetos/genética , Hemolinfa/metabolismo , Hormônios de Inseto/genética , Hormônios de Inseto/metabolismo , Larva/metabolismo , Oligopeptídeos/genética , Oligopeptídeos/metabolismo , Canais de Potássio/agonistas , Canais de Potássio/metabolismo , Ácido Pirrolidonocarboxílico/análogos & derivados , Receptores de Droga/agonistas , Receptores de Droga/metabolismo , Compostos de Sulfonilureia/farmacologia , Receptores de Sulfonilureias , Transgenes/genéticaRESUMO
Activins and inhibins, which belong to the TGF beta family, are composed of different combinations of alpha-, betaA-, and betaB-subunits, resulting in inhibin A (alphabetaA), inhibin B (alphabetaB), activin A (betaAbetaA), activin B (betaBbetaB), and activin AB (betaAbetaB). They regulate several cell functions, acting as paracrine/autocrine factors. Their actions, which depend on binding to specific receptors, are also modulated by follistatin. Gastroenteropancreatic (GEP) endocrine cells and endocrine tumors (ETs) produce several growth factors, but it is not well known whether they express follistatin and the various inhibin/activin subunits. We studied their expression in 65 GEP ETs using immunohistochemistry (IHC) and in situ hybridization (ISH). The alpha-subunit and follistatin were not identified in normal GEP endocrine cells and were poorly expressed in ETs. A betaA-subunit immunoreactivity (IR) was detected in A-, G-, EC-, and GIP-cells, while betaB-chain IR was present only in D-cells. The mRNAs encoding for these molecules were poorly expressed in normal tissues. BetaA- and betaB-subunits were identified in several ETs by both IHC and ISH: betaA-subunit mainly in G-cell and A-cell ETs, and betaB-subunit in D-cell, A-cell, and EC-cell ETs. Our results demonstrate a differential expression of activin/inhibin subunits among different types of GEP endocrine cells and related tumors, suggesting a role in modulation of biological functions of these normal and neoplastic endocrine cells.