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1.
PLoS Comput Biol ; 20(1): e1011811, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38289902

RESUMO

The endocrine cells of the pituitary gland are electrically active, and in vivo they form small networks where the bidirectional cell-cell coupling is through gap junctions. Numerous studies of dispersed pituitary cells have shown that typical behaviors are tonic spiking and bursting, the latter being more effective at evoking secretion. In this article, we use mathematical modeling to examine the dynamics of small networks of spiking and bursting pituitary cells. We demonstrate that intrinsic bursting cells are capable of converting intrinsic spikers into bursters, and perform a fast/slow analysis to show why this occurs. We then demonstrate the sensitivity of network dynamics to the placement of bursting cells within the network, and demonstrate strategies that are most effective at maximizing secretion from the population of cells. This study provides insights into the in vivo behavior of cells such as the stress-hormone-secreting pituitary corticotrophs that are switched from spiking to bursting by hypothalamic neurohormones. While much is known about the electrical properties of these cells when isolated from the pituitary, how they behave when part of an electrically coupled network has been largely unstudied.


Assuntos
Células Endócrinas , Hipófise , Modelos Teóricos , Potenciais de Ação
3.
Mol Metab ; 79: 101853, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38103636

RESUMO

OBJECTIVE: The consequences of mutations in genes associated with monogenic forms of diabetes on human pancreas development cannot be studied in a time-resolved fashion in vivo. More specifically, if recessive mutations in the insulin gene influence human pancreatic endocrine lineage formation is still an unresolved question. METHODS: To model the extremely reduced insulin levels in patients with recessive insulin gene mutations, we generated a novel knock-in H2B-Cherry reporter human induced pluripotent stem cell (iPSC) line expressing no insulin upon differentiation to stem cell-derived (SC-) ß cells in vitro. Differentiation of iPSCs into the pancreatic and endocrine lineage, combined with immunostaining, Western blotting and proteomics analysis phenotypically characterized the insulin gene deficiency in SC-islets. Furthermore, we leveraged FACS analysis and confocal microscopy to explore the impact of insulin shortage on human endocrine cell induction, composition, differentiation and proliferation. RESULTS: Interestingly, insulin-deficient SC-islets exhibited low insulin receptor (IR) signaling when stimulated with glucose but displayed increased IR sensitivity upon treatment with exogenous insulin. Furthermore, insulin shortage did not alter neurogenin-3 (NGN3)-mediated endocrine lineage induction. Nevertheless, lack of insulin skewed the SC-islet cell composition with an increased number in SC-ß cell formation at the expense of SC-α cells. Finally, insulin deficiency reduced the rate of SC-ß cell proliferation but had no impact on the expansion of SC-α cells. CONCLUSIONS: Using iPSC disease modelling, we provide first evidence of insulin function in human pancreatic endocrine lineage formation. These findings help to better understand the phenotypic impact of recessive insulin gene mutations during pancreas development and shed light on insulin gene function beside its physiological role in blood glucose regulation.


Assuntos
Células Endócrinas , Células-Tronco Pluripotentes Induzidas , Humanos , Insulina/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Diferenciação Celular/genética , Pâncreas/metabolismo , Insulina Regular Humana/metabolismo , Células Endócrinas/metabolismo
4.
Cell Metab ; 35(11): 1944-1960.e7, 2023 11 07.
Artigo em Inglês | MEDLINE | ID: mdl-37898119

RESUMO

Human pancreatic plasticity is implied from multiple single-cell RNA sequencing (scRNA-seq) studies. However, these have been invariably based on static datasets from which fate trajectories can only be inferred using pseudotemporal estimations. Furthermore, the analysis of isolated islets has resulted in a drastic underrepresentation of other cell types, hindering our ability to interrogate exocrine-endocrine interactions. The long-term culture of human pancreatic slices (HPSs) has presented the field with an opportunity to dynamically track tissue plasticity at the single-cell level. Combining datasets from same-donor HPSs at different time points, with or without a known regenerative stimulus (BMP signaling), led to integrated single-cell datasets storing true temporal or treatment-dependent information. This integration revealed population shifts consistent with ductal progenitor activation, blurring of ductal/acinar boundaries, formation of ducto-acinar-endocrine differentiation axes, and detection of transitional insulin-producing cells. This study provides the first longitudinal scRNA-seq analysis of whole human pancreatic tissue, confirming its plasticity in a dynamic fashion.


Assuntos
Células Endócrinas , Análise da Expressão Gênica de Célula Única , Humanos , Pâncreas , Diferenciação Celular
6.
Nature ; 622(7981): 164-172, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37674082

RESUMO

Development of immunocompetent T cells in the thymus is required for effective defence against all types of pathogens, including viruses, bacteria and fungi. To this end, T cells undergo a very strict educational program in the thymus, during which both non-functional and self-reactive T cell clones are eliminated by means of positive and negative selection1.Thymic epithelial cells (TECs) have an indispensable role in these processes, and previous studies have shown the notable heterogeneity of these cells2-7. Here, using multiomic analysis, we provide further insights into the functional and developmental diversity of TECs in mice, and reveal a detailed atlas of the TEC compartment according to cell transcriptional states and chromatin landscapes. Our analysis highlights unconventional TEC subsets that are similar to functionally well-defined parenchymal populations, including endocrine cells, microfold cells and myocytes. By focusing on the endocrine and microfold TEC populations, we show that endocrine TECs require Insm1 for their development and are crucial to maintaining thymus cellularity in a ghrelin-dependent manner; by contrast, microfold TECs require Spib for their development and are essential for the generation of thymic IgA+ plasma cells. Collectively, our study reveals that medullary TECs have the potential to differentiate into various types of molecularly distinct and functionally defined cells, which not only contribute to the induction of central tolerance, but also regulate the homeostasis of other thymus-resident populations.


Assuntos
Tolerância a Antígenos Próprios , Linfócitos T , Timo , Animais , Camundongos , Diferenciação Celular , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Tolerância a Antígenos Próprios/imunologia , Tolerância a Antígenos Próprios/fisiologia , Linfócitos T/classificação , Linfócitos T/citologia , Linfócitos T/imunologia , Timo/citologia , Timo/imunologia , Tecido Parenquimatoso , Células Musculares , Células Endócrinas , Cromatina , Transcrição Gênica , Grelina
7.
Nat Commun ; 14(1): 5554, 2023 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-37689751

RESUMO

NEUROD1 is a transcription factor that helps maintain a mature phenotype of pancreatic ß cells. Disruption of Neurod1 during pancreatic development causes severe neonatal diabetes; however, the exact role of NEUROD1 in the differentiation programs of endocrine cells is unknown. Here, we report a crucial role of the NEUROD1 regulatory network in endocrine lineage commitment and differentiation. Mechanistically, transcriptome and chromatin landscape analyses demonstrate that Neurod1 inactivation triggers a downregulation of endocrine differentiation transcription factors and upregulation of non-endocrine genes within the Neurod1-deficient endocrine cell population, disturbing endocrine identity acquisition. Neurod1 deficiency altered the H3K27me3 histone modification pattern in promoter regions of differentially expressed genes, which resulted in gene regulatory network changes in the differentiation pathway of endocrine cells, compromising endocrine cell potential, differentiation, and functional properties.


Assuntos
Células Endócrinas , Células Secretoras de Insulina , Diferenciação Celular/genética , Fatores de Transcrição , Ativação Transcricional
8.
Sci Adv ; 9(33): eadf5142, 2023 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-37595046

RESUMO

In contrast to mice, zebrafish have an exceptional yet elusive ability to replenish lost ß cells in adulthood. Understanding this framework would provide mechanistic insights for ß cell regeneration, which may be extrapolated to humans. Here, we characterize a krt4-expressing ductal cell type, which is distinct from the putative Notch-responsive cells, showing neogenic competence and giving rise to the majority of endocrine cells during postembryonic development. Furthermore, we demonstrate a marked ductal remodeling process featuring a Notch-responsive to krt4+ luminal duct transformation during late development, indicating several origins of krt4+ ductal cells displaying similar transcriptional patterns. Single-cell transcriptomics upon a series of time points during ß cell regeneration unveil a previously unrecognized dlb+ transitional endocrine precursor cell, distinct regulons, and a differentiation trajectory involving cellular shuffling through differentiation and dedifferentiation dynamics. These results establish a model of zebrafish pancreatic endocrinogenesis and highlight key values of zebrafish for translational studies of ß cell regeneration.


Assuntos
Células Endócrinas , Células Secretoras de Insulina , Humanos , Animais , Camundongos , Peixe-Zebra , Diferenciação Celular , Regeneração
9.
Cell Tissue Res ; 393(2): 393-399, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37314493

RESUMO

In the past years, it has become clear that the family of Mas-related G protein-coupled receptors plays a central role in neuro-immune communication at mucosal barrier surfaces, in particular in the skin. Remarkably, MRGPR expression at other mucosal surfaces remains poorly characterized. To fill this gap in our understanding, the present study was undertaken to screen and verify the expression of the human MRGPR family members in the mucosal biopsies of the human gastrointestinal (GI) tract. Our findings revealed that, of all human MRGPRs family members, only MRGPRF mRNA is expressed at detectable levels in human mucosal biopsies of both terminal ileum and sigmoid colon. Furthermore, immunohistochemical stainings revealed that MRGPRF is specifically expressed by mucosal entero-endocrine cells (EECs). Overall, this study showed for the first time that the human ileum and colonic mucosa represent a novel expression site for the orphan MRGPRF, more specifically in EECs.


Assuntos
Células Endócrinas , Mucosa Intestinal , Humanos , Mucosa Intestinal/metabolismo , Trato Gastrointestinal/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Colo/metabolismo , Células Endócrinas/metabolismo , Células Enteroendócrinas/metabolismo
10.
Islets ; 15(1): 2219477, 2023 12 31.
Artigo em Inglês | MEDLINE | ID: mdl-37258189

RESUMO

Previous studies indicated that ductal cells can contribute to endocrine neogenesis in adult rodents after alpha cells convert into beta cells. This can occur through Pax4 mis-expression in alpha cells or through long-term administration of gamma-aminobutyric acid (GABA) to healthy mice. GABA has also been reported to increase the number of beta cells through direct effects on their proliferation, but only in specific genetic mouse backgrounds. To test whether GABA induces neogenesis of beta cells from ductal cells or affects pancreatic cell proliferation, we administered GABA or saline over 2 or 6 months to Sox9CreER;R26RYFP mice in which 60-80% of large or small ducts were efficiently lineage labeled. We did not observe any increases in islet neogenesis from ductal cells between 1 and 2 months of age in saline treated mice, nor between 2 and 6 months of saline treatment, supporting previous studies indicating that adult ductal cells do not give rise to new endocrine cells during homeostasis. Unlike previous reports, we did not observe an increase in beta cell neogenesis after 2 or 6 months of GABA administration. Nor did we observe a significant increase in the pancreatic islet area, the number of insulin and glucagon double positive cells, or cell proliferation in the pancreas. This indicates that the effect of long term GABA administration on the pancreas is minimal or highly context dependent.


Assuntos
Células Endócrinas , Células Secretoras de Glucagon , Células Secretoras de Insulina , Ilhotas Pancreáticas , Camundongos , Animais , Ductos Pancreáticos , Ácido gama-Aminobutírico/farmacologia
11.
Stem Cell Res Ther ; 14(1): 101, 2023 04 25.
Artigo em Inglês | MEDLINE | ID: mdl-37098639

RESUMO

The efficiency of inducing human embryonic stem cells into NEUROG3+ pancreatic endocrine cells is a bottleneck in stem cell therapy for diabetes. To understand the cell properties and fate decisions during differentiation, we analyzed the modified induction method using single-cell transcriptome and found that DAPT combined with four factors (4FS): nicotinamide, dexamethasone, forskolin and Alk5 inhibitor II (DAPT + 4FS) increased the expression of NEUROG3 to approximately 34.3%. The increased NEUROG3+ cells were mainly concentrated in Insulin + Glucagon + (INS + GCG+) and SLAC18A1 + Chromogranin A+(SLAC18A1 + CHGA +) populations, indicating that the increased NEUROG3+ cells promoted the differentiation of pancreatic endocrine cells and enterochromaffin-like cells. Single-cell transcriptome analysis provided valuable clues for further screening of pancreatic endocrine cells and differentiation of pancreatic islet cells. The gene set enrichment analysis (GSEA) suggest that we can try to promote the expression of INS + GCG+ population by up-regulating G protein-coupled receptor (GPCR) and mitogen-activated protein kinase signals and down-regulating Wnt, NIK/NF-KappaB and cytokine-mediated signal pathways. We can also try to regulate GPCR signaling through PLCE1, so as to increase the proportion of NEUROG3+ cells in INS+GCG+ populations. To exclude non-pancreatic endocrine cells, ALCAMhigh CD9low could be used as a marker for endocrine populations, and ALCAMhigh CD9lowCDH1low could remove the SLC18A1 + CHGA+ population.


Assuntos
Molécula de Adesão de Leucócito Ativado , Células Endócrinas , Humanos , Molécula de Adesão de Leucócito Ativado/genética , Molécula de Adesão de Leucócito Ativado/metabolismo , Inibidores da Agregação Plaquetária/metabolismo , Análise da Expressão Gênica de Célula Única , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas do Tecido Nervoso/metabolismo , Diferenciação Celular/genética , Glucagon , Células Endócrinas/metabolismo , Proteínas Vesiculares de Transporte de Monoamina/genética , Proteínas Vesiculares de Transporte de Monoamina/metabolismo
13.
Anim Sci J ; 94(1): e13814, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36752108

RESUMO

Chicken adenohypophyseal cells were cultured in plates coated with different materials, and their morphologies were examined to confirm the characteristics of chicken folliculo-stellate (FS) cells in vitro. The adenohypophyseal cells were dispersed with a collagenase/trypsin mixture in media and seeded in plates coated in either poly L-lysine (PLL), collagen, or laminin. After 7 days of culture, the cells were fixed and immunocytochemistry was performed. 5-Bromo-2'-deoxyuridine incorporation test indicated that the proliferation activity of the culture cells was different based on the coating materials, and it was higher in the collagen-coated plate than two other coating materials. Fluorescence immunocytochemistry was also performed using mixed antibodies against growth hormone, prolactin, luteinizing hormone ß-subunit, basic cytokeratin (bCK), and S100B. The culture cells on the PLL- and laminin-coated surfaces were round or oval in shape, and bCK-immunopositive FS cells were morphologically indistinguishable from endocrine cells. In the collagen-coated plate, many endocrine cells were round or oval in shape, but FS cells displayed a larger and flattened morphology. S100B-immunoreactions were localized in the nuclei of bCK-immunopositive FS cells. These results suggest that culturing the chicken adenohypophyseal cells in the collagen-coated plate enables the distinction of FS cells from endocrine cells.


Assuntos
Galinhas , Células Endócrinas , Animais , Galinhas/metabolismo , Laminina , Prolactina/metabolismo , Colágeno , Células Endócrinas/metabolismo , Células Cultivadas
14.
Methods Mol Biol ; 2565: 35-42, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36205885

RESUMO

Immunocytochemistry enables the detection and localization of proteins in cells that are acutely dissociated or in culture. There are advantages and disadvantages to the use of cultured cells for immunocytochemistry. One of the advantages is that cultured cells can be used for one or more weeks after the dissociation of cells, whereas one of the disadvantages is that the properties of cells in culture might change under artificial conditions. On the other hand, acutely dissociated cells are expected to have the original properties of cells because almost all procedures before fixation, except for enzymatic digestion, are carried out at low temperatures. Here, we describe how adrenal medullary cells of small animals are acutely dissociated for immunostaining.


Assuntos
Medula Suprarrenal , Células Cromafins , Células Endócrinas , Animais , Células Cultivadas , Imuno-Histoquímica
15.
Life Sci ; 311(Pt A): 121141, 2022 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-36341914

RESUMO

BACKGROUND AND AIMS: Although oral berberine, a natural compound extracted from the Chinese herbal medicine curcumin, has low bioavailability, it is still effective in suppressing obesity; however, the underlying mechanism is unclear. Berberine can bind to bitter-taste receptors (TAS2Rs) in intestinal endocrine secretin tumor (STC-1) cells to promote glucagon-like peptide-1 (GLP-1) secretion. Notably, TAS2Rs also exist in the tuft cells of the gut. Therefore, this study aimed to explore whether the beneficial effect of oral berberine on obesity is dependent on bitter-taste signaling in the tuft cells of the gut. METHODS AND RESULTS: Standard chow diet (SCD) or high-fat diet (HFD) was administered to C57BL/6 mice, with or without berberine (100 mg/kg, 200 mg/kg, p. o.). The PLCß2 inhibitor U73122 was used to verify whether the anti-obesity effect of berberine was dependent on the bitter-taste signaling pathway. In this study, we observed that the oral administration of berberine alleviated HFD-induced obesity in mice that U73122 partially inhibited. Both in vivo and ex vivo, berberine upregulated the release of GLP-1, promoted the proliferation of tuft cells and secretion of IL-25 in obesity via the TAS2R signaling pathway. CONCLUSIONS: Oral berberine ameliorated HFD-induced obesity through the TAS2R-IL-25 signaling pathway in tuft cells in the gut. SIGNIFICANCE: We identified and functionally characterized the TAS2Rs and Gα-gustducin/Gß1γ13 signaling pathway utilized by tuft cells in response to oral berberine in obese mice and proposed a new mechanism underlying the anti-obesity effect of berberine.


Assuntos
Berberina , Células Endócrinas , Animais , Camundongos , Berberina/farmacologia , Dieta Hiperlipídica , Células Endócrinas/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Obesos , Obesidade/tratamento farmacológico , Obesidade/metabolismo , Receptores Acoplados a Proteínas G/metabolismo
16.
Ter Arkh ; 94(2): 194-199, 2022 Feb 15.
Artigo em Russo | MEDLINE | ID: mdl-36286744

RESUMO

AIM: To establish the role of the main risk factors and endocrine cells of the antrum of the stomach producing motilin (M-cells) in the occurrence of cholelithiasis. MATERIALS AND METHODS: The first group included 122 patients with cholelithiasis. The second group consisted of 30 healthy individuals who underwent medical examination. The groups were matched for gender and age. The work analyzed anamnestic, biochemical and anthropometric data. All patients underwent esophagogastroduodenoscopy with targeted biopsy of the mucous membrane from the antrum. Biopsies were subjected to cytological and immunohistochemical studies in order to verify Helicobacter pylori and estimate the number of M-cells. RESULTS: Patients with cholelithiasis more often belonged to the group of people of mental labor, had low physical activity, were committed to inappropriate nutrition and more often indicated the presence of aggravated heredity for cholelithiasis. Patients with gallstone disease had higher body mass index, waist volume, total cholesterol, low-density lipoprotein cholesterol, triglycerides, glucose, lower high-density lipoprotein cholesterol, H. pylori infection was more often verified and M-cell hypoplasia in the mucous membrane was established. stomach in comparison with the representatives of the second group. CONCLUSION: Our results suggest that certain external factors, nutritional characteristics of the metabolic syndrome components, hypoplasia of M-cells in the gastric mucosa are important factors in the formation of calculi in the gallbladder.


Assuntos
Colelitíase , Células Endócrinas , Infecções por Helicobacter , Helicobacter pylori , Humanos , Motilina , Infecções por Helicobacter/complicações , Infecções por Helicobacter/epidemiologia , Estômago , Mucosa Gástrica , Colelitíase/epidemiologia , Fatores de Risco , Triglicerídeos , Células Endócrinas/patologia , Colesterol , Glucose , Lipoproteínas HDL , Lipoproteínas LDL
17.
Nat Commun ; 13(1): 4540, 2022 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-35927244

RESUMO

During pancreas development endocrine cells leave the ductal epithelium to form the islets of Langerhans, but the morphogenetic mechanisms are incompletely understood. Here, we identify the Ca2+-independent atypical Synaptotagmin-13 (Syt13) as a key regulator of endocrine cell egression and islet formation. We detect specific upregulation of the Syt13 gene and encoded protein in endocrine precursors and the respective lineage during islet formation. The Syt13 protein is localized to the apical membrane of endocrine precursors and to the front domain of egressing endocrine cells, marking a previously unidentified apical-basal to front-rear repolarization during endocrine precursor cell egression. Knockout of Syt13 impairs endocrine cell egression and skews the α-to-ß-cell ratio. Mechanistically, Syt13 is a vesicle trafficking protein, transported via the microtubule cytoskeleton, and interacts with phosphatidylinositol phospholipids for polarized localization. By internalizing a subset of plasma membrane proteins at the front domain, including α6ß4 integrins, Syt13 modulates cell-matrix adhesion and allows efficient endocrine cell egression. Altogether, these findings uncover an unexpected role for Syt13 as a morphogenetic driver of endocrinogenesis and islet formation.


Assuntos
Células Endócrinas , Ilhotas Pancreáticas , Integrinas , Morfogênese , Pâncreas , Sinaptotagminas/genética
18.
Front Endocrinol (Lausanne) ; 13: 936160, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35872987

RESUMO

The endocrine cells of the anterior pituitary gland are electrically active when stimulated or, in some cases, when not inhibited. The activity pattern thought to be most effective in releasing hormones is bursting, which consists of depolarization with small spikes that are much longer than single spikes. Although a majority of the research on cellular activity patterns has been performed on dispersed cells, the environment in situ is characterized by networks of coupled cells of the same type, at least in the case of somatotrophs and lactotrophs. This produces some degree of synchronization of their activity, which can be greatly increased by hormones and changes in the physiological state. In this computational study, we examine how electrical coupling among model cells influences synchronization of bursting oscillations among the population. We focus primarily on weak electrical coupling, since strong coupling leads to complete synchronization that is not characteristic of pituitary cell networks. We first look at small networks to point out several unexpected behaviors of the coupled system, and then consider a larger random scale-free network to determine what features of the structural network formed through gap junctional coupling among cells produce a high degree of functional coupling, i.e., clusters of synchronized cells. We employ several network centrality measures, and find that cells that are closely related in terms of their closeness centrality are most likely to be synchronized. We also find that structural hubs (cells with extensive coupling to other cells) are typically not functional hubs (cells synchronized with many other cells). Overall, in the case of weak electrical coupling, it is hard to predict the functional network that arises from a structural network, or to use a functional network as a means for determining the structural network that gives rise to it.


Assuntos
Células Endócrinas , Hipófise , Potenciais de Ação , Hormônios
19.
Front Endocrinol (Lausanne) ; 13: 904004, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35769082

RESUMO

Diabetes mellitus, a disease that affects nearly 536.6 million people worldwide, is characterized by the death or dysfunction of insulin-producing beta cells of the pancreas. The beta cells are found within the islets of Langerhans, which are composed of multiple hormone-producing endocrine cells including the alpha (glucagon), delta (somatostatin), PP (pancreatic polypeptide), and epsilon (ghrelin) cells. There is direct evidence that physical and paracrine interactions between the cells in the islet facilitate and support beta cell function. However, communication between endocrine and exocrine cells in the pancreas may also directly impact beta cell growth and function. Herein we review literature that contributes to the view that "crosstalk" between neighboring cells within the pancreas influences beta cell growth and function and the maintenance of beta cell health.


Assuntos
Diabetes Mellitus , Células Endócrinas , Células Secretoras de Insulina , Glucagon , Humanos , Pâncreas
20.
Acta Histochem ; 124(5): 151898, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-35526370

RESUMO

TWIK-related acid-sensitive K+ (TASK) channels are thought to contribute to the resting membrane potential in adrenal cortical (AC) cells. However, the molecular identity of TASK channels in AC cells have not yet been elucidated. Thus, immunocytochemical and molecular biological approaches were employed to investigate the expression and intracellular distribution of TASK1 and TASK3 in mouse AC cells and H295R cells derived from human adrenocortical carcinoma. Immunocytochemical study revealed that immunoreactive materials were mainly located in the cytoplasm for TASK1 and at the cell periphery for TASK3 in mouse AC cells. A similar pattern of localization was observed when GFP-TASK1 and GFP-TASK3 were exogenously expressed in H295R cells. In addition, p11 that is known to suppress the endoplasmic reticulum exit of TASK1 was localized in the cytoplasm in mouse AC and H295R cells, but not in adrenal medullary cells. Proximity ligation assay (PLA) suggested formation of heteromeric TASK1-3 channels that were found predominantly in the cytoplasm and weakly at the cell periphery. A similar distribution was observed following exogenous expression of tandem TASK1-3 channels in H295R cells. When stimulated by angiotensin II, however, tandem TASK1-3 channels were present mainly in the cytoplasm in all H295R cells. In contrast to that in H295R cells, tandem channels were exclusively located at the cell periphery in all non-stimulated and exclusively in the cytoplasm in stimulated PC12 cells, respectively. From these results, we conclude that TASK1 proteins are present mainly in the cytoplasm and minimally at the cell periphery as a heteromeric channel with TASK3, whereas the majority of TASK3 is at the cell periphery as homomeric and heteromeric channels.


Assuntos
Células Endócrinas , Proteínas do Tecido Nervoso/metabolismo , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Angiotensina II/metabolismo , Animais , Células Endócrinas/metabolismo , Humanos , Camundongos , Células PC12 , Canais de Potássio/metabolismo , Ratos
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