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1.
Endocrinology ; 160(5): 1359-1361, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-31144717
2.
Cell Metab ; 2018 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-30449685

RESUMO

Identification of cell-surface markers specific to human pancreatic ß cells would allow in vivo analysis and imaging. Here we introduce a biomarker, ectonucleoside triphosphate diphosphohydrolase-3 (NTPDase3), that is expressed on the cell surface of essentially all adult human ß cells, including those from individuals with type 1 or type 2 diabetes. NTPDase3 is expressed dynamically during postnatal human pancreas development, appearing first in acinar cells at birth, but several months later its expression declines in acinar cells while concurrently emerging in islet ß cells. Given its specificity and membrane localization, we utilized an NTPDase3 antibody for purification of live human ß cells as confirmed by transcriptional profiling, and, in addition, for in vivo imaging of transplanted human ß cells. Thus, NTPDase3 is a cell-surface biomarker of adult human ß cells, and the antibody directed to this protein should be a useful new reagent for ß cell sorting, in vivo imaging, and targeting.

3.
Cell Metab ; 25(6): 1362-1373.e5, 2017 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-28591638

RESUMO

Decreasing glucagon action lowers the blood glucose and may be useful therapeutically for diabetes. However, interrupted glucagon signaling leads to α cell proliferation. To identify postulated hepatic-derived circulating factor(s) responsible for α cell proliferation, we used transcriptomics/proteomics/metabolomics in three models of interrupted glucagon signaling and found that proliferation of mouse, zebrafish, and human α cells was mTOR and FoxP transcription factor dependent. Changes in hepatic amino acid (AA) catabolism gene expression predicted the observed increase in circulating AAs. Mimicking these AA levels stimulated α cell proliferation in a newly developed in vitro assay with L-glutamine being a critical AA. α cell expression of the AA transporter Slc38a5 was markedly increased in mice with interrupted glucagon signaling and played a role in α cell proliferation. These results indicate a hepatic α islet cell axis where glucagon regulates serum AA availability and AAs, especially L-glutamine, regulate α cell proliferation and mass via mTOR-dependent nutrient sensing.


Assuntos
Proliferação de Células , Glucagon/metabolismo , Glutamina/metabolismo , Fígado/metabolismo , Transdução de Sinais , Sistemas de Transporte de Aminoácidos Neutros/genética , Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Animais , Glucagon/genética , Glutamina/genética , Camundongos , Camundongos Knockout , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
5.
J Endocrinol ; 227(2): 93-103, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26446275

RESUMO

Glucagon antagonism is a potential treatment for diabetes. One potential side effect is α-cell hyperplasia, which has been noted in several approaches to antagonize glucagon action. To investigate the molecular mechanism of the α-cell hyperplasia and to identify the responsible factor, we created a zebrafish model in which glucagon receptor (gcgr) signaling has been interrupted. The genetically and chemically tractable zebrafish, which provides a robust discovery platform, has two gcgr genes (gcgra and gcgrb) in its genome. Sequence, phylogenetic, and synteny analyses suggest that these are co-orthologs of the human GCGR. Similar to its mammalian counterparts, gcgra and gcgrb are mainly expressed in the liver. We inactivated the zebrafish gcgra and gcgrb using transcription activator-like effector nuclease (TALEN) first individually and then both genes, and assessed the number of α-cells using an α-cell reporter line, Tg(gcga:GFP). Compared to WT fish at 7 days postfertilization, there were more α-cells in gcgra-/-, gcgrb-/-, and gcgra-/-;gcgrb-/- fish and there was an increased rate of α-cell proliferation in the gcgra-/-;gcgrb-/- fish. Glucagon levels were higher but free glucose levels were lower in gcgra-/-, gcgrb-/-, and gcgra-/-;gcgrb-/- fish, similar to Gcgr-/- mice. These results indicate that the compensatory α-cell hyperplasia in response to interruption of glucagon signaling is conserved in zebrafish. The robust α-cell hyperplasia in gcgra-/-;gcgrb-/- larvae provides a platform to screen for chemical and genetic suppressors, and ultimately to identify the stimulus of α-cell hyperplasia and its signaling mechanism.


Assuntos
Inativação Gênica , Células Secretoras de Glucagon/patologia , Receptores de Glucagon/genética , Animais , Animais Geneticamente Modificados , Proliferação de Células/genética , Clonagem Molecular , Embrião não Mamífero , Regulação da Expressão Gênica no Desenvolvimento , Células Secretoras de Glucagon/metabolismo , Hiperplasia/genética , Receptores de Glucagon/metabolismo , Peixe-Zebra
6.
Diabetes ; 62(4): 1196-205, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23160527

RESUMO

Glucagon is a critical regulator of glucose homeostasis; however, mechanisms regulating glucagon action and α-cell function and number are incompletely understood. To elucidate the role of the hepatic glucagon receptor (Gcgr) in glucagon action, we generated mice with hepatocyte-specific deletion of the glucagon receptor. Gcgr(Hep)(-/-) mice exhibited reductions in fasting blood glucose and improvements in insulin sensitivity and glucose tolerance compared with wild-type controls, similar in magnitude to changes observed in Gcgr(-/-) mice. Despite preservation of islet Gcgr signaling, Gcgr(Hep)(-/-) mice developed hyperglucagonemia and α-cell hyperplasia. To investigate mechanisms by which signaling through the Gcgr regulates α-cell mass, wild-type islets were transplanted into Gcgr(-/-) or Gcgr(Hep)(-/-) mice. Wild-type islets beneath the renal capsule of Gcgr(-/-) or Gcgr(Hep)(-/-) mice exhibited an increased rate of α-cell proliferation and expansion of α-cell area, consistent with changes exhibited by endogenous α-cells in Gcgr(-/-) and Gcgr(Hep)(-/-) pancreata. These results suggest that a circulating factor generated after disruption of hepatic Gcgr signaling can increase α-cell proliferation independent of direct pancreatic input. Identification of novel factors regulating α-cell proliferation and mass may facilitate the generation and expansion of α-cells for transdifferentiation into ß-cells and the treatment of diabetes.


Assuntos
Células Secretoras de Glucagon/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Fígado/metabolismo , Receptores de Glucagon/metabolismo , Animais , Glicemia , Feminino , Glucagon/administração & dosagem , Glucagon/sangue , Células Secretoras de Glucagon/citologia , Células Secretoras de Glucagon/patologia , Glucose/metabolismo , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hiperplasia , Resistência à Insulina , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Fígado/citologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Glucagon/genética , Transdução de Sinais
7.
PLoS One ; 7(7): e39227, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22768297

RESUMO

Recent clinical evidence supports a link between 25-hydroxyvitamin D insufficiency (serum 25-hydroxyvitamin D [25(OH)D] levels <30 ng/mL) and Parkinson's disease. To investigate the effect of 25(OH)D depletion on neuronal susceptibility to toxic insult, we induced a state of 25(OH)D deficiency in mice and then challenged them with the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). We found there was no significant difference between control and 25(OH)D-deficient animals in striatal dopamine levels or dopamine transporter and tyrosine hydroxylase expression after lesioning with MPTP. Additionally, we found no difference in tyrosine hydroxylase expression in the substantia nigra pars compacta. Our data suggest that reducing 25(OH)D serum levels in mice has no effect on the vulnerability of nigral dopaminergic neurons in vivo in this model system of parkinsonism.


Assuntos
1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/efeitos adversos , Corpo Estriado/metabolismo , Neurônios Dopaminérgicos/metabolismo , Intoxicação por MPTP/metabolismo , Neurotoxinas/efeitos adversos , Substância Negra/metabolismo , Vitamina D/análogos & derivados , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/farmacologia , Animais , Corpo Estriado/patologia , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Neurônios Dopaminérgicos/patologia , Intoxicação por MPTP/patologia , Masculino , Camundongos , Proteínas do Tecido Nervoso/metabolismo , Neurotoxinas/farmacologia , Substância Negra/patologia , Tirosina 3-Mono-Oxigenase/metabolismo , Vitamina D/metabolismo
8.
Neurotoxicology ; 29(5): 855-63, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18577399

RESUMO

Parkinson's disease (PD) is primarily thought of as a disease of aging. However, recent evidence points to the potential for exposure to xenobiotics during development to increase risk of PD. Here, we report that developmental exposure to the organochlorine pesticide heptachlor alters the dopamine system and increases neurotoxicity in an animal model of PD. Exposure of pregnant mice to heptachlor led to increased levels of the dopamine transporter (DAT) and vesicular monoamine transporter 2 (VMAT2) levels at both the protein and mRNA level in their offspring. Increased DAT and VMAT2 levels were accompanied by alterations of mRNA levels of nuclear transcription factors that control dopamine neuron development and regulate DAT and VMAT2 levels in adulthood. At 12 weeks of age, control and heptachlor-exposed offspring were administered a moderate dose (2 x 10mg/kg) of the parkinsonism-inducing agent MPTP. Greater neurotoxicity as evidenced by a greater loss of striatal dopamine and potentiation of increased levels of glial fibrillary acidic protein and alpha-synuclein was observed in heptachlor-exposed offspring. The neurotoxicity observed was greater in the male offspring than the female offspring, suggesting that males are more susceptible to the long-term effects of developmental heptachlor exposure. These data suggest that developmental heptachlor exposure causes long-term alterations of the dopamine system thereby rendering it more susceptible to dopaminergic damage in adulthood.


Assuntos
Dopamina/metabolismo , Heptacloro/toxicidade , Intoxicação por MPTP/patologia , Neurônios/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal , Análise de Variância , Animais , Animais Recém-Nascidos , Modelos Animais de Doenças , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Heptacloro/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Gravidez , Fatores Sexuais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Vesiculares de Transporte de Monoamina/genética , Proteínas Vesiculares de Transporte de Monoamina/metabolismo
9.
FASEB J ; 20(10): 1695-7, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16809432

RESUMO

Exposure to pesticides has been suggested to increase the risk of Parkinson's disease (PD), but the mechanisms responsible for this association are not clear. Here, we report that perinatal exposure of mice during gestation and lactation to low levels of dieldrin (0.3, 1, or 3 mg/kg every 3 days) alters dopaminergic neurochemistry in their offspring and exacerbates MPTP toxicity. At 12 wk of age, protein and mRNA levels of the dopamine transporter (DAT) and vesicular monoamine transporter 2 (VMAT2) were increased by perinatal dieldrin exposure in a dose-related manner. We then administered MPTP (2 x 10 mg/kg s.c) at 12 wk of age and observed a greater reduction of striatal dopamine in dieldrin-exposed offspring, which was associated with a greater DAT:VMAT2 ratio. Additionally, dieldrin exposure during development potentiated the increase in GFAP and alpha-synuclein levels induced by MPTP, indicating increased neurotoxicity. In all cases there were greater effects observed in the male offspring than the female, similar to that observed in human cases of PD. These data suggest that developmental exposure to dieldrin leads to persistent alterations of the developing dopaminergic system and that these alterations induce a "silent" state of dopamine dysfunction, thereby rendering dopamine neurons more vulnerable later in life.


Assuntos
Dieldrin/farmacologia , Dopamina/metabolismo , Síndromes Neurotóxicas/etiologia , Doença de Parkinson Secundária/induzido quimicamente , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/administração & dosagem , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/farmacologia , Animais , Animais Recém-Nascidos , Dieldrin/administração & dosagem , Modelos Animais de Doenças , Proteínas da Membrana Plasmática de Transporte de Dopamina/análise , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Sinergismo Farmacológico , Feminino , Lactação , Masculino , Camundongos , Doença de Parkinson Secundária/etiologia , Praguicidas/farmacologia , Gravidez , RNA Mensageiro/análise , Proteínas Vesiculares de Transporte de Monoamina/análise , Proteínas Vesiculares de Transporte de Monoamina/genética
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