RESUMO
Skeletal dysplasias are common disabling disorders characterized by aberrant growth of bone and cartilage leading to abnormal skeletal structures and functions, often attributable to defects in skeletal progenitor cells. The underlying molecular and cellular mechanisms of most skeletal dysplasias remain elusive. Although the Wnt/ß-catenin signaling pathway is required for skeletal progenitor cells to differentiate along the osteoblastic lineage, inappropriately elevated levels of signaling can also inhibit bone formation by suppressing osteoblast maturation. Here, we investigate interactions of the four major Gα protein families (Gα(s), Gα(i/o), Gα(q/11), and Gα(12/13)) with the Wnt/ß-catenin signaling pathway and identify a causative role of Wnt/ß-catenin signaling in fibrous dysplasia (FD) of bone, a disease that exhibits abnormal differentiation of skeletal progenitor cells. The activating Gα(s) mutations that cause FD potentiated Wnt/ß-catenin signaling, and removal of Gα(s) led to reduced Wnt/ß-catenin signaling and decreased bone formation. We further show that activation of Wnt/ß-catenin signaling in osteoblast progenitors results in an FD-like phenotype and reduction of ß-catenin levels rescued differentiation defects of FD patient-derived stromal cells. Gα proteins may act at the level of ß-catenin destruction complex assembly by binding Axin. Our results indicate that activated Gα proteins differentially regulate Wnt/ß-catenin signaling but, importantly, are not required core components of Wnt/ß-catenin signaling. Our data suggest that activated Gα proteins are playing physiologically significant roles during both skeletal development and disease by modulating Wnt/ß-catenin signaling strength.
Assuntos
Displasia Fibrosa Óssea/metabolismo , Subunidades alfa de Proteínas de Ligação ao GTP/metabolismo , Via de Sinalização Wnt , Adulto , Animais , Células da Medula Óssea/patologia , Displasia Fibrosa Óssea/patologia , Displasia Fibrosa Poliostótica/metabolismo , Displasia Fibrosa Poliostótica/patologia , Humanos , Camundongos , Osteoblastos/metabolismo , Osteoblastos/patologia , Fenótipo , Células-Tronco/metabolismo , Células-Tronco/patologia , Células Estromais/metabolismo , Células Estromais/patologia , Regulação para Cima , beta Catenina/metabolismoRESUMO
Impaired functioning of pancreatic beta cells is a key hallmark of type 2 diabetes. beta cell function is modulated by the actions of different classes of heterotrimeric G proteins. The functional consequences of activating specific beta cell G protein signaling pathways in vivo are not well understood at present, primarily due to the fact that beta cell G protein-coupled receptors (GPCRs) are also expressed by many other tissues. To circumvent these difficulties, we developed a chemical-genetic approach that allows for the conditional and selective activation of specific beta cell G proteins in intact animals. Specifically, we created two lines of transgenic mice each of which expressed a specific designer GPCR in beta cells only. Importantly, the two designer receptors differed in their G protein-coupling properties (G(q/11) versus G(s)). They were unable to bind endogenous ligand(s), but could be efficiently activated by an otherwise pharmacologically inert compound (clozapine-N-oxide), leading to the conditional activation of either beta cell G(q/11) or G(s) G proteins. Here we report the findings that conditional and selective activation of beta cell G(q/11) signaling in vivo leads to striking increases in both first- and second-phase insulin release, greatly improved glucose tolerance in obese, insulin-resistant mice, and elevated beta cell mass, associated with pathway-specific alterations in islet gene expression levels. Selective stimulation of beta cell G(s) triggered qualitatively similar in vivo metabolic effects. Thus, this developed chemical-genetic strategy represents a powerful approach to study G protein regulation of beta cell function in vivo.
Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Regulação da Expressão Gênica/fisiologia , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/anatomia & histologia , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais/fisiologia , Animais , Células COS , Chlorocebus aethiops , Clozapina/análogos & derivados , Clozapina/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Teste de Tolerância a Glucose , Insulina/metabolismo , Secreção de Insulina , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Transgênicos , Ensaio Radioligante , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacosRESUMO
We are creating families of designer G protein-coupled receptors (GPCRs) to allow for precise spatiotemporal control of GPCR signaling in vivo. These engineered GPCRs, called receptors activated solely by synthetic ligands (RASSLs), are unresponsive to endogenous ligands but can be activated by nanomolar concentrations of pharmacologically inert, drug-like small molecules. Currently, RASSLs exist for the three major GPCR signaling pathways (G(s), G(i) and G(q)). We review these advances here to facilitate the use of these powerful and diverse tools.
Assuntos
Engenharia de Proteínas/métodos , Receptores Acoplados a Proteínas G/análise , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais , Animais , Evolução Molecular , Humanos , Ligantes , Ligação Proteica , Receptores Acoplados a Proteínas G/química , Receptores Acoplados a Proteínas G/genéticaRESUMO
We present an application of a quantitative systems pharmacology (QSP) model to support a regulatory decision, specifically in assessing the adequacy of the proposed dosing regimen. On January 23, 2015, the US Food and Drug Administration (FDA) approved Natpara (human parathyroid hormone (PTH)) to control hypocalcemia in patients with hypoparathyroidism. Clinical trial results indicated that although once-daily PTH reduced calcium and vitamin D dose requirement while maintaining the normocalcemia, the regimen was not adequate to control hypercalciuria. We hypothesized that the lack of control on urinary calcium excretion was due to the short half-life of PTH. The QSP model-based simulations indicated that a more frequent dosing regimen may provide better control on hypercalciuria while maintaining normocalcemia. A postmarketing trial was recommended to assess pharmacokinetics (PKs) and pharmacodynamics (PDs) of PTH dose and dosing regimen. Although other modeling approaches may be feasible, in this specific case, QSP model-based simulations fulfilled the information gap to support recommendations of this postmarketing trial.
Assuntos
Hipoparatireoidismo/tratamento farmacológico , Modelos Biológicos , Hormônio Paratireóideo/administração & dosagem , Cálcio/metabolismo , Relação Dose-Resposta a Droga , Esquema de Medicação , Humanos , Hipoparatireoidismo/metabolismo , Hormônio Paratireóideo/metabolismoRESUMO
AIMS/HYPOTHESIS: Partial lipodystrophy (PL) is most commonly characterized by loss of subcutaneous fat in the extremities with preservation of truncal fat and is associated with insulin resistance, diabetes and hyperlipidaemia. Recombinant human leptin (r-metHuLeptin) therapy has been shown to be effective in treating metabolic abnormalities associated with congenital or acquired generalized lipodystrophy and PL associated with lamin A/C (LMNA) gene mutations or highly active antiretroviral therapy (HAART). Our aim was to assess the effectiveness of leptin therapy in treating metabolic complications of PL associated with heterozygous peroxisome proliferator activated receptor gamma (PPARG) mutations. This is the first report to detail the clinical response of a patient with PL due to a PPARG mutation treated with r-metHuLeptin. METHODS: A 36-year-old female with PL associated with a heterozygous PPARG mutation complicated by poorly controlled diabetes and severe, refractory hypertriglyceridaemia was enrolled in a National Institutes of Health (NIH) protocol to evaluate the role of r-metHuLeptin in lipodystrophy. The patient received escalating doses of r-metHuLeptin until a dose 0.12 mg/kg/day was reached. Metabolic parameters, including serum chemistries, fasting blood glucose, glycated haemoglobin (HbA1c), lipid profile, an oral glucose tolerance test (OGTT), an insulin tolerance test (ITT), liver volume, percentage body fat and energy expenditure were followed at regular time intervals over 18 months of therapy. RESULTS: Eighteen months of r-MetHuLeptin therapy was associated with a marked improvement in glucose homeostasis as evidenced by normalization of the fasting blood glucose (baseline = 8.3 mmol/l; 18 months = 4.9 mmol/l), lowering of HbA1c (baseline = 9.9%; 18 months = 7.2%) and improved tolerance to an oral glucose load. In addition, a striking amelioration in the patient's refractory, severe hypertriglyceridaemia was observed (baseline = 21.15 mmol/l; 18 months = 5.96 mmol/l). CONCLUSION: r-MetHuLeptin is effective in treating metabolic complications associated with PL due to PPARG mutations. In the context of previously published work, our findings suggest that the response to r-MetHuLeptin is independent of the aetiology in lipodystrophy.
Assuntos
Leptina/análogos & derivados , Lipodistrofia/tratamento farmacológico , Lipodistrofia/genética , PPAR gama/genética , Adulto , Feminino , Humanos , Leptina/administração & dosagem , Resultado do TratamentoRESUMO
Under physiologic conditions, glucose plays a critical role in providing energy to the central nervous system. A precipitous drop in the availability of this substrate results in dramatic symptoms that signal a medical emergency and warrant immediate therapy aimed at restoring plasma glucose to normal levels. A systemic approach to the differential diagnosis is useful in identifying the cause of hypoglycemia. Once established, a specific and/or definitive intervention that addresses that underlying problem can be implemented. In most cases, this systemic approach to diagnosis and therapy is rewarded with a good outcome for the patient.
Assuntos
Hipoglicemia , Glicemia/metabolismo , Humanos , Hipoglicemia/sangue , Hipoglicemia/diagnóstico , Hipoglicemia/etiologia , Hipoglicemia/terapiaRESUMO
The Chennai Urban Population Study investigates a South Indian population with a high prevalence of cardiovascular disease associated with the metabolic syndrome (MS). The Ala54Thr polymorphism in the fatty acid-binding protein 2 (FABP2) gene as well as the T-455C and C-482T polymorphisms in the apolipoprotein C-III (APOC3) gene promoter have been associated with features of the MS in specific populations. This study evaluates in Asian-Indians the association between these polymorphisms with MS and dyslipidemia, defined according to National Cholesterol Education Program Adult Treatment Panel III. Allelic frequencies in 70 controls and 110 patients with diabetes from the Chennai Urban Population Study were 52.9% for FABP2 Thr54, 73.0% for APOC3 -482T, and 80.2% for APOC3 -455C. The polymorphisms were in agreement with Hardy-Weinberg equilibrium. Controls carrying FABP2 Thr54 were more likely to have MS than noncarriers (Fisher's exact test P = 0.031; odds ratio = 6.9 with a 95% confidence interval of 1.1, 43.9). Those carrying at least one polymorphic allele in both genes had a higher likelihood of having MS than wild type (Fisher's exact test P = 0.003; odds ratio = 12.1 with a 95% confidence interval of 1.88, 77.6). Dyslipidemia was associated with the polymorphism as well. The polymorphisms were not associated with MS in patients with diabetes. The association of the polymorphisms with MS and dyslipidemia could contribute to the high cardiovascular disease prevalence in this population.
Assuntos
Apolipoproteínas C/genética , Proteínas de Transporte/genética , Hiperlipidemias/genética , Síndrome Metabólica/genética , Polimorfismo Genético , Adulto , Idoso , Apolipoproteína C-III , Povo Asiático/etnologia , Proteínas de Ligação a Ácido Graxo , Feminino , Frequência do Gene , Predisposição Genética para Doença/etnologia , Humanos , Hiperlipidemias/etnologia , Índia/epidemiologia , Masculino , Síndrome Metabólica/etnologia , Pessoa de Meia-Idade , Prevalência , Regiões Promotoras Genéticas/genética , Fatores de RiscoRESUMO
CONTEXT: An end of fast insulin ≥ 3 µIU/mL and a proinsulin concentration ≥ 5 pmol/L have been suggested as useful cutoffs for the diagnosis of insulinoma. OBJECTIVE: The main objective was to evaluate the diagnostic performance of an end of fast insulin concentration ≥ 3 µIU/mL and an end of fast proinsulin concentration ≥ 5 pmol/L. DESIGN: The design was a case-control series. SETTING: The setting was a tertiary-care center. PATIENTS: Fifty-six subjects with a positive 48-hour supervised fast had an insulinoma between June 2000 and April 2011. During this same time period, a diagnosis of insulinoma was excluded in 29 subjects who underwent a supervised fast. INTERVENTION: 48-hour supervised fast. MAIN OUTCOME MEASURE: The main outcome measures were serum insulin concentration and plasma proinsulin concentration. RESULTS: Ninety-one percent of the patients with an insulinoma had a measured insulin concentration ≥5 µIU/mL at the end of fast. The sensitivity increased to 98% if the threshold to define inadequate insulin suppression was lowered to ≥3 µIU/mL. The median (interquartile range) end of fast proinsulin was 100 (53-270) pmol/L for cases and 6.8 (4.2-12.0) pmol/L for controls. An end of fast proinsulin value of ≥ 5 pmol/L could not distinguish cases from controls (59% false positive rate). All patients with an insulinoma (sensitivity 100%) and none of the control subject (specificity 100%) had end of fast proinsulin concentration ≥ 27 pmol/L. CONCLUSIONS: Using a current insulin assay 9% of insulinoma cases end the supervised fast with an insulin concentration below 5 µIU/mL. Inadequate insulin suppression defined using a threshold of ≥ 3 µIU/mL increases the sensitivity of the test. The value of the proinsulin test lies in its unique ability to distinguish cases from controls. A proinsulin concentration of ≥22 pmol/L best discriminates cases from controls. Reliance on an end of fast proinsulin cutoff value of 5 pmol/L does not augment sensitivity but greatly reduces specificity of the test.
Assuntos
Insulina/sangue , Insulinoma/diagnóstico , Neoplasias Pancreáticas/diagnóstico , Proinsulina/sangue , Estudos de Casos e Controles , Jejum , Feminino , Humanos , Insulinoma/sangue , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/sangue , Guias de Prática Clínica como Assunto , Estudos Retrospectivos , Sensibilidade e Especificidade , Sociedades Científicas , Centros de Atenção TerciáriaRESUMO
Type 2 diabetes (T2D) has emerged as a major threat to human health in most parts of the world. Therapeutic strategies aimed at improving pancreatic ß cell function are predicted to prove beneficial for the treatment of T2D. In the present study, we demonstrate that drug-mediated, chronic, and selective activation of ß cell G(q) signaling greatly improve ß cell function and glucose homeostasis in mice. These beneficial metabolic effects were accompanied by the enhanced expression of many genes critical for ß cell function, maintenance, and differentiation. By employing a combination of in vivo and in vitro approaches, we identified a novel ß cell pathway through which receptor-activated G(q) leads to the sequential activation of ERK1/2 and IRS2 signaling, thus triggering a series of events that greatly improve ß cell function. Importantly, we found that chronic stimulation of a designer G(q)-coupled receptor selectively expressed in ß cells prevented both streptozotocin-induced diabetes and the metabolic deficits associated with the consumption of a high-fat diet in mice. Since ß cells are endowed with numerous receptors that mediate their cellular effects via activation of G(q)-type G proteins, our findings provide a rational basis for the development of novel antidiabetic drugs targeting this class of receptors.
Assuntos
Clozapina/análogos & derivados , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/metabolismo , Hipoglicemiantes/farmacologia , Células Secretoras de Insulina/efeitos dos fármacos , Receptores Acoplados a Proteínas G/agonistas , Animais , Linhagem Celular Tumoral , Proliferação de Células , Clozapina/farmacologia , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Experimental/prevenção & controle , Avaliação Pré-Clínica de Medicamentos , Feminino , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/genética , Expressão Gênica/efeitos dos fármacos , Proteínas Substratos do Receptor de Insulina/genética , Proteínas Substratos do Receptor de Insulina/metabolismo , Proteínas Substratos do Receptor de Insulina/fisiologia , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/fisiologia , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Terapia de Alvo Molecular , Agonistas Muscarínicos/farmacologia , Engenharia de Proteínas , Receptor Muscarínico M3/agonistas , Receptor Muscarínico M3/fisiologia , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Proteínas Recombinantes/agonistas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismoRESUMO
Here, we describe a newly generated transgenic mouse in which the Gs DREADD (rM3Ds), an engineered G protein-coupled receptor, is selectively expressed in striatopallidal medium spiny neurons (MSNs). We first show that in vitro, rM3Ds can couple to Gαolf and induce cAMP accumulation in cultured neurons and HEK-T cells. The rM3Ds was then selectively and stably expressed in striatopallidal neurons by creating a transgenic mouse in which an adenosine2A (adora2a) receptor-containing bacterial artificial chromosome was employed to drive rM3Ds expression. In the adora2A-rM3Ds mouse, activation of rM3Ds by clozapine-N-oxide (CNO) induces DARPP-32 phosphorylation, consistent with the known consequence of activation of endogenous striatal Gαs-coupled GPCRs. We then tested whether CNO administration would produce behavioral responses associated with striatopallidal Gs signaling and in this regard CNO dose-dependently decreases spontaneous locomotor activity and inhibits novelty induced locomotor activity. Last, we show that CNO prevented behavioral sensitization to amphetamine and increased AMPAR/NMDAR ratios in transgene-expressing neurons of the nucleus accumbens shell. These studies demonstrate the utility of adora2a-rM3Ds transgenic mice for the selective and noninvasive modulation of Gαs signaling in specific neuronal populations in vivo.This unique tool provides a new resource for elucidating the roles of striatopallidal MSN Gαs signaling in other neurobehavioral contexts.
Assuntos
Corpo Estriado/citologia , AMP Cíclico/metabolismo , Subunidades alfa Gs de Proteínas de Ligação ao GTP/genética , Neurônios/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Muscarínicos/genética , Inibidores da Captação Adrenérgica/farmacologia , Anfetamina/farmacologia , Animais , Animais Recém-Nascidos , Clozapina/análogos & derivados , Clozapina/farmacologia , Cocaína/farmacologia , Inibidores da Captação de Dopamina/farmacologia , Fosfoproteína 32 Regulada por cAMP e Dopamina/metabolismo , Proteínas de Fluorescência Verde/genética , Locomoção/efeitos dos fármacos , Locomoção/genética , Proteínas Luminescentes/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fosforilação/efeitos dos fármacos , Fosforilação/genética , Receptor A2A de Adenosina/genética , Receptores de Dopamina D1/genética , Receptores de Dopamina D2/genética , Proteína Vermelha FluorescenteRESUMO
Muscarinic acetylcholine (ACh) receptors (mAChRs; M(1)-M(5)) regulate the activity of an extraordinarily large number of important physiological processes. We and others previously demonstrated that pancreatic ß-cells are endowed with M(3) mAChRs which are linked to G proteins of the G(q) family. The activation of these receptors by ACh or other muscarinic agonists leads to the augmentation of glucose-induced insulin release via multiple mechanisms. Interestingly, in humans, ACh acting on human ß-cell mAChRs is released from adjacent α-cells which express both choline acetyltransferase (ChAT) and the vesicular acetylcholine transporter (vAChT), indicative of the presence of a non-neuronal cholinergic system in human pancreatic islets. In order to shed light on the physiological roles of ß-cell M(3) receptors, we recently generated and analyzed various mutant mouse models. Specifically, we carried out studies with mice which overexpressed M(3) receptors or mutant M(3) receptors in pancreatic ß-cells or which selectively lacked M(3) receptors or M(3)-receptor-associated proteins in pancreatic ß-cells. Our findings indicate that ß-cell M(3) receptors play a key role in maintaining proper insulin release and whole body glucose homeostasis and that strategies aimed at enhancing signaling through ß-cell M(3) receptors may prove useful to improve ß-cell function for the treatment of type 2 diabetes (T2D).
Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Células Secretoras de Insulina/metabolismo , Receptor Muscarínico M3/metabolismo , Animais , Diabetes Mellitus Tipo 2/genética , Regulação da Expressão Gênica , Glucose/metabolismo , Humanos , Insulina/metabolismo , Camundongos , Mutação , Proteínas RGS/metabolismo , Receptor Muscarínico M3/genéticaRESUMO
Impaired function of pancreatic ß-cells is one of the hallmarks of type 2 diabetes. ß-cell function is regulated by the activity of many hormones and neurotransmitters, which bind to specific cell surface receptors. The M(3) muscarinic acetylcholine receptor (M3R) belongs to the superfamily of G protein-coupled receptors and, following ligand dependent activation, selectively activates G proteins of the G(q/11) family. Recent studies with M3R mutant mice strongly suggest that ß-cell M3Rs play a central role in promoting insulin release and maintaining correct glucose homeostasis. In this review, we highlight recent studies indicating that ß-cell M3Rs and components of downstream signaling pathways might represent promising new targets for the treatment of type 2 diabetes.
Assuntos
Células Secretoras de Insulina/fisiologia , Receptor Muscarínico M3/fisiologia , Animais , Glicemia/metabolismo , Proteínas de Ligação ao GTP/fisiologia , Homeostase/fisiologia , Humanos , Insulina/metabolismo , Secreção de Insulina , Células Secretoras de Insulina/química , Camundongos , Camundongos Knockout , Camundongos Mutantes , Mutação , Proteínas RGS/genética , Proteínas RGS/fisiologia , Receptor Muscarínico M3/genética , Transdução de Sinais/fisiologiaRESUMO
Insulinomas are rare neuroendocrine tumors of pancreatic islet cells that retain the ability to produce and secrete insulin. In contrast to normally differentiated ß-cells, insulinoma cells continue to secrete insulin and proinsulin at low blood glucose. This deregulated insulin secretion manifests clinically as fasting hypoglycemia. The molecular pathways that characterize normal insulin secretion and ß-cell growth are reviewed and contrasted to the biology of insulinomas. The second half of this review summarizes the clinical approach to the disorder. The diagnosis of insulinoma is established by demonstrating inappropriately high insulin levels with coincident hypoglycemia at the time of a supervised fast. Localization of insulinomas is challenging owing to their small size but should be attempted to maximize the chance for successful surgical resection and avoid risks associated with reoperation. In the majority of cases, successful surgical resection leads to lifelong cure.
RESUMO
Previous studies have shown that ß-cell M(3) muscarinic acetylcholine receptors (M3Rs) play a key role in maintaining blood glucose homeostasis by enhancing glucose-dependent insulin release. In this study, we tested the hypothesis that long-term, persistent activation of ß-cell M3Rs can improve glucose tolerance and ameliorate the metabolic deficits associated with the consumption of a high-fat diet. To achieve the selective and persistent activation of ß-cell M3Rs in vivo, we generated transgenic mice that expressed the Q490L mutant M3R in their pancreatic ß-cells (ß-M3-Q490L Tg mice). The Q490L point mutation is known to render the M3R constitutively active. The metabolic phenotypes of the transgenic mice were examined in several in vitro and in vivo metabolic tests. In the presence of 15 mm glucose and the absence of M3R ligands, isolated perifused islets prepared from ß-M3-Q490L Tg mice released considerably more insulin than wild-type control islets. This effect could be completely blocked by incubation of the transgenic islets with atropine (10 µm), an inverse muscarinic agonist, indicating that the Q490L mutant M3R exhibited ligand-independent signaling (constitutive activity) in mouse ß-cells. In vivo studies showed that ß-M3-Q490L Tg mice displayed greatly improved glucose tolerance and increased serum insulin levels as well as resistance to diet-induced glucose intolerance and hyperglycemia. These results suggest that chronic activation of ß-cell M3Rs may represent a useful approach to boost insulin output in the long-term treatment of type 2 diabetes.
Assuntos
Glicemia/metabolismo , Células Secretoras de Insulina/metabolismo , Receptor Muscarínico M3/metabolismo , Análise de Variância , Animais , Atropina/farmacologia , Glicemia/efeitos dos fármacos , Genótipo , Homeostase/efeitos dos fármacos , Insulina/sangue , Insulina/metabolismo , Secreção de Insulina , Células Secretoras de Insulina/efeitos dos fármacos , Camundongos , Camundongos Transgênicos , Antagonistas Muscarínicos/farmacologia , Fenótipo , Receptor Muscarínico M3/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
CONTEXT: Selective intraarterial calcium injection of the major pancreatic arteries with hepatic venous sampling [calcium arterial stimulation (CaStim)] has been used as a localizing tool for insulinomas at the National Institutes of Health (NIH) since 1989. The accuracy of this technique for localizing insulinomas was reported for all cases until 1996. OBJECTIVES: The aim of the study was to assess the accuracy and track record of the CaStim over time and in the context of evolving technology and to review issues related to result interpretation and procedure complications. CaStim was the only invasive preoperative localization modality used at our center. Endoscopic ultrasound (US) was not studied. DESIGN AND SETTING: We conducted a retrospective case review at a referral center. PATIENTS: Twenty-nine women and 16 men (mean age, 47 yr; range, 13-78) were diagnosed with an insulinoma from 1996-2008. INTERVENTION: A supervised fast was conducted to confirm the diagnosis of insulinoma. US, computed tomography (CT), magnetic resonance imaging (MRI), and CaStim were used as preoperative localization studies. Localization predicted by each preoperative test was compared to surgical localization for accuracy. MAIN OUTCOME: We measured the accuracy of US, CT, MRI, and CaStim for localization of insulinomas preoperatively. RESULTS: All 45 patients had surgically proven insulinomas. Thirty-eight of 45 (84%) localized to the correct anatomical region by CaStim. In five of 45 (11%) patients, the CaStim was falsely negative. Two of 45 (4%) had false-positive localizations. CONCLUSION: The CaStim has remained vastly superior to abdominal US, CT, or MRI over time as a preoperative localizing tool for insulinomas. The utility of the CaStim for this purpose and in this setting is thus validated.
Assuntos
Cálcio/farmacologia , Hipoglicemia/etiologia , Insulinoma/patologia , Neoplasias Pancreáticas/patologia , Adolescente , Adulto , Idoso , Cálcio/administração & dosagem , Jejum , Feminino , Veias Hepáticas , Humanos , Injeções Intra-Arteriais , Insulina/sangue , Insulinoma/cirurgia , Laparoscopia , Laparotomia , Masculino , Pessoa de Meia-Idade , Pancreatectomia , Neoplasias Pancreáticas/cirurgia , Adulto JovemRESUMO
OBJECTIVE: An increase in the rate of hepatic glucose production is the major determinant of fasting hyperglycemia in type 2 diabetes. A better understanding of the signaling pathways and molecules that regulate hepatic glucose metabolism is therefore of great clinical importance. Recent studies suggest that an increase in vagal outflow to the liver leads to decreased hepatic glucose production and reduced blood glucose levels. Since acetylcholine (ACh) is the major neurotransmitter of the vagus nerve and exerts its parasympathetic actions via activation of muscarinic ACh receptors (mAChRs), we examined the potential metabolic relevance of hepatocyte mAChRs. RESEARCH DESIGN AND METHODS: We initially demonstrated that the M(3) mAChR is the only mAChR subtype expressed by mouse liver/hepatocytes. To assess the physiological role of this receptor subtype in regulating hepatic glucose fluxes and glucose homeostasis in vivo, we used gene targeting and transgenic techniques to generate mutant mice lacking or overexpressing M(3) receptors in hepatocytes only. RESULTS: Strikingly, detailed in vivo phenotyping studies failed to reveal any significant metabolic differences between the M(3) receptor mutant mice and their control littermates, independent of whether the mice were fed regular or a high-fat diet. Moreover, the expression levels of genes for various key transcription factors, signaling molecules, and enzymes regulating hepatic glucose fluxes were not significantly altered in the M(3) receptor mutant mice. CONCLUSIONS: This rather surprising finding suggests that the pronounced metabolic effects mediated by activation of hepatic vagal nerves are mediated by noncholinergic signaling pathways.