RESUMO
Islet cell surface autoantibodies were previously found in subjects with type 1 diabetes mellitus (T1DM), but their target antigens and pathogenic mechanisms remain elusive. The glutamate transporter solute carrier family 1, member 2 (GLT1/EAAT2) is expressed on the membrane of pancreatic ß-cells and physiologically controls extracellular glutamate concentrations thus preventing glutamate-induced ß-cell death. We hypothesized that GLT1 could be an immunological target in T1DM and that autoantibodies against GLT1 could be pathogenic. Immunoprecipitation and ELISA experiments showed that sera from T1DM subjects recognized GLT1 expressed in brain, pancreatic islets, and GLT1-transfected COS7-cell extracts. We validated these findings in two cohorts of T1DM patients by quantitative immunofluorescence assays. Analysis of the combined data sets indicated the presence of autoantibodies against GLT1 in 32 of the 87 (37%) T1DM subjects and in none of healthy controls (n = 64) (p < 0.0001). Exposure of pancreatic ßTC3 cells and human islets to purified IgGs from anti-GLT1 positive sera supplemented with complement resulted in plasma membrane ruffling, cell lysis and death. The cytotoxic effect was prevented when sera were depleted from IgGs. Furthermore, in the absence of complement, 6 out of 16 (37%) anti-GLT1 positive sera markedly reduced GLT1 transport activity in ßTC3 cells by inducing GLT1 internalization, also resulting in ß-cell death. In conclusion, we provide evidence that GLT1 is a novel T1DM autoantigen and that anti-GLT1 autoantibodies cause ß-cell death through complement-dependent and independent mechanisms. GLT1 seems an attractive novel therapeutic target for the prevention of ß-cell death in individuals with diabetes and prediabetes.
Assuntos
Sistema X-AG de Transporte de Aminoácidos , Diabetes Mellitus Tipo 1 , Autoanticorpos , Diabetes Mellitus Tipo 1/terapia , Ácido Glutâmico/metabolismo , Humanos , Neuroglia/metabolismoRESUMO
While a number of structural and cellular abnormalities occur in the islet of Langerhans in diabetes, and in particular in type 2 diabetes, the focus has been mostly on the insulin producing ß-cells and only more recently on glucagon producing α- and δ-cells. There is ample evidence that in type 2 diabetes mellitus (T2DM), in addition to a progressive decline in ß-cell function and associated insulin resistance in a number of insulin-sensitive tissues, alterations in glucagon secretion are also present and may play an important role in the pathogenesis of hyperglycemia both in the fasting and in the postprandial state. Recently, a number of studies have showed that there are also functional and structural alterations in glucagon-producing α-cells and somatostatin-producing δ-cells. Thus, it is becoming increasingly clear that multiple cellular alterations of multiple cell types occur, which adds even more complexity to our understanding of the pathophysiology of this common and severe disease. We believe that persistent efforts to increase the understanding of the pathophysiology of hormone secretion in the islets of Langerhans will also improve our capability to better prevent and treat diabetes mellitus.
Assuntos
Glicemia/metabolismo , Diabetes Mellitus Tipo 2/patologia , Ilhotas Pancreáticas/citologia , Amiloide/metabolismo , Animais , Células Secretoras de Glucagon/ultraestrutura , Haplorrinos , Humanos , Ilhotas Pancreáticas/ultraestrutura , Camundongos , Modelos Animais , Células Secretoras de Polipeptídeo Pancreático/ultraestrutura , Papio , Ratos , Células Secretoras de Somatostatina/ultraestruturaRESUMO
AIMS/HYPOTHESIS: Potentiation of glucose-induced insulin secretion is the main mechanism of exenatide (EXE) antidiabetic action, however, increased glucose utilization by peripheral tissues has been also reported. We here studied the effect of EXE on glucose uptake by skeletal muscle cells. METHODS: 2-deoxy-glucose (2DG) uptake and intracellular signal pathways were measured in rat L6 skeletal muscle myotubes exposed to 100 nmol/l EXE for up to 48 h. Mechanisms of EXE action were explored by inhibiting AMPK activity with compound C (CC, 40 µmol/l) or siRNAs (2 µmol/l). RESULTS: Time course experiments show that EXE increases glucose uptake up to 48 h achieving its maximal effect, similar to that induced by insulin, after 20 min (2- vs 2.5-fold-increase, respectively). Differently from insulin, EXE does not stimulate: (i) IR ß-subunit- and IRS1 tyrosine phosphorylation and binding to p85 regulatory subunit of PI-3kinase; (ii) AKT activation; and (iii) ERK1/2 and JNK1/2 phosphorylation. Conversely, EXE increases phosphorylation of α-subunit of AMPK at Thr172 by 2.5-fold (p < 0.01). Co-incubation of EXE and insulin does not induce additive effects on 2DG-uptake. Inhibition of AMPK with CC, and reduction of AMPK protein expression by siRNA, completely abolish EXE-induced 2DG-uptake. Liraglutide, another GLP-1 receptor agonist, also stimulates AMPK phosphorylation and 2DG-uptake. Moreover, EXE stimulates 2DG-uptake also by L6 myotubes rendered insulin-resistant with methylglyoxal. Finally, EXE also induces glucose transporter Glut-4 translocation to the plasma membrane. CONCLUSIONS/INTERPRETATION: In L6 myotubes, EXE and liraglutide increase glucose uptake in an insulin-independent manner by activating AMPK.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Glucose/metabolismo , Liraglutida/farmacologia , Peptídeos/farmacologia , Peçonhas/farmacologia , Animais , Transporte Biológico/efeitos dos fármacos , Linhagem Celular , Desoxiglucose/metabolismo , Ativação Enzimática/efeitos dos fármacos , Exenatida , Perfilação da Expressão Gênica , Inativação Gênica/efeitos dos fármacos , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Humanos , Resistência à Insulina , Camundongos , Fibras Musculares Esqueléticas/efeitos dos fármacos , Fibras Musculares Esqueléticas/enzimologia , Fibras Musculares Esqueléticas/metabolismo , Papio , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Aldeído Pirúvico/farmacologia , Ratos , Transdução de Sinais/efeitos dos fármacos , Fatores de TempoRESUMO
AIMS/HYPOTHESIS: The cellular composition of the islet of Langerhans is essential to ensure its physiological function. Morphophysiological islet abnormalities are present in type 2 diabetes but the relationship between fasting plasma glucose (FPG) and islet cell composition, particularly the role of delta cells, is unknown. We explored these questions in pancreases from baboons (Papio hamadryas) with FPG ranging from normal to type 2 diabetic values. METHODS: We measured the volumes of alpha, beta and delta cells and amyloid in pancreatic islets of 40 baboons (Group 1 [G1]: FPG < 4.44 mmol/l [n = 10]; G2: FPG = 4.44-5.26 mmol/l [n = 9]; G3: FPG = 5.27-6.94 mmol/l [n = 9]; G4: FPG > 6.94 mmol/l [n = 12]) and correlated islet composition with metabolic and hormonal variables. We also performed confocal microscopy including TUNEL, caspase-3, and anti-caspase cleavage product of cytokeratin 18 (M30) immunostaining, electron microscopy, and immuno-electron microscopy with anti-somatostatin antibodies in baboon pancreases. RESULTS: Amyloidosis preceded the decrease in beta cell volume. Alpha cell volume increased â¼ 50% in G3 and G4 (p < 0.05), while delta cell volume decreased in these groups by 31% and 39%, respectively (p < 0.05). In G4, glucagon levels were higher, while insulin and HOMA index of beta cell function were lower than in the other groups. Immunostaining of G4 pancreatic sections with TUNEL, caspase-3 and M30 showed apoptosis of beta and delta cells, which was also confirmed by immuno-electron microscopy with anti-somatostatin antibodies. CONCLUSIONS/INTERPRETATION: In diabetic baboons, changes in islet composition correlate with amyloid deposition, with increased alpha cell and decreased beta and delta cell volume and number due to apoptosis. These data argue for an important role of delta cells in type 2 diabetes.
Assuntos
Morte Celular , Diabetes Mellitus Tipo 2/patologia , Resistência à Insulina/fisiologia , Ilhotas Pancreáticas/patologia , Células Secretoras de Somatostatina/patologia , Animais , Glicemia/metabolismo , Caspase 3/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Progressão da Doença , Feminino , Células Secretoras de Glucagon/metabolismo , Células Secretoras de Glucagon/patologia , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Ilhotas Pancreáticas/metabolismo , Masculino , Papio hamadryas , Células Secretoras de Somatostatina/metabolismoRESUMO
Glutamate is the major excitatory neurotransmitter of the central nervous system (CNS) and may induce cytotoxicity through persistent activation of glutamate receptors and oxidative stress. Its extracellular concentration is maintained at physiological concentrations by high affinity glutamate transporters of the solute carrier 1 family (SLC1). Glutamate is also present in islet of Langerhans where it is secreted by the α-cells and acts as a signaling molecule to modulate hormone secretion. Whether glutamate plays a role in islet cell viability is presently unknown. We demonstrate that chronic exposure to glutamate exerts a cytotoxic effect in clonal ß-cell lines and human islet ß-cells but not in α-cells. In human islets, glutamate-induced ß-cell cytotoxicity was associated with increased oxidative stress and led to apoptosis and autophagy. We also provide evidence that the key regulator of extracellular islet glutamate concentration is the glial glutamate transporter 1 (GLT1). GLT1 localizes to the plasma membrane of ß-cells, modulates hormone secretion, and prevents glutamate-induced cytotoxicity as shown by the fact that its down-regulation induced ß-cell death, whereas GLT1 up-regulation promoted ß-cell survival. In conclusion, the present study identifies GLT1 as a new player in glutamate homeostasis and signaling in the islet of Langerhans and demonstrates that ß-cells critically depend on its activity to control extracellular glutamate levels and cellular integrity.
Assuntos
Transportador 2 de Aminoácido Excitatório/biossíntese , Regulação da Expressão Gênica , Proteínas de Transporte de Glutamato da Membrana Plasmática/biossíntese , Células Secretoras de Insulina/citologia , Animais , Apoptose , Autofagia , Sobrevivência Celular , Transportador 2 de Aminoácido Excitatório/fisiologia , Proteínas de Transporte de Glutamato da Membrana Plasmática/fisiologia , Ácido Glutâmico/química , Ácido Glutâmico/metabolismo , Homeostase , Humanos , Ilhotas Pancreáticas/citologia , Camundongos , Modelos Biológicos , Estresse OxidativoRESUMO
beta-Cell dysfunction is an important factor in the development of hyperglycemia of type-2 diabetes mellitus, and pancreatic islet amyloidosis (IA) has been postulated to be one of the main contributors to impaired insulin secretion. The aim of this study was to evaluate the correlation of IA with metabolic parameters and its effect on islets of Langerhans remodeling and relative endocrine-cell volume in baboons. We sequenced the amylin peptide, determined the fibrillogenic propensities, and evaluated pancreatic histology, clinical and biochemical characteristics, and endocrine cell proliferation and apoptosis in 150 baboons with different metabolic status. Amylin sequence in the baboon was 92% similar to humans and showed superimposable fibrillogenic propensities. IA severity correlated with fasting plasma glucose (FPG) (r = 0.662, P < 0.001) and HbA1c (r = 0.726, P < 0.001), as well as with free fatty acid, glucagon values, decreased homeostasis model assessment (HOMA) insulin resistance, and HOMA-B. IA severity was associated with a decreased relative beta-cell volume, and increased relative alpha-cell volume and hyperglucagonemia. These results strongly support the concept that IA and beta-cell apoptosis in concert with alpha-cell proliferation and hypertrophy are key determinants of islets of Langerhans "dysfunctional remodeling" and hyperglycemia in the baboon, a nonhuman primate model of type-2 diabetes mellitus. The most important determinants of IA were age and FPG (R(2) = 0.519, P < 0.0001), and different FPG levels were sensitive and specific to predict IA severity. Finally, a predictive model for islet amyloid severity was generated with age and FPG as required variables.
Assuntos
Amiloidose/patologia , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Células Secretoras de Glucagon/metabolismo , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/patologia , Amiloide/metabolismo , Animais , Apoptose , Glicemia/metabolismo , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/patologia , Ácidos Graxos/metabolismo , Feminino , Resistência à Insulina , Polipeptídeo Amiloide das Ilhotas Pancreáticas , Masculino , PapioRESUMO
The glucagon-like peptide-1 receptor agonist exenatide improves glycemic control by several and not completely understood mechanisms. Herein, we examined the effects of chronic intravenous exenatide infusion on insulin sensitivity, ß cell and α cell function and relative volumes, and islet cell apoptosis and replication in nondiabetic nonhuman primates (baboons). At baseline, baboons received a 2-step hyperglycemic clamp followed by an l-arginine bolus (HC/A). After HC/A, baboons underwent a partial pancreatectomy (tail removal) and received a continuous exenatide (n = 12) or saline (n = 12) infusion for 13 weeks. At the end of treatment, HC/A was repeated, and the remnant pancreas (head-body) was harvested. Insulin sensitivity increased dramatically after exenatide treatment and was accompanied by a decrease in insulin and C-peptide secretion, while the insulin secretion/insulin resistance (disposition) index increased by about 2-fold. ß, α, and δ cell relative volumes in exenatide-treated baboons were significantly increased compared with saline-treated controls, primarily as the result of increased islet cell replication. Features of cellular stress and secretory dysfunction were present in islets of saline-treated baboons and absent in islets of exenatide-treated baboons. In conclusion, chronic administration of exenatide exerts proliferative and cytoprotective effects on ß, α, and δ cells and produces a robust increase in insulin sensitivity in nonhuman primates.
Assuntos
Diabetes Mellitus Tipo 2/tratamento farmacológico , Exenatida/farmacologia , Hipoglicemiantes/farmacologia , Resistência à Insulina , Ilhotas Pancreáticas/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Glicemia/análise , Proliferação de Células/efeitos dos fármacos , Transdiferenciação Celular/efeitos dos fármacos , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/patologia , Modelos Animais de Doenças , Exenatida/uso terapêutico , Feminino , Técnica Clamp de Glucose , Humanos , Hipoglicemiantes/uso terapêutico , Infusões Intravenosas , Insulina/metabolismo , Ilhotas Pancreáticas/patologia , Masculino , PapioRESUMO
To learn more about nonimmune-mediated islet graft failure, we transplanted different preparations (preps) of isolated human islets under the kidney capsule of streptozotocin (STZ)-diabetic nude mice. One month after the implantation of 1,000 or 2,000 islets, grafts were harvested for morphological, immunohistochemical, and ultrastructural analysis. Only a single islet prep cured the diabetes out of all the recipients, while the remaining preps showed only partial function after the implantation of 2,000 islets. Transplanted mice showed high circulating proinsulin levels but, with the exclusion of those bearing curative grafts, relatively low mature insulin levels. Engrafted beta-cells showed positive carboxypeptidase E (CPE) and prohormone convertase 1 (PC1) staining, while prohormone convertase 2 (PC2) was undetectable. In contrast, PC2 was abundantly expressed by engrafted alpha-cells. Moreover, engrafted beta-cells did not show evidence of replication, and preapoptotic beta-cells, with intra- and extracellular amyloid deposition, were detected with electron microscopy. Cell cycle inhibitors p16(INK4), p21(WAF1), and p27(Kip1) were abundantly expressed in the islet grafts and showed a predominant nuclear localization. In conclusion, diabetic nude mice transplanted with human islets showed disproportionate hyperproinsulinemia and graft evidence of beta-cell restricted PC2 depletion, amyloid deposition and beta-cell death, and lack of beta-cell replication with nuclear translocation of p27(Kip1) and p21(WAF1) that together may contribute to delayed graft failure.
Assuntos
Ciclo Celular/fisiologia , Diabetes Mellitus Experimental/cirurgia , Hiperinsulinismo/etiologia , Transplante das Ilhotas Pancreáticas/efeitos adversos , Ilhotas Pancreáticas/enzimologia , Ilhotas Pancreáticas/metabolismo , Proinsulina/metabolismo , Pró-Proteína Convertase 2/deficiência , Transplante Heterólogo/efeitos adversos , Animais , Autopsia , Humanos , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/enzimologia , Células Secretoras de Insulina/patologia , Ilhotas Pancreáticas/patologia , Transplante das Ilhotas Pancreáticas/patologia , Masculino , Camundongos , Camundongos Nus , Pâncreas/patologia , Proinsulina/sangue , Valores de Referência , Falha de TratamentoRESUMO
We have previously reported that in tumorigenic pancreatic beta-cells, calcitriol exerts a potent antitumorigenic effect by inducing apoptosis, cell growth inhibition, and reduction of solid beta-cell tumors. Here we have studied the molecular pathways involved in the antineoplastic activity of calcitriol on mouse insulinoma beta TC(3) cells, mouse insulinoma beta TC expressing or not expressing the oncogene p53, and beta TC-tet cells overexpressing or not the antiapoptotic gene Bcl2. Our results indicate that calcitriol-induced apoptosis was dependent on the function of p53 and was associated with a biphasic increase in protein levels of transcription factor nuclear factor-kappa B. Calcitriol decreased cell viability by about 40% in p53-retaining beta TC and in beta TC(3) cells; in contrast, beta TC p53(-/-) cells were only minimally affected. Calcitriol-induced cell death was regulated by members of the Bcl-2 family of apoptosis regulatory proteins, as shown by calcitriol-induced up-regulation of proapoptotic Bax and Bak and the lack of calcitriol-induced cytotoxicity in Bcl-2-overexpressing insulinoma cells. Moreover, calcitriol-mediated arrest of beta TC(3) cells in the G(1) phase of the cell cycle was associated with the abnormal expression of p21 and G(2)/M-specific cyclin B2 genes and involved the DNA damage-inducible factor GADD45. Finally, in beta TC(3) cells, calcitriol modulated the expression of IGF-I and IGF-II genes. In conclusion, these findings contribute to the understanding of the antitumorigenic effects of calcitriol on tumorigenic pancreatic beta-cells and further support the rationale of its utilization in the treatment of patients with malignant insulinomas.
Assuntos
Antineoplásicos/farmacologia , Calcitriol/farmacologia , Insulinoma/metabolismo , Neoplasias Pancreáticas/metabolismo , Prolina/análogos & derivados , Animais , Apoptose/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21 , Ciclinas/metabolismo , Resistência a Medicamentos , Expressão Gênica/efeitos dos fármacos , Proteínas I-kappa B/metabolismo , Insulinoma/genética , Insulinoma/fisiopatologia , Peptídeos e Proteínas de Sinalização Intracelular , Camundongos , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/fisiopatologia , Fosforilação , Prolina/farmacologia , Proteínas/genética , Tiocarbamatos/farmacologia , Fatores de Tempo , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/metabolismo , Proteínas GADD45RESUMO
Malignant insulinoma is a rare form of cancer with a poor prognosis because of metastatic dissemination and untreatable hypoglycemia. Effective chemotherapy of patients who are not cured by surgery is needed. Calcitriol has known anticancer properties on different neoplastic cell lines, but no data are available regarding its activity on tumorigenic pancreatic beta-cells. We analyzed the in vitro effects of calcitriol on the murine insulinoma cell line betaTC(3) and primary cultures of human isolated islets and benign insulinoma. The effect of in vivo calcitriol administration on insulinoma of recombinant insulin/Simian virus 40 oncogene-expressing transgenic mice was also investigated. In betaTC(3), calcitriol induced growth inhibition; apoptosis; down-regulation of insulin gene expression; and nongenomic activation of the MAPK pathway. MAPK kinase inhibitor (UO126) and staurosporine reduced calcitriol-mediated betaTC(3) death, and down-regulation of insulin gene transcription was prevented by staurosporine but not UO126. Calcitriol significantly decreased insulin release and mRNA levels of human islets and insulinoma cells. Finally, recombinant insulin/Simian virus 40 oncogene-expressing transgenic mice treated with calcitriol showed reduced insulinoma volumes because of increased apoptosis of adenomatous cells. Together, these findings provide the rationale for testing the efficacy of calcitriol in the treatment of patients with solid beta-cell tumors.
Assuntos
Antineoplásicos/farmacologia , Calcitriol/farmacologia , Antagonistas da Insulina/farmacologia , Insulinoma/metabolismo , Insulinoma/patologia , Ilhotas Pancreáticas , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Animais , Apoptose , Divisão Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Expressão Gênica/efeitos dos fármacos , Humanos , Insulina/genética , Insulina/metabolismo , Insulinoma/fisiopatologia , Ilhotas Pancreáticas/enzimologia , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/fisiopatologia , Camundongos , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Neoplasias Pancreáticas/fisiopatologia , Fosforilação/efeitos dos fármacos , Proteína Quinase C/antagonistas & inibidoresRESUMO
We recently developed two rat pituitary GH3 cell clones engineered to secrete human insulin (InsGH3). InsGH3 cells convert proinsulin into mature insulin, which is partially stored into a readily releasable pool of secretory granules. The efficiency of these processes, however, is relatively low in these cells, either in vitro or in vivo. This study was aimed at determining whether carboxypeptidase E (Cpe) overexpression can increase proinsulin processing and regulated secretion by InsGH3 clones. Indeed, in its membrane-bound form Cpe works as sorting receptor for the regulated secretory pathway of many hormones while, in its soluble form, Cpe takes part to the late step of insulin maturation. We obtained two Cpe-overexpressing cell lines from two different InsGH3 clones (InsGH3/C1 and C7). In the Cpe-overexpressing cell lines, derived from InsGH3 of clone 1 (InsGH3/C1-HACpe), in which the membrane-bound form of exogenous Cpe is accounted for by 90% of total Cpe immunoreactivity, we observed an increase in proinsulin gene expression, and in basal and stimulated insulin secretion compared with the original clone. In contrast, in the Cpe-overexpressing cell line derived from InsGH3 of clone 7 (InsGH3/C7-HACpe), where the exogenous membrane-bound form was only 60% of total Cpe, we detected a decrease in basal insulin release and a modest, albeit significant, increase in intracellular proinsulin processing. In conclusion, Cpe overexpression can increase regulated insulin secretion and proinsulin processing in InsGH3 cells; however, such improvements appear quantitatively and qualitatively modest.
Assuntos
Carboxipeptidases/genética , Transplante de Células/métodos , Insulina/genética , Hipófise/citologia , Proinsulina/metabolismo , Animais , Carboxipeptidase H , Linhagem Celular , Colforsina/farmacologia , DNA Complementar , Furina , Regulação Enzimológica da Expressão Gênica , Imuno-Histoquímica , Insulina/metabolismo , Secreção de Insulina , Hipófise/fisiologia , RNA Mensageiro/análise , Ratos , Subtilisinas , Engenharia Tecidual , TransfecçãoRESUMO
Insulin-deficient rats are characterized by multiple defects in the pathway of glycogen synthesis and breakdown in both liver and skeletal muscle. The aim of this study was to clarify whether islet transplantation under the kidney capsule, which is associated with delivery of insulin into the peripheral circulation, is able to normalize glycogen metabolism in liver and muscle of streptozotocin-diabetic rats. Three groups of male Lewis rats were studied under fasting condition: controls, untreated diabetics, and islet transplanted diabetics. Glycogen content, glucose-6-phosphate concentration, and glycogen synthase activity were measured in both liver and skeletal muscle. Untreated diabetic rats were characterized by an increase in glycogen content of 178% and a reduction of glucose-6-phosphate level of 50%. Both glycogen and glucose-6-phosphate contents were restored to normal in transplanted diabetic rats. Active glycogen synthase (0.35 +/- 0.1 nmol/min/mg) and activity ratio (0.22 +/- 0.04) were significantly impaired compared with controls (0.99 +/- 0.2 nmol/min/mg and 0.43 +/- 0.06, respectively) and were normalized by islet transplantation. In the skeletal muscle, glycogen content was similar in the three groups of animals, whereas muscle glucose-6-phosphate level was reduced by 28% and glycogen synthase was in a less active state in the untreated diabetic rats. Both the glucose-6-phosphate concentration and the kinetic profile of glycogen synthase were normalized by islet transplantation. In conclusion, islet transplantation under the kidney capsule corrects the diabetes-induced abnormalities in glycogen and glucose-6-phosphate content and glycogen synthase activity in both liver and skeletal muscle.
Assuntos
Diabetes Mellitus Experimental/cirurgia , Glicogênio/metabolismo , Transplante das Ilhotas Pancreáticas/métodos , Transplante das Ilhotas Pancreáticas/tendências , Ilhotas Pancreáticas/metabolismo , Rim/cirurgia , Fígado/metabolismo , Músculo Esquelético/metabolismo , Animais , Glicemia/metabolismo , Glucose-6-Fosfato/metabolismo , Glicogênio Sintase/metabolismo , Insulina/metabolismo , Secreção de Insulina , Transplante das Ilhotas Pancreáticas/normas , Rim/fisiologia , Fígado/fisiopatologia , Masculino , Músculo Esquelético/fisiopatologia , Ratos , Ratos Endogâmicos LewRESUMO
INTRODUCTION: The purpose of this study was to compare and validate the use of SenseWear Armband (SWA) placed on the arm (SWA ARM) and on the back (SWA BACK) in healthy humans during resting and a cycle-ergometer exercise and to evaluate the SWA to estimate Resting Energy Expenditure (REE) and Total Energy Expenditure (TEE) in healthy baboons. METHODS: We studied 26 (15F/11M) human subjects wearing SWA in two different anatomical sites (arm and back) during resting and a cycle-ergometer test and directly compared these results with indirect calorimetry evaluation (IC), performed at the same time. We then inserted the SWA in a metabolic jacket for baboons and evaluated the TEE and REE in free living condition for 6 days in 21 (8F/13M) non-human primates. RESULTS: In humans we found a good correlation between SWA place on the ARM and on the BACK with IC during the resting experiment (1.1±0.3 SWAs, 1±0.2 IC kcal/min) and a slight underestimation in the SWAs data compared with IC during the cycle-ergometer exercise (5±1.9 SWA ARM, 4.5±1.5 SWA BACK and 5.4±2.1 IC kcal/min). In the non-human primate (baboons) experiment SWA estimated a TEE of 0.54±0.009 kcal/min during free living and a REE of 0.82±0.06 kcal/min. CONCLUSION: SWA, an extremely simple and inexpensive apparatus, provides quite accurate measurements of energy expenditure in humans and in baboons. Energy expenditure data obtained with SWA are highly correlated with the data obtained with "gold standard", IC, in humans.
Assuntos
Calorimetria Indireta/métodos , Metabolismo Energético/fisiologia , Adulto , Idoso , Animais , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeAssuntos
Fibromatose Agressiva/tratamento farmacológico , Resistência à Insulina , Piperazinas/uso terapêutico , Pirimidinas/uso terapêutico , Anemia/induzido quimicamente , Benzamidas , Feminino , Fibromatose Agressiva/patologia , Fibromatose Agressiva/fisiopatologia , Humanos , Hipoglicemiantes/uso terapêutico , Mesilato de Imatinib , Insulina/uso terapêutico , Leucopenia/induzido quimicamente , Pessoa de Meia-Idade , Piperazinas/efeitos adversos , Inibidores de Proteínas Quinases/efeitos adversos , Inibidores de Proteínas Quinases/uso terapêutico , Pirimidinas/efeitos adversos , Resultado do TratamentoRESUMO
We report a case of unusually long-lasting remission of type 1 diabetes (T1D). The patient, a Caucasian man, at the age of 43 years developed a ketotic diabetes, classified as type 1 based on clinical presentation and positivity for islet autoantibodies. Shortly after diabetes onset, oral topiramate was added to preexisting valproic acid for generalized seizures and maintained thereafter. Initial intensive insulin treatment was rapidly reduced to low doses (3 Units/day) maintained for a long time and then discontinued at month 55; fasting glucose and glycosylated hemoglobin were basically normalized at 58 months. An oral glucose tolerance test performed at month 53 showed an impaired fasting glucose (6.0 mmol/l) and a value slightly above the threshold for the diagnosis of diabetes at 2 h (11.2 mmol/l). We hypothesize that this unusually prolonged preservation of ß-cell function might be ascribed to the concomitant therapy with topiramate, an antiepileptic agent with demonstrated efficacy as antidiabetic in type 2 diabetes (T2D). Topiramate should be further investigated as candidate agent for the preservation of ß-cell function also in T1D.
Assuntos
Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 1/tratamento farmacológico , Frutose/análogos & derivados , Convulsões/complicações , Convulsões/tratamento farmacológico , Adulto , Anticonvulsivantes/uso terapêutico , Diabetes Mellitus Tipo 1/reabilitação , Frutose/uso terapêutico , Humanos , Masculino , Indução de Remissão , TopiramatoRESUMO
In the present article, we propose the perspective that abnormal glutamate homeostasis might contribute to diabetes pathogenesis. Previous reports and our recent data indicate that chronically high extracellular glutamate levels exert direct and indirect effects that might participate in the progressive loss of ß-cells occurring in both T1D and T2D. In addition, abnormal glutamate homeostasis may impact all the three accelerators of the "accelerator hypothesis" and could partially explain the rising frequency of T1D and T2D.
Assuntos
Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Ácido Glutâmico/metabolismo , Células Secretoras de Insulina/metabolismo , Diabetes Mellitus Tipo 1/epidemiologia , Diabetes Mellitus Tipo 2/epidemiologia , Epidemias , Homeostase/fisiologia , HumanosRESUMO
BACKGROUND: The reasons for the long-term complete or partial loss of islet graft function are unknown, but there are obviously other reasons than just pure allogeneic graft rejection. Earlier studies have shown that deposition of islet amyloid polypeptide amyloid in transplanted islets may indicate a mechanism for loss of ß cells. MATERIALS AND METHODS: Sections from liver material from four deceased islet-bearing recipients have been scrutinized for the presence of amyloid. Clinical data and certain aspects of the islet graft pathology of these patients have been published previously. RESULT: With this extended histological analysis, we demonstrate the occurrence of amyloid deposits in islets transplanted into the liver in three of four patients with type 1 diabetes. CONCLUSION: The finding adds evidence to the assumption that aggregation of islet amyloid polypeptide might be an important cause of progressing ß-cell dysfunction in clinically transplanted islets.
Assuntos
Polipeptídeo Amiloide das Ilhotas Pancreáticas/metabolismo , Transplante das Ilhotas Pancreáticas/efeitos adversos , Adulto , Diabetes Mellitus Tipo 1/cirurgia , Evolução Fatal , Feminino , Humanos , Imuno-Histoquímica , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Transplante das Ilhotas Pancreáticas/mortalidade , Transplante das Ilhotas Pancreáticas/patologia , Transplante das Ilhotas Pancreáticas/fisiologia , Fígado/metabolismo , Fígado/patologia , Masculino , Pessoa de Meia-Idade , Pró-Proteína Convertase 2/metabolismo , Doadores de TecidosRESUMO
UNLABELLED: Non-alcoholic fatty liver disease (NAFLD) is characterized by accumulation of triglycerides (TG) in hepatocytes, which may also trigger cirrhosis. The mechanisms of NAFLD are not fully understood, but insulin resistance has been proposed as a key determinant. AIMS: To determine the TG content and long chain fatty acyl CoA composition profile in liver from obese non-diabetic insulin resistant (IR) and lean insulin sensitive (IS) baboons in relation with hepatic and peripheral insulin sensitivity. METHODS: Twenty baboons with varying grades of adiposity were studied. Hepatic (liver) and peripheral (mainly muscle) insulin sensitivity was measured with a euglycemic clamp and QUICKI. Liver biopsies were performed at baseline for TG content and LCFA profile by mass spectrometry, and histological analysis. Findings were correlated with clinical and biochemical markers of adiposity and insulin resistance. RESULTS: Obese IR baboons had elevated liver TG content compared to IS. Furthermore, the concentration of unsaturated (LC-UFA) was greater than saturated (LC-SFA) fatty acyl CoA in the liver. Interestingly, LC-FA UFA and SFA correlated with waist, BMI, insulin, NEFA, TG, QUICKI, but not M/I. Histological findings of NAFLD ranging from focal to diffuse hepatic steatosis were found in obese IR baboons. CONCLUSION: Liver TG content is closely related with both hepatic and peripheral IR, whereas liver LC-UFA and LC-SFA are closely related only with hepatic IR in non-human primates. Mechanisms leading to the accumulation of TG, LC-UFA and an altered UFA: LC-SFA ratio may play an important role in the pathophysiology of fatty liver disease in humans.
Assuntos
Ácidos Graxos/metabolismo , Resistência à Insulina , Fígado/metabolismo , Triglicerídeos/metabolismo , Acil Coenzima A/metabolismo , Adiposidade , Animais , Ácidos Graxos Insaturados/metabolismo , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Feminino , Glucose/metabolismo , Técnica Clamp de Glucose , Humanos , Modelos Lineares , Fígado/patologia , Masculino , Hepatopatia Gordurosa não Alcoólica , Papio , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
Intra-graft infiltrating cells apoptosis was evaluated in 20 consecutive kidney-pancreas transplanted (KP) patients without kidney rejection. Two fine-needle aspirated biopsy (FNAB) and two peripheral blood lymphocytes (PBL) samples were obtained 14 days after transplantation. Immunosuppression was based on anti-Thymoglobulins (ATG) induction for 7 days and cyclosporine/mofetil mycophenolate as maintenance therapy. Ten matched healthy subjects were chosen as controls for PBL. Lymphocyte phenotypes and activation markers, apoptotic rate and lymphocyte expression of pro/anti-apoptotic molecules were analysed by flow cytometry analysis (FACS). Lymphocyte phenotypes and activation markers: higher levels of CD8 and CD4DR were evident in the graft (p < 0.05) than in PBL, CD3CD25 in PBL were higher in transplanted patients than in controls. Apoptotic rate and lymphocyte expression of pro- and anti-apoptotic molecules: a higher expression of annexin V, together with reduced lymphocytes CD95L, iNOs and Bcl-2 expression (PBL = 97.7+/-1.1% vs FNAB = 81.9+/-15.1%; p < 0.05) were evident in the graft than in PBL. In KP patients intra-graft apoptosis and reduced anti-apoptotic molecules were evident after ATG induction.