Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 69
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Diabetologia ; 56(1): 22-30, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23011351

RESUMO

AIMS/HYPOTHESIS: Recent studies suggest that proton pump inhibitor treatment may increase insulin secretion and improve glucose metabolism in type 2 diabetes. In a randomised double-blind prospective placebo-controlled 2 × 2 factorial study, we examined the effect of esomeprazole on insulin secretion, HbA(1c) and cardiovascular risk factors in type 2 diabetes. METHODS: Forty-one patients with type 2 diabetes using dietary control or oral glucose-lowering treatment were randomised to receive add-on esomeprazole 40 mg (n = 20) or placebo (n = 21) for 12 weeks. Randomisation was carried out prior to inclusion on the basis of a computer-generated random-number list. The allocation sequence was concealed in sealed envelopes from the researcher enrolling and assessing participants. The study was undertaken at Steno Diabetes Center, Gentofte, Denmark. The primary outcome was change in AUC for insulin levels during a meal test. Secondary outcomes were the levels of HbA(1c) and biochemical markers of cardiovascular risk, including lipids, coagulation factors, inflammation markers, markers of endothelial function and 24 h ambulatory BP measurements. RESULTS: Forty-one participants were analysed. In the esomeprazole-treated group the AUC for insulin did not change (before vs after treatment: 28,049 ± 17,659 vs 27,270 ± 32,004 pmol/l × min (p = 0.838). In the placebo group AUC for insulin decreased from 27,392 ± 14,348 pmol/l × min to 22,938 ± 11,936 pmol/l × min (p = 0.002). Esomeprazole treatment (n = 20) caused a ninefold increase in the AUC for gastrin. HbA(1c) increased from 7.0 ± 0.6% (53 ± 5 mmol/mol) to 7.3 ± 0.8% (56 ± 6 mmol/mol) in the esomeprazole-treated group and from 7.0 ± 0.6% (53 ± 5 mmol/mol) to 7.4 ± 0.8% (57 ± 6 mmol/mol) in the placebo group (n = 21) (p for difference in change >0.05). Except for BP, there were no differences between the groups in the markers of cardiovascular risk (p > 0.05). Monitoring of 24 h ambulatory BP showed a significant decrease in daytime systolic BP, daytime diastolic BP and 24 h diastolic BP in the placebo group (p < 0.05). No change in BP was seen in the patients treated with esomeprazole. CONCLUSIONS/INTERPRETATION: Treatment with esomeprazole over 12 weeks did not improve insulin secretion, glycaemic control or cardiovascular disease biomarkers in patients with type 2 diabetes.


Assuntos
Doenças Cardiovasculares/prevenção & controle , Diabetes Mellitus Tipo 2/tratamento farmacológico , Esomeprazol/uso terapêutico , Hiperglicemia/prevenção & controle , Insulina/metabolismo , Inibidores da Bomba de Prótons/uso terapêutico , Idoso , Biomarcadores/sangue , Monitorização Ambulatorial da Pressão Arterial , Doenças Cardiovasculares/epidemiologia , Terapia Combinada , Dinamarca/epidemiologia , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/dietoterapia , Diabetes Mellitus Tipo 2/metabolismo , Método Duplo-Cego , Quimioterapia Combinada/efeitos adversos , Esomeprazol/administração & dosagem , Esomeprazol/efeitos adversos , Gastrinas/sangue , Gastrinas/metabolismo , Hemoglobina A Glicada/análise , Humanos , Hipertensão/prevenção & controle , Insulina/sangue , Secreção de Insulina , Masculino , Pessoa de Meia-Idade , Efeito Placebo , Inibidores da Bomba de Prótons/administração & dosagem , Inibidores da Bomba de Prótons/efeitos adversos , Fatores de Risco , Iogurte
2.
Diabetologia ; 54(6): 1379-87, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21347622

RESUMO

AIMS/HYPOTHESIS: The hormone glucagon-like peptide 1 (GLP-1) is released in response to a meal from the intestinal L-cells, where it is processed from proglucagon by the proconvertase (PC)1/3. In contrast, in the adult islets proglucagon is processed to glucagon by the PC2 enzyme. The aim of the study was to evaluate if, during the development of diabetes, alpha cells produce GLP-1 that, in turn, might trigger beta cell growth. METHODS: Beta cell mass, GLP-1 and insulin levels were measured in the gerbil Psammomys obesus (P. obesus), a rodent model of nutritionally induced diabetes. Furthermore, the presence of biologically active forms of GLP-1 and PC1/3 in alpha cells was demonstrated by immunofluorescence, and the release of GLP-1 from isolated P. obesus, mouse and human islets was investigated. RESULTS: During the development of diabetes in P. obesus, a significant increase in GLP-1 was detected in the portal vein (9.8 ± 1.5 vs 4.3 ± 0.7 pmol/l, p < 0.05), and in pancreas extracts (11.4 ± 2.2 vs 5.1 ± 1.3 pmol/g tissue, p < 0.05). Freshly isolated islets from hyperglycaemic animals released more GLP-1 following 24 h culture than islets from control animals (28.2 ± 4.4 pmol/l vs 5.8 ± 2.4, p < 0.01). GLP-1 release was increased from healthy P. obesus islets following culture in high glucose for 6 days (91 ± 9.1 pmol/l vs 28.8 ± 6.6, p < 0.01). High levels of GLP-1 were also found to be released from human islets. PC1/3 colocalised weakly with alpha cells. CONCLUSIONS/INTERPRETATION: GLP-1 release from alpha cells is upregulated in P. obesus during the development of diabetes. A similar response is seen in islets exposed to high glucose, which supports the hypothesis that GLP-1 released from alpha cells promotes an increase in beta cell mass and function during metabolic challenge such as diabetes.


Assuntos
Diabetes Mellitus/metabolismo , Gerbillinae/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Células Secretoras de Glucagon/metabolismo , Hiperglicemia/metabolismo , Obesidade/metabolismo , Regulação para Cima/fisiologia , Adaptação Fisiológica/fisiologia , Adulto , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Diabetes Mellitus/etiologia , Diabetes Mellitus/patologia , Dieta/efeitos adversos , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Células Secretoras de Glucagon/efeitos dos fármacos , Células Secretoras de Glucagon/patologia , Glucose/farmacologia , Humanos , Hiperglicemia/patologia , Insulina/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Obesidade/genética , Obesidade/patologia
3.
Immunol Lett ; 136(1): 74-9, 2011 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-21237203

RESUMO

We investigated the impact of ß-cell specific overexpression of suppressor of cytokine signalling-3 (SOCS-3) on the development of multiple low dose streptozotocin (MLDSTZ) induced Type 1 diabetes and the possible mechanisms involved. MLDSTZ treatment was administered to RIP-SOCS-3 transgenic and wild-type (wt) mice and progression of hyperglycemia monitored. Isolated islets from both strains were exposed to human IL-1ß (25U/ml) or a combination of human IL-1ß (25U/ml) and murine IFN-γ (1000U/ml) for 24h or 48h and we investigated the expression of IL-1 receptor antagonist (IL-1Ra) mRNA in islet cells and secretion of IL-1Ra into culture medium. MLDSTZ treatment caused gradual hyperglycemia both in the wt mice and in the transgenic mice with the latter tending to be more sensitive. In vitro experiments on wt and transgenic islets did not reveal any differences in sensitivity to damaging effects of STZ. Exposure of wt islets to IL-1ß or IL-1ß+IFN-γ seemed to lead to a failing IL-1Ra response from SOCS-3 transgenic islets. It could be that an increased expression of a possible protective molecule against ß-cell destruction may lead to a dampered response of another putative protective molecule. This may have counteracted a protective effect against MLDSTZ in SOCS-3 transgenic mice.


Assuntos
Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Células Secretoras de Insulina/metabolismo , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Animais , Células Cultivadas , Diabetes Mellitus Tipo 1/induzido quimicamente , Diabetes Mellitus Tipo 1/genética , Regulação da Expressão Gênica , Hiperglicemia/induzido quimicamente , Hiperglicemia/genética , Hiperglicemia/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor de Interferon alfa e beta/genética , Estreptozocina , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/genética
4.
Diabetologia ; 53(10): 2220-3, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20585936

RESUMO

AIMS/HYPOTHESIS: Gastrin has been implicated in islet growth/neogenesis, and proton pump inhibitors (PPIs) have been shown to increase endogenous gastrin levels in animals and humans. Therefore, we investigated the effect of PPIs in a model of type 2 diabetes, Psammomys obesus. METHODS: P. obesus (morning blood glucose [mBG] 16.9 +/- 0.6 mmol/l) were treated with vehicle or different doses (1-15 mg/kg) of lansoprazole for 17 days. RESULTS: Treatment with lansoprazole resulted in up to ninefold dose-dependent increases in endogenous gastrin levels (p < 0.05 for 10 mg/kg lansoprazole vs vehicle). There was a significant reduction in mBG levels in all animals in the high-dose lansoprazole groups during the 17 day treatment period, whereas there was no significant improvement in mBG in animals in the vehicle groups. The mBG at end of study was 18.2 +/- 2.1, 8.7 +/- 2.2 (p < 0.01), and 6.1 +/- 2.3 (p < 0.001) mmol/l for vehicle and lansoprazole 10 and 15 mg/kg, respectively. The animals treated with 15 mg/kg lansoprazole, compared with vehicle, had a 2.3-fold increase in the intensity of insulin staining in beta cells (p=0.0002) and 50% higher beta cell mass (p=0.04). CONCLUSIONS/INTERPRETATIONS: The PPI lansoprazole had significant glucose-lowering effects in an animal model of type 2 diabetes, an effect that is most likely mediated through an increase in endogenous gastrin levels.


Assuntos
2-Piridinilmetilsulfinilbenzimidazóis/uso terapêutico , Glicemia/efeitos dos fármacos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Hiperglicemia/tratamento farmacológico , Inibidores da Bomba de Prótons , 2-Piridinilmetilsulfinilbenzimidazóis/administração & dosagem , Análise de Variância , Animais , Glicemia/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/uso terapêutico , Feminino , Gastrinas/sangue , Gerbillinae , Imuno-Histoquímica , Insulina/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Lansoprazol , Masculino
5.
Diabetes Obes Metab ; 11(3): 196-203, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19215277

RESUMO

AIMS/HYPOTHESIS: The suppressor of cytokine signalling 1 (SOCS1) is a natural inhibitor of cytokine and insulin signalling pathways and may also play a role in obesity. In addition, SOCS1 is considered a candidate gene in the pathogenesis of both type 1 diabetes (T1D) and type 2 diabetes (T2D). The objective was to perform mutation analysis of SOCS1 and to test the identified variations for association to T2D-related quantitative traits, T2D or T1D. METHODS: Mutation scanning was performed by direct sequencing in 27 white Danish subjects. Genotyping was carried out by TaqMan allelic discrimination. A total of more than 8100 individuals were genotyped. RESULTS: Eight variations were identified in the 5' untranslated region (UTR) region. Two of these had allele frequencies below 1% and were not further examined. The six other variants were analysed in groups of T1D families (n = 1461 subjects) and T2D patients (n = 1430), glucose tolerant first-degree relatives of T2D patients (n = 212) and normal glucose tolerant (NGT) subjects. The rs33977706 polymorphism (-820G > T) was associated with a lower body mass index (BMI) (p = 0.004). In a second study (n = 4625 NGT subjects), significant associations of both the rs33977706 and the rs243330 (-1656G > A) variants to obesity were found (p = 0.047 and p = 0.015) respectively. The rs33977706 affected both binding of a nuclear protein to and the transcriptional activity of the SOCS1 promoter, indicating a relationship between this polymorphism and gene regulation. CONCLUSIONS/INTERPRETATION: This study demonstrates that functional variations in the SOCS1 promoter may associate with alterations in BMI in the general white population.


Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Grupo com Ancestrais do Continente Europeu/genética , Resistência à Insulina/genética , Obesidade/genética , Polimorfismo Genético/genética , Proteínas Supressoras da Sinalização de Citocina/genética , Adulto , Índice de Massa Corporal , Diabetes Mellitus Tipo 1/etnologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 2/etnologia , Diabetes Mellitus Tipo 2/genética , Feminino , Humanos , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Obesidade/metabolismo , Proteína 1 Supressora da Sinalização de Citocina , Proteínas Supressoras da Sinalização de Citocina/metabolismo
6.
Diabetologia ; 51(10): 1873-82, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18648765

RESUMO

AIMS/HYPOTHESIS: The pro-inflammatory cytokines IL-1 and IFNgamma are critical molecules in immune-mediated beta cell destruction leading to type 1 diabetes mellitus. Suppressor of cytokine signalling (SOCS)-3 inhibits the cytokine-mediated destruction of insulinoma-1 cells. Here we investigate the effect of SOCS3 in primary rodent beta cells and diabetic animal models. METHODS: Using mice with beta cell-specific Socs3 expression and a Socs3-encoding adenovirus construct, we characterised the protective effect of SOCS3 in mouse and rat islets subjected to cytokine stimulation. In transplantation studies of NOD mice and alloxan-treated mice the survival of Socs3 transgenic islets was investigated. RESULTS: Socs3 transgenic islets showed significant resistance to cytokine-induced apoptosis and impaired insulin release. Neither glucose-stimulated insulin release, insulin content or glucose oxidation were affected by SOCS3. Rat islet cultures transduced with Socs3-adenovirus displayed reduced cytokine-induced nitric oxide and apoptosis associated with inhibition of the IL-1-induced nuclear factor-kappaB and mitogen-activated protein kinase (MAPK) pathways. Transplanted Socs3 transgenic islets were not protected in diabetic NOD mice, but showed a prolonged graft survival when transplanted into diabetic allogenic BALB/c mice. CONCLUSIONS/INTERPRETATION: SOCS3 inhibits IL-1-induced signalling through the nuclear factor-kappaB and MAPK pathways and apoptosis induced by cytokines in primary beta cells. Moreover, Socs3 transgenic islets are protected in an allogenic transplantation model. SOCS3 may represent a target for pharmacological or genetic engineering in islet transplantation for treatment of type 1 diabetes mellitus.


Assuntos
Apoptose/fisiologia , Citocinas/metabolismo , Ilhotas Pancreáticas/metabolismo , Proteínas Supressoras da Sinalização de Citocina/fisiologia , Aloxano , Animais , Animais Recém-Nascidos , Apoptose/genética , Western Blotting , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/fisiopatologia , Sobrevivência de Enxerto/genética , Sobrevivência de Enxerto/fisiologia , Humanos , Marcação In Situ das Extremidades Cortadas , Ilhotas Pancreáticas/citologia , Transplante das Ilhotas Pancreáticas/métodos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/genética , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Transplante Homólogo
7.
Genes Immun ; 8(3): 232-8, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17330137

RESUMO

We recently finemapped a type 1 diabetes (T1D)-linked region on chromosome 21, indicating that one or more T1D-linked genes exist in this region with 33 annotated genes. In the current study, we have taken a novel approach using transcriptional profiling in predicting and prioritizing the most likely candidate genes influencing beta-cell function in this region. Two array-based approaches were used, a rat insulinoma cell line (INS-1alphabeta) overexpressing pancreatic duodenum homeobox 1 (pdx-1) and treated with interleukin 1beta (IL-1beta) as well as human pancreatic islets stimulated with a mixture of cytokines. Several candidate genes with likely functional significance in T1D were identified. Genes showing differential expression in the two approaches were highly similar, supporting the role of these specific gene products in cytokine-induced beta-cell damage. These were genes involved in cytokine signaling, oxidative phosphorylation, defense responses and apoptosis. The analyses, furthermore, revealed several transcription factor binding sites shared by the differentially expressed genes and by genes demonstrating highly similar expression profiles with these genes. Comparable findings in the rat beta-cell line and human islets support the validity of the methods used and support this as a valuable approach for gene mapping and identification of genes with potential functional significance in T1D, within a region of linkage.


Assuntos
Cromossomos Humanos Par 21/genética , Diabetes Mellitus Tipo 1/genética , Ilhotas Pancreáticas/metabolismo , Adolescente , Adulto , Animais , Linhagem Celular Tumoral , Criança , Feminino , Perfilação da Expressão Gênica , Predisposição Genética para Doença , Proteínas de Homeodomínio/genética , Humanos , Técnicas In Vitro , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Insulinoma/genética , Interleucina-1beta/farmacologia , Ilhotas Pancreáticas/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/genética , Ratos , Transativadores/genética
8.
Scand J Immunol ; 64(6): 639-45, 2006 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17083620

RESUMO

Secretagogin is a newly identified calcium-binding protein selectively expressed in neuroendocrine tissue and pancreatic beta-cells. The function of secretagogin is unknown, but it has been suggested in beta-cells to influence calcium-influx, insulin secretion and proliferation, and has been observed downregulated in diabetes-prone BB rat islets exposed to cytokines. In the present study, we identified and characterized promoter activity of a human 1498 bp sequence upstream the transcription start site. The promoter sequence showed subtle but significant regulation by glucose within the normo-physiological range. Glucose also led to changes in expression of secretagogin protein in INS-1e cells, but not in primary cells from non-diabetes-prone Wistar Furth rats. No effects of cytokines neither on promoter activity nor protein expression were observed. The promoter region was furthermore screened by direct sequencing, and 11 polymorphisms were identified. Genotyping in a large homogenous Type 1 diabetes (T1D) family collection did not reveal association with T1D.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Regulação da Expressão Gênica , Regiões Promotoras Genéticas/fisiologia , Regiões 5' não Traduzidas/genética , Regiões 5' não Traduzidas/fisiologia , Animais , Sequência de Bases , Proteínas de Ligação ao Cálcio/metabolismo , Linhagem Celular Tumoral , Citocinas/farmacologia , Diabetes Mellitus Tipo 1/genética , Glucose/farmacologia , Humanos , Ilhotas Pancreáticas/metabolismo , Dados de Sequência Molecular , Polimorfismo Genético , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Ratos , Secretagoguinas , Sítio de Iniciação de Transcrição
9.
Acta Diabetol ; 42(2): 95-8, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15944843

RESUMO

Type 1 diabetes (T1D) is an autoimmune disease in genetically predisposed individuals characterised by selective destruction of the beta-cells. Development of diabetes is in the asymptomatic pre-diabetic period characterised by impaired first-phase insulin response and the first clinical symptom is elevated blood glucose (BG). It is still uncertain whether stress or incidental hyperglycaemia can be regarded as predictors for development of T1D or not, even when immunologic and genetic markers for T1D are considered. The aim of this study was to investigate if there was any relationship between elevated BG in 30-day-old anaesthetised pre-diabetic diabetes-prone Bio Breeding (BB-DP) rats and later development of diabetes. Rats anaesthetised by intraperitoneal (ip) injection for islet transplantation displayed significantly higher BG values (Delta1.27 mmol/l, p=8.27x10(-12)) compared to non-anaesthetised non-transplanted rats, indicating that ip injection and/or anaesthesia induce a higher BG level. Linear regression analysis of BG and time of onset of diabetes in transplanted and non-transplanted BB-DP rats revealed no correlation (R(2) at 0.0075 and 0.0324 and p-values at 0.56 and 0.23 respectively). We were not able to identify any association or correlation between the induced temporary hyperglycaemia in 30-day-old BB-DP rats and later development of diabetes.


Assuntos
Anestesia/efeitos adversos , Glicemia/metabolismo , Diabetes Mellitus Tipo 1/fisiopatologia , Hiperglicemia/etiologia , Animais , Hiperglicemia/epidemiologia , Estado Pré-Diabético , Prevalência , Ratos , Ratos Endogâmicos BB
10.
Diabetologia ; 47(12): 2185-99, 2004 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-15605246

RESUMO

AIM/HYPOTHESIS: Maturation of the beta cells in the islets of Langerhans is dependent upon sequential activation of different transcription factors such as Pdx-1 and Nkx6.1. This maturation is associated with an acquired sensitivity to cytokines and may eventually lead to type 1 diabetes. The aims of this study were to characterise changes in mRNA expression during beta cell maturation as well as after interleukin-1beta (IL-1beta) exposure. METHODS: Transcriptome analyses were performed on two phenotypes characterised as a glucagon-producing pre-beta-cell phenotype (NHI-glu), which matures to an IL-1beta-sensitive insulin-producing beta cell phenotype (NHI-ins). Beta cell lines over-expressing Pdx-1 or Nkx6.1, respectively, were used for functional characterisation of acquired IL-1beta sensitivity. RESULTS: During beta cell maturation 98 fully annotated mRNAs changed expression levels. Of these, 50 were also changed after 24 h of IL-1beta exposure. In addition, 522 and 197 fully annotated mRNAs, not affected by maturation, also changed expression levels following IL-1beta exposure of the beta cell and the pre-beta-cell phenotype, respectively. Beta cell maturation was associated with an increased expression of Nkx6.1, whereas both Pdx-1 and Nkx6.1 expression were decreased following IL-1beta exposure. Over-expression of Nkx6.1 or Pdx-1 in cell lines resulted in a significantly increased sensitivity to IL-1beta. CONCLUSIONS/INTERPRETATION: These results suggest that the final beta cell maturation accompanied by increased IL-1beta sensitivity is, in part, dependent upon the expression of genes regulated by Pdx-1 and Nkx6.1. Future classification of the genes regulated by these transcription factors and changed during beta cell maturation should elucidate their role in the acquired sensitivity to IL-1beta and may be helpful in identifying new targets for intervention/prevention strategies.


Assuntos
Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Interleucina-1/farmacologia , Ilhotas Pancreáticas/fisiologia , Transativadores/genética , Animais , Sequência de Bases , Linhagem Celular , Primers do DNA , Ilhotas Pancreáticas/efeitos dos fármacos , Transplante das Ilhotas Pancreáticas , RNA Complementar/genética , RNA Mensageiro/genética , Ratos , Transcrição Genética
11.
Diabetologia ; 47(11): 1998-2011, 2004 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-15578154

RESUMO

AIMS/HYPOTHESIS: The proinflammatory cytokine IL-1beta induces apoptosis in pancreatic beta cells via pathways dependent on nuclear factor-kappaB (NF-kappaB), mitogen-activated protein kinase, and protein kinase C. We recently showed suppressor of cytokine signalling (SOCS)-3 to be a natural negative feedback regulator of IL-1beta- and IFN-gamma-mediated signalling in rat islets and beta cell lines, preventing their deleterious effects. However, the mechanisms underlying SOCS-3 inhibition of IL-1beta signalling and prevention against apoptosis remain unknown. METHODS: The effect of SOCS-3 expression on the global gene-expression profile following IL-1beta exposure was microarray-analysed using a rat beta cell line (INS-1) with inducible SOCS-3 expression. Subsequently, functional analyses were performed. RESULTS: Eighty-two known genes and several expressed sequence tags (ESTs) changed expression level 2.5-fold or more in response to IL-1beta alone. Following 6 h of IL-1beta exposure, 23 transcripts were up-regulated. Of these, several, including all eight transcripts relating to immune/inflammatory response pathways, were suppressed by SOCS-3. Following 24 h of IL-1beta exposure, secondary response genes were detected, affecting metabolism, energy generation, protein synthesis and degradation, growth arrest, and apoptosis. The majority of these changes were prevented by SOCS-3 expression. Multiple IL-1beta-induced NF-kappaB-dependent proapoptotic early response genes were inhibited by SOCS-3 expression, suggesting that SOCS-3 inhibits NF-kappaB-mediated signalling. These observations were experimentally confirmed in functional analyses. CONCLUSIONS/INTERPRETATION: This study suggests that there is an unexpected cross-talk between the SOCS/IFN and the IL-1beta pathways of signalling in pancreatic beta cells, which could lead to a novel perspective of blocking two important proapoptotic pathways in pancreatic beta cells by influencing a single signalling molecule, namely SOCS-3.


Assuntos
Apoptose/efeitos dos fármacos , Interleucina-1/toxicidade , NF-kappa B/fisiologia , Proteínas Repressoras/fisiologia , Transdução de Sinais/fisiologia , Fatores de Transcrição/fisiologia , Animais , Sequência de Bases , Linhagem Celular , Primers do DNA , Regulação da Expressão Gênica , Óxido Nítrico/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Ratos , Proteínas Repressoras/genética , Transdução de Sinais/efeitos dos fármacos , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina , Fatores de Transcrição/genética
12.
Diabetologia ; 47(7): 1273-1277, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15249995

RESUMO

AIMS/HYPOTHESIS: Beta cell loss in Type 1 and Type 2 diabetes mellitus may result from apoptosis and necrosis induced by inflammatory mediators. The suppressor of cytokine signalling (SOCS)-3 is a natural inhibitor of cytokine signalling and also influences insulin signalling. SOCS3 could therefore be a candidate gene in the development of Type 1 and Type 2 diabetes mellitus. METHODS: Mutation analysis of the SOCS3 gene was performed in 21 patients with Type 1 diabetes mellitus and in seven healthy subjects. An identified promoter variant was examined in (i) 250 families with Type 1 diabetic family members (1097 individuals); (ii) 212 glucose-tolerant first-degree relatives of Type 2 diabetic patients; and (iii) 370 population-based young, healthy subjects who were unrelated. RESULTS: Three mutations were identified in the promoter region, but none in the coding region or the 3'UTR. Two of the three mutations had allele frequencies below 1% whereas the C -920-->A substitution had a minor allele frequency of 8%. In the group of young healthy subjects the insulin sensitivity index was higher among homozygous carriers of the A-allele than among heterozygous and wild-type subjects ( p=0.027, uncorrected). The same trend was found in the group of first-degree relatives of Type 2 diabetic patients. No association or linkage was found to Type 1 diabetes mellitus. CONCLUSIONS/INTERPRETATION: Homozygosity for the A-allele of the C -920-->A promoter polymorphism of the SOCS3 gene may be associated with increased whole-body insulin sensitivity, but deserves further investigation.


Assuntos
Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 2/genética , Insulina/fisiologia , Polimorfismo de Nucleotídeo Único/genética , Proteínas Repressoras/genética , Fatores de Transcrição/genética , Adolescente , Adulto , Sequência de Bases , Criança , Pré-Escolar , Análise Mutacional de DNA , Primers do DNA , Família , Humanos , Lactente , Reação em Cadeia da Polimerase , Transdução de Sinais , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina
13.
Scand J Immunol ; 59(6): 582-91, 2004 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15182254

RESUMO

CD4 is a candidate gene in autoimmune diseases, including Type 1 diabetes mellitus (T1DM), because the CD4 receptor is crucial for appropriate antigen responses of CD4(+) T cells. We previously found linkage between a CD4-1188(TTTTC)(5-14) promoter polymorphism and T1DM. In the present study, we screened the human CD4 promoter for mutations and identified three frequent single nucleotide polymorphisms (SNPs): CD4-181C/G, CD4-521C/G and CD4-1050T/C. The SNPs are in strong linkage disequilibrium (LD) and association with the CD4-1188(TTTTC)(5-14) alleles, and we observed nine CD4 promoter haplotypes, of which four are frequent. We genotyped the SNPs in 253 Danish T1DM families (1129 individuals) and found evidence for linkage and association of a CD4 (A4(-1188)T(-1050)G(-521)C(-181)) haplotype to T1DM. In reporter studies, we show that (1) the T1DM-associated CD4 haplotype encodes high constitutive promoter activity and (2) the CD4-181G variant encodes higher stimulated promoter activity than the CD4-181C variant. This difference is in part neutralized in the frequently occurring CD4 promoter haplotypes by the more upstream genetic variants. Thus, we report functional impact of a novel CD4-181C/G SNP on stimulated CD4 promoter activity and the identification of a novel CD4 haplotype with high constitutive promoter activity that is linked and associated with T1DM.


Assuntos
Antígenos CD4/genética , Diabetes Mellitus Tipo 1/genética , Predisposição Genética para Doença , Regiões Promotoras Genéticas , Adolescente , Antígenos CD4/imunologia , Criança , Dinamarca , Diabetes Mellitus Tipo 1/imunologia , Feminino , Frequência do Gene , Haplótipos , Humanos , Desequilíbrio de Ligação/genética , Masculino , Mutação , Reação em Cadeia da Polimerase , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único/genética , Sequências de Repetição em Tandem/genética
14.
Diabetologia ; 47(5): 892-908, 2004 May.
Artigo em Inglês | MEDLINE | ID: mdl-15105991

RESUMO

AIMS/HYPOTHESIS: Type 1 diabetes mellitus is a multifactorial autoimmune disease characterised by selective destruction of beta cells in the islets of Langerhans. We have previously shown that IL-1 beta modulates beta cell function, causes beta cell death and induces expression changes in 82 out of 1815 protein spots detected by two-dimensional gel electrophoresis (2-DGE) in diabetes-prone bio-breeding (BB-DP) rat islets in vitro. The aim of this study was to describe the relevance of these proteins in the development of diabetes in vivo. METHODS: Syngeneic neonatal islets ( n=200) were transplanted under the kidney capsule of 30-day-old BB-DP and control rats, removed to different time points after transplantation or at the onset of diabetes, and metabolically labelled with S(35)-methionine for 2-DGE. The 82 proteins were re-localised and followed. In addition, transplants were examined for expression of IL-1 beta mRNA by in situ hybridisation. RESULTS: All 82 proteins could be re-localised in all syngeneic transplants from BB-DP and control rats. A total of 60 of the 82 proteins were changed during development of diabetes. Of the 82 proteins, 32 were changed in expression at the onset of diabetes compared to non-diabetic BB-DP rats, and 25 of these were changed as by IL-1 beta in vitro. Highest expression of IL-1 beta mRNA was found at the onset of diabetes. CONCLUSIONS/INTERPRETATION: IL-1 beta-induced protein expression changes in islets in vitro also occur in vivo and change in a complex pattern during the development of diabetes in the BB-DP rat. No single protein seems to be responsible for the development of diabetes, but rather the cumulative numbers of changes seem to interfere with the intracellular stability of the beta cell.


Assuntos
Diabetes Mellitus Tipo 1/genética , Regulação da Expressão Gênica/imunologia , Interleucina-1/genética , Animais , Separação Celular/métodos , Hibridização In Situ , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/imunologia , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos BB
15.
Diabetologia ; 47(1): 62-74, 2004 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-14652719

RESUMO

AIM/HYPOTHESIS: Type 1 diabetes mellitus (T1DM) is caused by specific destruction of the pancreatic beta cells in the islets of Langerhans. Increased sensitivity to cytokines, in particular to interleukin-1beta (IL-1beta) seems to be an acquired trait during beta-cell maturation. In response to cytokines both protective and deleterious mechanisms are induced in beta cells, and when the deleterious prevail, T1DM develops. The aims of this study were to identify perturbation in protein patterns (PiPP) associated with beta-cell maturation, and compare these changes to previous analyses of IL-1beta exposed rat islets. For this purpose, proteome analyses were carried out using a cell-line, which matures from a glucagon-producing pre-beta-cell phenotype (NHI-glu) to an insulin-producing beta-cell phenotype (NHI-ins). We have previously shown that this maturation is accompanied by acquired sensitivity to the toxic effects of IL-1beta. METHODS: 2D-gel electrophoresis was used to separate the proteins and MALDI-MS and database searches were performed to identify the proteins. RESULTS: During beta-cell maturation 135 protein spots out of 2239 detectable changed expression levels. Of these, 74 were down-regulated, 44 up-regulated, 16 were suppressed and 1 was expressed de novo. Using MALDI-MS, positive identification was obtained for 93 out of the 135 protein-spots revealing 97 different proteins. Of these, 22 proteins were in common with changes identified in previous proteome analysis of perturbation in protein pattern in IL-1beta exposed rat islets. Several of the proteins were present in more than one spot suggesting post-translational modification. CONCLUSION/INTERPRETATION: Several proteins and protein modifications were identified that could be critically involved in beta-cell maturation, insulin-gene expression and the acquired IL-1beta sensitivity.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Interleucina-1/farmacologia , Ilhotas Pancreáticas/fisiologia , Animais , Células Cultivadas , Eletroforese em Gel Bidimensional , Metabolismo Energético , Glutationa Transferase/metabolismo , Glicólise , Ilhotas Pancreáticas/efeitos dos fármacos , Metionina/metabolismo , Oxirredução , Biossíntese de Proteínas , Dobramento de Proteína , Proteínas/genética , Proteínas/isolamento & purificação , Ratos , Ratos Endogâmicos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
16.
Cytokine ; 24(1-2): 13-24, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-14561487

RESUMO

The stress-activated protein kinase c-Jun NH2-terminal kinase (JNK) is a central signal for interleukin-1beta (IL-1beta)-induced apoptosis in insulin-producing beta-cells. The cell-permeable peptide inhibitor of JNK (JNKI1), that introduces the JNK binding domain (JBD) of the scaffold protein islet-brain 1 (IB1) inside cells, effectively prevents beta-cell death caused by this cytokine. To define the molecular targets of JNK involved in cytokine-induced beta-cell apoptosis we investigated whether JNKI1 or stable expression of JBD affected the expression of selected pro- and anti-apoptotic genes induced in rat (RIN-5AH-T2B) and mouse (betaTC3) insulinoma cells exposed to IL-1beta. Inhibition of JNK significantly reduced phosphorylation of the specific JNK substrate c-Jun (p<0.05), IL-1beta-induced apoptosis (p<0.001), and IL-1beta-mediated c-fos gene expression. However, neither JNKI1 nor JBD did influence IL-1beta-induced NO synthesis or iNOS expression or the transcription of the genes encoding mitochondrial manganese superoxide dismutase (MnSOD), catalase (CAT), glutathione peroxidase (GPx), glutathione-S-transferase rho (GSTrho), heat shock protein (HSP) 70, IL-1beta-converting enzyme (ICE), caspase-3, apoptosis-inducing factor (AIF), Bcl-2 or Bcl-xL. We suggest that the anti-apoptotic effect of JNK inhibition by JBD is independent of the transcription of major pro- and anti-apoptotic genes, but may be exerted at the translational or posttranslational level.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Apoptose/fisiologia , Ilhotas Pancreáticas/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas Nucleares/metabolismo , Transativadores/metabolismo , Animais , Sítios de Ligação , Insulina/metabolismo , Interleucina-1/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno , Camundongos , Óxido Nítrico , Óxido Nítrico Sintase/metabolismo , Estrutura Terciária de Proteína , Ratos
17.
Diabetologia ; 45(11): 1550-61, 2002 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-12436339

RESUMO

AIMS/HYPOTHESIS: Type I (insulin-dependent) diabetes mellitus is characterized by selective destruction of the insulin producing beta cells. Interleukin-1beta (IL-1beta) modulates the beta-cell function, protein synthesis, energy production and causes apoptosis. We have previously shown changes in the expression of 82 out of 1 815 protein spots detected by two dimensional gel electrophoresis in IL-1beta exposed diabetes prone Bio Breeding (BB-DP) rat islets of Langerhans in vitro. The aim of this study was to identify the proteins in these 82 spots by mass spectrometry and compare these changes with those seen in IL-1beta exposed Wistar Furth (WF) rat islets. METHODS: The 82 protein spots, that changed expression after IL-1beta exposure, were all re-identified on preparative gels of 200 000 neonatal WF rat islets, cut out and subjected to mass spectrometry for identification. RESULTS: Forty-five different proteins were identified from 51 spots and grouped according to function: (i) energy transduction and redox potentials; (ii) glycolytic and Krebs cycle enzymes; (iii) protein, DNA and RNA synthesis, chaperoning and protein folding; (iv) signal transduction, regulation, differentiation and apoptosis; (v) cellular defence; and (vi) other functions. Comparison of IL-1beta exposed BB-DP and WF islets showed common changes in 14 proteins and several proteins influencing similar pathways, suggesting that similar routes in the two strains lead to beta-cell destruction. CONCLUSION/INTERPRETATION: We demonstrate that proteome analysis is a powerful tool to identify proteins and pathways in BB-DP rat islets exposed to IL-1beta.


Assuntos
Diabetes Mellitus Tipo 1/genética , Interleucina-1/farmacologia , Ilhotas Pancreáticas/fisiologia , Proteínas/genética , Proteoma , Animais , Animais Recém-Nascidos , Células Cultivadas , Eletroforese em Gel Bidimensional , Enzimas/genética , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Proteínas/isolamento & purificação , Ratos , Ratos Endogâmicos BB , Ratos Endogâmicos WF
18.
Genes Immun ; 2(7): 398-400, 2001 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-11704806

RESUMO

Accumulating evidence has suggested a role for the anti-apoptotic protein BCL2 in the development of autoimmune diseases, including type 1 diabetes mellitus (T1DM). Recently, the first BCL2 polymorphism (Ala43Thr) with association to T1DM in a Japanese population was reported. The polymorphism was found significantly more frequent in control individuals (14.5%) than in T1DM patients (6.8%), and was furthermore found to be functionally relevant, promoting a increased sensitivity to apoptosis when overexpressed in an IL-7 dependent mouse pre-B cell line. To investigate the relevance of the polymorphism in Caucasians, we have genotyped nearly 1400 individuals comprising Danish, Finnish and Basque T1DM family materials, using a PCR-based RFLP assay. In contrast to what was observed in Japanese diabetic/control individuals, we find no evidence for association of the BCL2 Ala43Thr polymorphism to T1DM in Danish, Finnish and Basque Type 1 diabetes families.


Assuntos
Diabetes Mellitus Tipo 1/genética , Genes bcl-2/genética , Predisposição Genética para Doença/genética , Polimorfismo Genético/genética , Apoptose , Dinamarca/epidemiologia , Diabetes Mellitus Tipo 1/epidemiologia , Diabetes Mellitus Tipo 1/patologia , Feminino , Finlândia/epidemiologia , Humanos , Masculino , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Espanha/epidemiologia
19.
Proc Natl Acad Sci U S A ; 98(21): 12191-6, 2001 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-11593036

RESUMO

Suppressor of cytokine signaling 3 (SOCS-3) is a negative feedback regulator of IFN-gamma signaling, shown up-regulated in mouse bone marrow cells by the proinflammatory cytokines interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), and IFN-gamma. IL-1beta and IFN-gamma alone, or potentiated by TNF-alpha, are cytotoxic to the insulin producing pancreatic beta-cells and beta-cell lines in vitro and suggested to contribute to the specific beta-cell destruction in Type-1 diabetes mellitus (T1DM). Using a doxycycline-inducible SOCS-3 expression system in the rat beta-cell line INS-1, we demonstrate that the toxic effect of both IL-1beta or IFN-gamma at concentrations that reduced the viability by 50% over 3 days, was fully preventable when SOCS-3 expression was turned on in the cells. At cytokine concentrations or combinations more toxic to the cells, SOCS-3 overexpression yielded a partial protection. Whereas SOCS-3-mediated inhibition of IFN-gamma signaling is described in other cell systems, SOCS-3 mediated inhibition of IL-1beta signaling has not previously been described. In addition we show that SOCS-3 prevention of IL-1beta-induced toxicity is accompanied by inhibited transcription of the inducible nitric oxide synthase (iNOS) by 80%, resulting in 60% decreased formation of the toxic nitric oxide (NO). Analysis of isolated native rat islets exposed to IL-1beta revealed a naturally occurring but delayed up-regulated SOCS-3 transcription. Influencing SOCS-3 expression thus represents an approach for affecting cytokine-induced signal transduction at a proximal step in the signal cascade, potentially useful in future therapies aimed at reducing the destructive potential of beta-cell cytotoxic cytokines in T1DM, as well as other cytokine-dependent diseases.


Assuntos
Interferon gama/farmacologia , Interleucina-1/farmacologia , Ilhotas Pancreáticas/efeitos dos fármacos , Proteínas/fisiologia , Proteínas Repressoras , Transdução de Sinais/fisiologia , Fatores de Transcrição , Animais , Apoptose , Linhagem Celular , Sobrevivência Celular , Células Cultivadas , Proteínas de Ligação a DNA/metabolismo , Expressão Gênica , Humanos , Ilhotas Pancreáticas/citologia , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase Tipo II , Regiões Promotoras Genéticas , Proteínas/genética , RNA Mensageiro , Ratos , Ratos Endogâmicos WF , Fator de Transcrição STAT1 , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina , Transativadores/metabolismo
20.
Eur Cytokine Netw ; 12(3): 501-9, 2001.
Artigo em Inglês | MEDLINE | ID: mdl-11566631

RESUMO

UNLABELLED: Nitric oxide (NO) may be a necessary but not sufficient mediator of cytokine-mediated, selective beta-cell destruction. Previously, we have described a difference in NO-dependent IL-1beta sensitivity in vivo and in vitro of pancreatic islets from two rat strains, Brown Norway (BN) and Wistar Kyoto (WK), the latter being the more sensitive strain. Here we investigated whether strain-dependent, differential islet iNOS expression was associated with differences in islet expression of the IL-1 receptor type 1(IL-1RI) or interferon regulating factor 1 (IRF-1), and/or caused differences in HSP70 expression, a marker of cell defence against oxidative stress. METHODS: isolated islets from both rat strains were exposed to increasing concentrations of IL-1beta (0-150 pg/ml) for 24 hours or for varying culture periods (4-48 hours) to 15 pg/ml of IL-1beta. MEASUREMENTS: accumulated insulin and nitrite release into incubation medium, and islet mRNA and protein expression of iNOS, IL-1RI, IRF-1 and HSP70 by semi-quantitative RT-PCR and Western blotting. RESULTS: Higher insulin and lower nitrite release into the incubation medium were seen for BN compared to WK rats islets in both dose- and time-response experiments. IRF-1 expression preceded iNOS expression and was more pronounced in WK than in BN islets. No strain differences were observed for islet expression of IL-1RI. A strain-dependent HSP70 expression in response to IL-1beta with the highest levels in WK rat islets following iNOS expression was seen. CONCLUSION: There was a strain-dependent difference in iNOS expression which was associated with IRF-1 and HSP70 expression.


Assuntos
Proteínas de Ligação a DNA/genética , Interleucina-1/metabolismo , Ilhotas Pancreáticas/metabolismo , Óxido Nítrico Sintase/genética , Fosfoproteínas/genética , RNA Mensageiro/análise , Receptores de Interleucina-1/genética , Animais , Proteínas de Ligação a DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Perfilação da Expressão Gênica , Proteínas de Choque Térmico HSP70/agonistas , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Técnicas In Vitro , Insulina/metabolismo , Secreção de Insulina , Fator Regulador 1 de Interferon , Interleucina-1/farmacologia , Ilhotas Pancreáticas/citologia , Óxido Nítrico Sintase/efeitos dos fármacos , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo II , Nitritos/metabolismo , Fosfoproteínas/efeitos dos fármacos , Proteínas de Protozoários/agonistas , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , RNA Mensageiro/efeitos dos fármacos , Ratos , Ratos Wistar , Receptores de Interleucina-1/efeitos dos fármacos , Especificidade da Espécie
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA