RESUMO
The reactions of intestinal functional parameters to type 2 diabetes at a young age remain unclear. The study aimed to assess changes in the activity of intestinal enzymes, glucose absorption, transporter content (SGLT1, GLUT2) and intestinal structure in young Wistar rats with type 2 diabetes (T2D) and impaired glucose tolerance (IGT). To induce these conditions in the T2D (n = 4) and IGT (n = 6) rats, we used a high-fat diet and a low dose of streptozotocin. Rats fed a high-fat diet (HFD) (n = 6) or a standard diet (SCD) (n = 6) were used as controls. The results showed that in T2D rats, the ability of the small intestine to absorb glucose was higher in comparison to HFD rats (p < 0.05). This was accompanied by a tendency towards an increase in the number of enterocytes on the villi of the small intestine in the absence of changes in the content of SGLT1 and GLUT2 in the brush border membrane of the enterocytes. T2D rats also showed lower maltase and alkaline phosphatase (AP) activity in the jejunal mucosa compared to the IGT rats (p < 0.05) and lower AP activity in the colon contents compared to the HFD (p < 0.05) and IGT (p < 0.05) rats. Thus, this study provides insights into the adaptation of the functional and structural parameters of the small intestine in the development of type 2 diabetes and impaired glucose tolerance in young representatives.
Assuntos
Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Tipo 2/complicações , Intolerância à Glucose/complicações , Glucose/farmacocinética , Intestino Delgado/enzimologia , Intestino Delgado/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Enterócitos/química , Glucana 1,4-alfa-Glucosidase/metabolismo , Transportador de Glucose Tipo 2/análise , Absorção Intestinal , Masculino , Ratos , Ratos Wistar , Transportador 1 de Glucose-Sódio/análise , alfa-Glucosidases/metabolismoRESUMO
Protozoan parasites of the genus Leishmania are causative agents of leishmaniasis, a wide range of diseases affecting 12 million people worldwide. The species L. infantum and L. amazonensis are etiologic agents of visceral and cutaneous leishmaniasis, respectively. Most proteome analyses of Leishmania have been carried out on whole-cell extracts, but such an approach tends to underrepresent membrane-associated proteins due to their high hydrophobicity and low solubility. Considering the relevance of this category of proteins in virulence, invasiveness and the host-parasite interface, this study applied label-free proteomics to assess the plasma membrane sub-proteome of L. infantum and L. amazonensis. The number of proteins identified in L. infantum and L. amazonensis promastigotes was 1168 and 1455, respectively. After rigorous data processing and mining, 157 proteins were classified as putative plasma membrane-associated proteins, of which 56 proteins were detected in both species, six proteins were detected only in L. infantum and 39 proteins were exclusive to L. amazonensis. The quantitative analysis revealed that two proteins were more abundant in L. infantum, including the glucose transporter 2, and five proteins were more abundant in L. amazonensis. The identified proteins associated with distinct processes and functions. In this regard, proteins of L. infantum were linked to metabolic processes whereas L. amazonensis proteins were involved in signal transduction. Moreover, transmembrane transport was a significant process among the group of proteins detected in both species and members of the superfamily of ABC transporters were highly represented. Interestingly, some proteins of this family were solely detected in L. amazonensis, such as ABCA9. GP63, a well-known virulence factor, was the only GPI-anchored protein identified in the membrane preparations of both species. Finally, we found several proteins with uncharacterized functions, including differentially abundant ones, highlighting a gap in the study of Leishmania proteins. Proteins characterization could provide a better biological understanding of these parasites and deliver new possibilities regarding the discovery of therapeutic targets, drug resistance and vaccine candidates.
Assuntos
Leishmania infantum/química , Leishmania mexicana/química , Proteínas de Membrana/análise , Proteômica/métodos , Proteínas de Protozoários/análise , Animais , Membrana Celular/química , Cromatografia Líquida , Biologia Computacional , Cricetinae , Transportador de Glucose Tipo 2/análise , Interações Hospedeiro-Parasita , Leishmania infantum/metabolismo , Leishmania infantum/patogenicidade , Leishmania infantum/ultraestrutura , Leishmania mexicana/ultraestrutura , Macrófagos Peritoneais/parasitologia , Espectrometria de Massas , Mesocricetus , Metaloendopeptidases/análise , Camundongos , Camundongos Endogâmicos BALB C , Transdução de Sinais , Espectrometria de Massas em Tandem , VirulênciaRESUMO
Liver plays a central role in modulating blood glucose level. Our most recent findings suggested that supplementation with microbiota metabolite sodium butyrate (NaB) could ameliorate progression of type 2 diabetes mellitus (T2DM) and decrease blood HbA1c in db/db mice. To further investigate the role of butyrate in homeostasis of blood glucose and glycogen metabolism, we carried out the present study. In db/db mice, we found significant hypertrophy and steatosis in hepatic lobules accompanied by reduced glycogen storage, and expression of GPR43 was significantly decreased by 59.38 ± 3.33%; NaB administration significantly increased NaB receptor G-protein coupled receptor 43 (GPR43) level and increased glycogen storage in both mice and HepG2 cells. Glucose transporter 2 (GLUT2) and sodium-glucose cotransporter 1 (SGLT1) on cell membrane were upregulated by NaB. The activation of intracellular signaling Protein kinase B (PKB), also known as AKT, was inhibited while glycogen synthase kinase 3 (GSK3) was activated by NaB in both in vivo and in vitro studies. The present study demonstrated that microbiota metabolite NaB possessed beneficial effects on preserving blood glucose homeostasis by promoting glycogen metabolism in liver cells, and the GPR43-AKT-GSK3 signaling pathway should contribute to this effect.
Assuntos
Ácido Butírico/administração & dosagem , Diabetes Mellitus Tipo 2/metabolismo , Glicogênio Hepático/metabolismo , Animais , Glicemia/análise , Ácido Butírico/metabolismo , Imunofluorescência , Microbioma Gastrointestinal/fisiologia , Transportador de Glucose Tipo 2/análise , Hemoglobinas Glicadas/análise , Quinase 3 da Glicogênio Sintase/metabolismo , Células Hep G2 , Homeostase/efeitos dos fármacos , Humanos , Fígado/química , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Acoplados a Proteínas G/análise , Transdução de Sinais/efeitos dos fármacos , Transportador 1 de Glucose-Sódio/análiseRESUMO
SCOPE: (-)-Epicatechin (EC) and main colonic phenolic acids derived from flavonoid intake, such as 2,3-dihydroxybenzoic acid (DHBA), 3,4-dihydroxyphenylacetic acid (DHPAA), 3-hydroxyphenylpropionic acid (HPPA), and vanillic acid (VA), have been suggested to exert beneficial effects in diabetes, although the mechanism for their actions remains unknown. In this study, the modulation of glucose homeostasis and insulin signaling by the mentioned compounds on renal proximal tubular NRK-52E cells is investigated. METHODS AND RESULTS: Levels of the glucose transporters SGLT-2 and GLUT-2, as well as glucose uptake, glucose production, and key proteins of the insulin pathways, namely insulin receptor (IR), insulin receptor substrate-1 (IRS-1), and PI3K/AKT pathway are analyzed. EC (5-20 µm) and DHBA (20 µm) reduced both renal glucose uptake and production. Interestingly, EC and DHBA did not modify the levels of SGLT-2 and GLUT-2, and modulated the expression of phosphoenolpyruvate carboxykinase via AKT leading to a diminished glucose production. EC and DHBA also enhanced the tyrosine phosphorylation and total IR and IRS-1 levels, and activated the PI3K/AKT pathway in NRK-52E cells. CONCLUSION: EC and DHBA regulate the renal glucose homeostasis by modulating both glucose uptake and production, and strengthen the insulin signaling by activating key proteins of that pathway in NRK-52E cells.
Assuntos
Catequina/farmacologia , Colo/metabolismo , Glucose/metabolismo , Hidroxibenzoatos/farmacologia , Insulina/fisiologia , Rim/metabolismo , Transdução de Sinais/fisiologia , Animais , Células Cultivadas , Gluconeogênese/efeitos dos fármacos , Transportador de Glucose Tipo 2/análise , Quinase 3 da Glicogênio Sintase/fisiologia , Hidroxibenzoatos/metabolismo , Rim/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , RatosRESUMO
The assessment of the ß-cell mass in experimental models of diabetes and ultimately in patients is a hallmark to understand the relationship between reduced ß-cell mass/function and the onset of diabetes. It has been shown before that the GLUT-2 transporter is highly expressed in both ß-cells and hepatocytes and that D-mannoheptulose (DMH) has high uptake specificity for the GLUT-2 transporter. As 19-fluorine MRI has emerged as a new alternative method for MRI cell tracking because it provides potential non-invasive localization and quantification of labeled cells, the purpose of this project is to validate ß-cell and pancreatic islet imaging by using fluorinated, GLUT-2 targeting mannoheptulose derivatives (19 FMH) both in vivo and ex vivo. In this study, we confirmed that, similar to DMH, 19 FMHs inhibit insulin secretion and increase the blood glucose level in mice temporarily (approximately two hours). We were able to assess the distribution of 19 FMHs in vivo with a temporal resolution of about 20 minutes, which showed a quick removal of 19 FMH from the circulation (within two hours). Ex vivo MR spectroscopy confirmed a preferential uptake of 19 FMH in tissue with high expression of the GLUT-2 transporter, such as liver, endocrine pancreas and kidney. No indication of further metabolism was found. In summary, 19 FMHs are potentially suitable for visualizing and tracking of GLUT-2 expressed cells. However, current bottlenecks of this technique related to the quick clearance of the compound and relative low sensitivity of 19 F MRI need to be overcome. Copyright © 2016 John Wiley & Sons, Ltd.
Assuntos
Meios de Contraste/química , Flúor , Transportador de Glucose Tipo 2/análise , Células Secretoras de Insulina/citologia , Ilhotas Pancreáticas/citologia , Imageamento por Ressonância Magnética/métodos , Animais , Transportador de Glucose Tipo 2/metabolismo , Células Secretoras de Insulina/química , Ilhotas Pancreáticas/química , Rim/química , Rim/metabolismo , Fígado/metabolismo , Espectroscopia de Ressonância Magnética , Manoeptulose/metabolismo , Manoeptulose/farmacocinética , Camundongos , Imagem Molecular/métodos , Pâncreas/metabolismoRESUMO
SCOPE: We aimed at investigating the mechanisms linking hyperlipidemia (HL) with dysfunctional HDL and its main antioxidant enzyme, paraoxonase1 (PON1). PON1 expression and activity was determined in the small intestine, liver, and sera of normal and HL hamsters and associated with the ER stress (ERS) and the development of aortic valve lesions. METHODS AND RESULTS: Male Golden Syrian hamsters were fed standard chow (N) or standard diet with 3% cholesterol and 15% butter for 16 weeks. All hamsters on fat diet developed HL, 50% also hyperglycemia (HLHG) and a fourfold increased homeostasis model assessment of insuline resistance. PON1 expression was reduced in the small intestine and liver (N > HL > HLHG) along with the increased extent of ERS, oxidized lipids, and decreased expression of liver X receptors beta (LXRß) in the small intestine, peroxisome proliferator-activated receptor-γ (PPARγ) in the liver, and of the glucose transporter 4 in the myocardium. Serum PON1 levels decreased along with the increase of oxidized LDL and lesion areas of the aortic valves (N > HL > HLHG). CONCLUSION: The fat diet activates the ERS and oxidative stress, decreases LXRß, PPARγ, and PON1 in the small intestine, liver, and sera of all HL animals, in parallel with the appearance of atherosclerotic lesions in the aortic valves.
Assuntos
Arildialquilfosfatase/metabolismo , Estresse do Retículo Endoplasmático , Hiperlipidemias/metabolismo , Intestino Delgado/metabolismo , Lipoproteínas HDL/fisiologia , Fígado/metabolismo , Animais , Arildialquilfosfatase/genética , Cricetinae , Dieta Hiperlipídica , Transportador de Glucose Tipo 2/análise , Masculino , Mesocricetus , Miocárdio/metabolismoRESUMO
GLUT2 is a facilitative glucose transporter, expressed in polarized epithelial cells of the liver, intestine, kidney and pancreas, where it plays a critical role in glucose homeostasis. Together with SGLT1/2, it mediates glucose absorption in metabolic epithelial tissues, where it can be translocated apically upon high glucose exposure. To track the subcellular localization and dynamics of GLUT2, we created an mCherry-hGLUT2 fusion protein and expressed it in multicellular kidney cysts, a major site of glucose reabsorption. Live imaging of GLUT2 enabled us to avoid the artefactual localization of GLUT2 in fixed cells and to confirm the apical GLUT2 model. Live cell imaging showed a rapid 15 ± 3 min PKC-dependent basal-to-apical translocation of GLUT2 in response to glucose stimulation and a fourfold slower basolateral translocation under starvation. These results mark the physiological importance of responding quickly to rising glucose levels. Importantly, we show that phloretin, an apple polyphenol, inhibits GLUT2 translocation in both directions, suggesting that it exerts its effect by PKC inhibition. Subcellular localization studies demonstrated that GLUT2 is endocytosed through a caveolae-dependent mechanism, and that it is at least partly recovered in Rab11A-positive recycling endosome. Our work illuminates GLUT2 dynamics, providing a platform for drug development for diabetes and hyperglycaemia.
Assuntos
Células Epiteliais/citologia , Transportador de Glucose Tipo 2/análise , Transportador de Glucose Tipo 2/metabolismo , Glucose/metabolismo , Rim/citologia , Imagem Óptica/métodos , Animais , Linhagem Celular , Cães , Endocitose , Células Epiteliais/metabolismo , Transportador de Glucose Tipo 2/genética , Humanos , Rim/metabolismo , Proteínas Luminescentes/análise , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Microscopia de Fluorescência/métodos , Transporte Proteico , Proteínas Recombinantes de Fusão/análise , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteína Vermelha FluorescenteRESUMO
To elucidate the molecular mechanisms underlying the insufficient sensitivity in the detection of hepatocellular carcinoma (HCC) by [18F] 2-ï¬uoro-2-deoxy-D-glucose positron emission tomography (FDG-PET), the characteristics of glucose metabolism-related protein expression in HCC were examined in liver metastasis from colorectal cancer (Meta). Thirty-four patients (14 Meta and 20 HCC) who underwent FDG-PET and hepatectomy were studied. The relationships between the maximum standardized uptake value (SUV) in tumors and the mRNA expression of glucose metabolism-related proteins [hexokinase (HK), glucose transporter 1 (GLUT1), and glucose-6-phosphatase (G6Pase)] and proliferating cell nuclear antigen (PCNA) were examined in snap-frozen specimens with quantitative PCR. Tumor detection rates were lower in HCC (15/20) compared to Meta (13/14) patients. HK and GLUT1 expression was lower and G6Pase expression was higher in HCC compared to Meta. In particular, GLUT1 overexpression was 92-fold in Meta and 11-fold in HCC compared to the surrounding liver. The SUV correlated with GLUT1 and PCNA expression in HCC, but not Meta patients. Of note, four cases of poorly differentiated (P/D) HCC compared to moderately differentiated (M/D) HCC produced completely different results for FDG uptake (SUV, 14.4 vs. 4.0) and mRNA expression (G6Pase expression, 0.007 vs. 1.5). Variations in the expression of glucose metabolism-related enzymes between HCC and Meta patients are attributed to origin or degree of differentiation. Low FDG uptake in M/D HCC reflected low GLUT1 and high G6Pase expression, while high FDG accumulation in P/D HCC could reflect increased GLUT1 and decreased G6Pase expression. These results may explain why M/D HCC is not detected as sensitively by FDG-PET.
Assuntos
Carcinoma Hepatocelular/diagnóstico por imagem , Fluordesoxiglucose F18 , Glucose/metabolismo , Neoplasias Hepáticas/diagnóstico por imagem , Tomografia por Emissão de Pósitrons/métodos , Idoso , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/cirurgia , Feminino , Fluordesoxiglucose F18/química , Transportador de Glucose Tipo 1/análise , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 2/análise , Glucose-6-Fosfatase/genética , Glucose-6-Fosfatase/metabolismo , Glucose-6-Fosfato/análogos & derivados , Glucose-6-Fosfato/química , Hepatectomia , Hexoquinase/análise , Hexoquinase/genética , Humanos , Fígado/diagnóstico por imagem , Fígado/metabolismo , Fígado/patologia , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/cirurgia , Masculino , Antígeno Nuclear de Célula em Proliferação/genética , RNA Mensageiro/biossíntese , Compostos RadiofarmacêuticosRESUMO
Islet equivalent (IE), the standard estimate of isolated islet volume, is an essential measure to determine the amount of transplanted islet tissue in the clinic and is used in research laboratories to normalize results, yet it is based on the false assumption that all islets are spherical. Here, we developed and tested a new easy-to-use method to quantify islet volume with greater accuracy. Isolated rat islets were dissociated into single cells, and the total cell number per islet was determined by using computer-assisted cytometry. Based on the cell number per islet, we created a regression model to convert islet diameter to cell number with a high R2 value (0.8) and good validity and reliability with the same model applicable to young and old rats and males or females. Conventional IE measurements overestimated the tissue volume of islets. To compare results obtained using IE or our new method, we compared Glut2 protein levels determined by Western Blot and proinsulin content via ELISA between small (diameter≤100 µm) and large (diameter≥200 µm) islets. When normalized by IE, large islets showed significantly lower Glut2 level and proinsulin content. However, when normalized by cell number, large and small islets had no difference in Glut2 levels, but large islets contained more proinsulin. In conclusion, normalizing islet volume by IE overestimated the tissue volume, which may lead to erroneous results. Normalizing by cell number is a more accurate method to quantify tissue amounts used in islet transplantation and research.
Assuntos
Contagem de Células , Tamanho Celular , Transplante das Ilhotas Pancreáticas , Ilhotas Pancreáticas/citologia , Animais , DNA/análise , Feminino , Transportador de Glucose Tipo 2/análise , Humanos , Ilhotas Pancreáticas/química , Masculino , Proinsulina/análise , Ratos , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
The present study aims to investigate the combined and individual treatment of aliskiren and olmesartan for 8 weeks in streptozotocin induced diabetic rats provide an effective blockade of RAAS by improving glucose homeostasis, glomerular filtration rate along with renal variables thereby delaying the progression of the disease. Streptozotocin induced diabetic rats were administered with aliskiren (10mg/kg/day), olmesartan (6mg/kg/day) alone and in combination. To identify the glucose homeostasis, translocation of glucose transporter proteins in liver and muscle was observed by their expression after treatment. Glomerular filtration rate is estimated using serum creatinine, cystatin C and beta 2 microglobulin. This study also examined the effects of combination and monotherapy on various renal variables viz. albumin, total proteins, TGF-ß, TNF-α, VEGF, nitric oxide, adiponectin and erythropoeitin. In addition, histopathological and anti-apoptotic profile of kidney was also investigated. The present study indicates that dual blockade of RAAS improved glucose homeostasis and confirms the nephroprotective effects of the combined treatment of aliskiren and olmesartan independent of their antihypertensive property in the STZ induced diabetes. In addition, its antifibrotic, antiproteinuric effects indicate that combination treatment might be potential as an important therapeutic option for chronic fibrotic diseases in renal complications.
Assuntos
Amidas/administração & dosagem , Bloqueadores do Receptor Tipo 1 de Angiotensina II/administração & dosagem , Glicemia/metabolismo , Diabetes Mellitus Experimental/tratamento farmacológico , Fumaratos/administração & dosagem , Taxa de Filtração Glomerular/efeitos dos fármacos , Imidazóis/administração & dosagem , Sistema Renina-Angiotensina/efeitos dos fármacos , Tetrazóis/administração & dosagem , Amidas/uso terapêutico , Bloqueadores do Receptor Tipo 1 de Angiotensina II/uso terapêutico , Animais , Anti-Hipertensivos/administração & dosagem , Diabetes Mellitus Experimental/fisiopatologia , Quimioterapia Combinada , Fumaratos/uso terapêutico , Transportador de Glucose Tipo 2/análise , Transportador de Glucose Tipo 4/análise , Humanos , Imidazóis/uso terapêutico , Insulina/sangue , Rim/efeitos dos fármacos , Rim/fisiopatologia , Fígado/química , Músculo Esquelético/química , Ratos , Ratos Wistar , Albumina Sérica/análise , Tetrazóis/uso terapêutico , Fatores de TempoRESUMO
In this study, we investigated the significance of ß(2)-adrenergic receptor (ß(2)AR) in age-related impaired insulin secretion and glucose homeostasis. We characterized the metabolic phenotype of ß(2)AR-null C57Bl/6N mice (ß(2)AR(-/-)) by performing in vivo and ex vivo experiments. In vitro assays in cultured INS-1E ß-cells were carried out in order to clarify the mechanism by which ß(2)AR deficiency affects glucose metabolism. Adult ß(2)AR(-/-) mice featured glucose intolerance, and pancreatic islets isolated from these animals displayed impaired glucose-induced insulin release, accompanied by reduced expression of peroxisome proliferator-activated receptor (PPAR)γ, pancreatic duodenal homeobox-1 (PDX-1), and GLUT2. Adenovirus-mediated gene transfer of human ß(2)AR rescued these defects. Consistent effects were evoked in vitro both upon ß(2)AR knockdown and pharmacologic treatment. Interestingly, with aging, wild-type (ß(2)AR(+/+)) littermates developed impaired insulin secretion and glucose tolerance. Moreover, islets from 20-month-old ß(2)AR(+/+) mice exhibited reduced density of ß(2)AR compared with those from younger animals, paralleled by decreased levels of PPARγ, PDX-1, and GLUT2. Overexpression of ß(2)AR in aged mice rescued glucose intolerance and insulin release both in vivo and ex vivo, restoring PPARγ/PDX-1/GLUT2 levels. Our data indicate that reduced ß(2)AR expression contributes to the age-related decline of glucose tolerance in mice.
Assuntos
Insulina/metabolismo , Receptores Adrenérgicos beta 2/fisiologia , Fatores Etários , Animais , Teste de Tolerância a Glucose , Transportador de Glucose Tipo 2/análise , Transportador de Glucose Tipo 2/fisiologia , Proteínas de Homeodomínio/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , PPAR gama/fisiologia , Fenótipo , Transativadores/fisiologiaRESUMO
Diets containing corn starch may improve glucose supply by providing significant amounts of intestinal starch and increasing intestinal glucose absorption in dairy cows. Glucose absorption in the small intestine requires specific glucose transporters; that is, sodium-dependent glucose co-transporter-1 (SGLT1) and facilitated glucose transporter (GLUT2), which are usually downregulated in the small intestine of functional ruminants but are upregulated when luminal glucose is available. We tested the hypothesis that mRNA and protein expression of intestinal glucose transporters and mRNA expression of enzymes related to gluconeogenesis are affected by variable starch supply. Dairy cows (n=9/group) were fed for 4 wk total mixed rations (TMR) containing either high (HS) or low (LS) starch levels in the diet. Feed intake and milk yield were measured daily. After slaughter, tissue samples of the small intestinal mucosa (mid-duodenum and mid-jejunum) were taken for determination of mRNA concentrations of SGLT1 and GLUT2 as well as pyruvate carboxylase, cytosolic phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase by real-time reverse transcription PCR relative to a housekeeping gene. Protein expression of GLUT2 in crude mucosal membranes and of SGLT1 and GLUT2 in brush-border membrane vesicles was quantified by sodium dodecyl sulfate-PAGE and immunoblot. A mixed model was used to examine feeding and time-related changes on feed intake and milk yield and to test feeding and gut site effects on gene or protein expression of glucose transporters and enzymes in the intestinal mucosa. Dry matter intake, but not energy intake, was higher in cows fed HS compared with LS. Abundance of SGLT1 mRNA tended to be higher in duodenal than in jejunal mucosa, and mRNA abundances of pyruvate carboxylase tended to be higher in jejunal than in duodenal mucosa. In brush-border membrane vesicles, SGLT1 and GLUT2 protein expression could be demonstrated. No diet-dependent differences were found concerning mRNA and protein contents of glucose transporter or mRNA level of gluconeogenic enzymes. In conclusion, our investigations on glucose transporters and gluconeogenic enzymes in the small intestinal mucosa of dairy cows did not show significant diet regulation when TMR with different amounts of intestinal starch were fed. Therefore, predicted intestinal glucose absorption after enhanced starch feeding is probably not supported by changes of intestinal glucose transporters in dairy cows.
Assuntos
Dieta/veterinária , Glucose/biossíntese , Mucosa Intestinal/enzimologia , Lactação/metabolismo , Proteínas de Transporte de Sódio-Glucose/análise , Amido/farmacologia , Animais , Bovinos , Relação Dose-Resposta a Droga , Duodeno/química , Duodeno/efeitos dos fármacos , Duodeno/enzimologia , Duodeno/metabolismo , Feminino , Transportador de Glucose Tipo 2/análise , Glucose-6-Fosfatase/análise , Mucosa Intestinal/química , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Jejuno/química , Jejuno/efeitos dos fármacos , Jejuno/enzimologia , Jejuno/metabolismo , Lactação/efeitos dos fármacos , Piruvato Carboxilase/análise , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Transportador 1 de Glucose-Sódio/análise , Amido/administração & dosagemRESUMO
Dioscorea is a traditional medicinal food in Asia. This study investigated the anti-insulin resistance of dioscorea polysaccharide (DPS) in inflammatory factor (tumor necrosis factor-α; TNF-α) induced mouse normal liver FL83B cells. Insulin resistance was induced by treating cells with TNF-α (20 ng/mL) for 5 h; subsequently, the medium was replaced with insulin and DPS for 60 min of incubation (model 1; alleviating group). In addition, cells were cotreated with TNF-α and DPS for 5 h in model 2 (preventing group). DPS effectively increased glucose uptake and glucose transporter 2 (GLUT2) expression of insulin-resistant cells. Furthermore, DPS stimulated insulin receptor substrate (IRS) tyrosyl phosphorylation and increased p-Akt level to alleviate insulin resistance in models 1 and 2. Finally, the possible mechanism of DPS promoting insulin sensitivity in TNF-α-induced FL83B cells was investigated in this study. DPS may attenuate c-Jun N-terminal kinases (JNK) and insulin resistance caused by TNF-α induction; therefore, DPS also elevated the levels of p-IRS(Tyr) and p-Akt(Ser) to improve insulin sensitivity in the TNF-α-induced FL83B cells.
Assuntos
Dioscorea/química , Resistência à Insulina , Polissacarídeos/farmacologia , Fator de Necrose Tumoral alfa/farmacologia , 4-Cloro-7-nitrobenzofurazano/análogos & derivados , 4-Cloro-7-nitrobenzofurazano/metabolismo , Animais , Linhagem Celular , Desoxiglucose/análogos & derivados , Desoxiglucose/metabolismo , Transportador de Glucose Tipo 2/análise , Fígado , CamundongosRESUMO
BACKGROUND: Pancreatic cancer is among the most dismal of human malignancies. The 5-year survival rate is lower than 5%. The identification of precursor lesions would be the main step to improve this fatal outcome. One precursor lesions are called pancreatic intraepithelial neoplasia (PanIN) and are graduated in grade 1 to 3, whereas grade 3 is classified as carcinoma in situ. Currently, no reliable, noninvasive imaging technique (e.g., ultrasound, computed tomography, magnet resonance imaging) exists to verify PanINs. METHODS: Recently, a transgenic mouse model of pancreatic cancer was established in which the tumor progression of human pancreatic carcinoma is reproduced. These so-called Pdx-1-Cre; LSL-KrasG12D/+; LSL-Trp53R172H/+mice develop PanINs, which transform to invasive growing pancreatic carcinoma. The pancreata of mice of different ages were immunohistochemically stained using α-GLUT-2 antibodies. Furthermore, mice underwent positron emission tomography (PET)-computed tomography (CT) with (18)F-fluorodeoxyglucose (FDG) to evaluate early detection of PanIN lesions. RESULTS: An expression of GLUT-2 in murine PanINs was found in PanINs of grade 1B and higher. This finding is associated with an elevated glucose metabolism, leading to the detection of precursor lesions of pancreatic cancer in the FDG PET-CT scan. In addition, immunohistochemical staining of GLUT-2 was detectable in 45 (75%) of 60 human PanINs, whereas PanINs of grade 1B and higher showed a very extensive expression. CONCLUSIONS: In conclusion, we demonstrate for the first time that an elevated glucose metabolism occurs already in precursor lesions of murine and human pancreatic carcinoma. These findings are the basis for the detection of precursor lesions by PET-CT, thereby helping improving the prognosis of this devastating disease.
Assuntos
Adenocarcinoma/diagnóstico por imagem , Carcinoma in Situ/diagnóstico por imagem , Glucose/metabolismo , Neoplasias Pancreáticas/diagnóstico por imagem , Tomografia por Emissão de Pósitrons/métodos , Lesões Pré-Cancerosas/diagnóstico por imagem , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Carcinoma in Situ/metabolismo , Carcinoma in Situ/patologia , Modelos Animais de Doenças , Progressão da Doença , Fluordesoxiglucose F18/análise , Fluordesoxiglucose F18/metabolismo , Engenharia Genética , Transportador de Glucose Tipo 2/análise , Humanos , Camundongos , Camundongos Transgênicos , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Lesões Pré-Cancerosas/metabolismo , Lesões Pré-Cancerosas/patologiaAssuntos
Proteínas Facilitadoras de Transporte de Glucose/análise , Odontogênese/fisiologia , Dente/embriologia , Ameloblastos/citologia , Animais , Órgão do Esmalte/embriologia , Células Epiteliais/citologia , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas Facilitadoras de Transporte de Glucose/fisiologia , Transportador de Glucose Tipo 1/análise , Transportador de Glucose Tipo 2/análise , Camundongos , Camundongos Endogâmicos ICR , Odontoblastos/citologia , Técnicas de Cultura de Órgãos , Dente/crescimento & desenvolvimentoRESUMO
Two F(2) Charolais x German Holstein families comprising full and half sibs share identical but reciprocal paternal and maternal Charolais grandfathers differ in milk production. We hypothesized that differences in milk production were related to differences in nutritional partitioning revealed by glucose metabolism and carcass composition. In 18F(2) cows originating from mating Charolais bulls to German Holstein cows and a following intercross of the F(1) individuals (n=9 each for family Ab and Ba; capital letters indicate the paternal and lowercase letter the maternal grandsire), glucose tolerance tests were performed at 10 d before calving and 30 and 93 d in milk (DIM) during second lactation. Glucose half-time as well as areas under the concentration curve for plasma glucose and insulin were calculated. At 94 DIM cows were infused intravenously with 18.3 micromol of d-[U-(13)C(6)]glucose/kg(0.75) of BW, and blood samples were taken to measure rate of glucose appearance and glucose oxidation as well as plasma concentrations of metabolites and hormones. Cows were slaughtered at 100 DIM and carcass size and composition was evaluated. Liver samples were taken to measure glycogen and fat content, gene expression levels, and enzyme activities of pyruvate carboxylase, phosphoenolpyruvate carboxykinase, and glucose 6-phosphatase as well as gene expression of glucose transporter 2. Milk yield was higher and milk protein content at 30 DIM was lower in Ba than in Ab cows. Glucose half-life was higher but insulin secretion after glucose challenge was lower in Ba than in Ab cows. Cows of Ab showed higher glucose oxidation, and plasma concentrations at 94 DIM were lower for glucose and insulin, whereas beta-hydroxybutyrate was higher in Ba cows. Hepatic gene expression of pyruvate carboxylase, glucose 6-phosphatase, and glucose transporter 2 were higher whereas phosphoenolpyruvate carboxykinase activities were lower in Ba than in Ab cows. Carcass weight as well as fat content of the carcass were higher in Ab than in Ba cows, whereas mammary gland mass was lower in Ab than in Ba cows. Fat classification indicated leaner carcass composition in Ba than in Ab cows. In conclusion, the 2 families showed remarkable differences in milk production that were accompanied by changes in glucose metabolism and body composition, indicating capacity for milk production as main metabolic driving force. Sex chromosomal effects provide an important regulatory mechanism for milk performance and nutrient partitioning that requires further investigation.
Assuntos
Composição Corporal/genética , Cruzamento , Bovinos/genética , Bovinos/metabolismo , Glucose/metabolismo , Lactação/genética , Leite/metabolismo , Tecido Adiposo , Animais , Glicemia/análise , Peso Corporal/genética , Feminino , Transportador de Glucose Tipo 2/análise , Glicogênio/análise , Insulina/sangue , Fígado/química , Fígado/enzimologia , Fígado/metabolismo , Masculino , Leite/químicaRESUMO
Islet transplantation is a potential cure for type 1 diabetes, but clinical results have been disappointing. Currently, islet isolation is by enzymatic digestion of the pancreas which has significant pitfalls: warm ischemia exposure, collagenase-induced damage to the islet mass and viability, poor reproducibility, high cost, a relatively low number of islets obtained per whole pancreas, and selection of islets for collagenase resistance rather than for glucose responsiveness. In the present study we performed a series of experiments in a porcine model to demonstrate the feasibility of a new isolation method based on selective osmotic shock (SOS) using very high glucose solutions, doubling or tripling physiological osmotic strength. The SOS method can be carried out at room temperature or in the cold eliminating warm ischemia time which damages the islets. The SOS method does not depend on the texture of the pancreas so all pancreases can be processed identically and the process can be fully automated. The SOS method isolates all the islets of the pancreas regardless of size and shape allowing a greater number of islets to be harvested. The SOS method avoids exposure to toxins in collagenase solutions, is inexpensive and selects for islets with high concentrations of Glut 2 transporters, representing the best glucose responding islets. The SOS method showed a comparable recovery of islets from young pig pancreas and the islets showed improved viability. We conclude that the selective osmotic shock (SOS) method of separating islets from the pancreatic tissue is superior to the collagenase method.
Assuntos
Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/fisiologia , Animais , Morte Celular , Separação Celular/métodos , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/cirurgia , Glucose/farmacologia , Transportador de Glucose Tipo 2/análise , Humanos , Hipoglicemiantes/uso terapêutico , Insulina/metabolismo , Insulina/uso terapêutico , Secreção de Insulina , Células Secretoras de Insulina/imunologia , Células Secretoras de Insulina/patologia , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Transplante das Ilhotas Pancreáticas , Pressão Osmótica , SuínosRESUMO
Diabetes ultimately results from an inadequate number of functional beta cells in the islets of Langerhans. Enhancing proliferation of functional endogenous beta cells to treat diabetes remains underexplored. Here, we report that excision of the Men1 gene, whose loss-of-function mutation leads to inherited multiple endocrine neoplasia type 1 (MEN1), rendered resistant to streptozotocin-induced hyperglycemia in a tamoxifen-inducible and temporally controlled Men1 excision mouse model as well as in a tissue-specific Men1 excision mouse model. Men1 excision prevented mice from streptozotocin-induced hyperglycemia mainly through increasing the number of functional beta cells. BrdU incorporation by beta cells, islet size, and circulating insulin levels were significantly increased in Men1-excised mice. Membrane localization of glucose transporter 2 was largely preserved in Men1-excised beta cells, but not in Men1-expressing beta cells. Our findings suggest that repression of menin, a protein encoded by the Men1 gene, might be a valuable means to maintain or increase the number of functional endogenous beta cells to prevent or ameliorate diabetes.
Assuntos
Diabetes Mellitus Experimental/prevenção & controle , Hiperglicemia/prevenção & controle , Proteínas Proto-Oncogênicas/fisiologia , Animais , Proliferação de Células , Transportador de Glucose Tipo 2/análise , Células Secretoras de Insulina/fisiologia , Masculino , Camundongos , Proteínas Proto-Oncogênicas/genética , EstreptozocinaRESUMO
PURPOSE: Hypoxia is detected in most solid tumors and is associated with malignant progression and adverse treatment outcomes. However, the oxygenation status of malignant salivary gland tumors has not been previously studied. The aim of this study was to investigate the potential clinical relevance of hypoxia in this tumor type. METHODS AND MATERIALS: Twelve patients scheduled for surgical resection of a salivary gland tumor were preoperatively injected with the hypoxia marker pimonidazole and the proliferation marker iododeoxyuridine. Tissue samples of the dissected tumor were immunohistochemically stained for blood vessels, pimonidazole, carbonic anhydrase-IX, glucose transporters-1 and -3 (Glut-1, Glut-3), hypoxia-inducible factor-1alpha, iododeoxyuridine, and epidermal growth factor receptor. The tissue sections were quantitatively assessed by computerized image analysis. RESULTS: The tissue material from 8 patients was of sufficient quality for quantitative analysis. All tumors were negative for pimonidazole binding, as well as for carbonic anhydrase-IX, Glut-1, Glut-3, and hypoxia-inducible factor-1alpha. The vascular density was high, with a median value of 285 mm(-2) (range, 209-546). The iododeoxyuridine-labeling index varied from <0.1% to 12.2% (median, 2.2%). Epidermal growth factor receptor expression levels were mostly moderate to high. In one-half of the cases, nuclear expression of epidermal growth factor receptor was observed. CONCLUSION: The absence of detectable pimonidazole binding, as well as the lack of expression of hypoxia-associated proteins in all tumors, indicates that malignant salivary gland tumors are generally well oxygenated. It is unlikely that hypoxia is a relevant factor for their clinical behavior and treatment responsiveness.
Assuntos
Hipóxia Celular , Neoplasias das Glândulas Salivares/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Neoplasias/análise , Anidrase Carbônica IX , Anidrases Carbônicas/análise , Proliferação de Células , Desoxiuridina/análogos & derivados , Transportador de Glucose Tipo 1/análise , Transportador de Glucose Tipo 2/análise , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/análise , Pessoa de Meia-Idade , Nitroimidazóis/metabolismo , Neoplasias das Glândulas Salivares/irrigação sanguínea , Neoplasias das Glândulas Salivares/química , Neoplasias das Glândulas Salivares/patologiaRESUMO
Intestinal function in young animals is influenced by maternal factors, such as alterations in the maternal diet. Glucagon-like peptide 2 (GLP-2) enhances intestinal growth and absorption in mature animals. Glucocorticosteroids induce intestinal maturation in neonates and increase sugar uptake in adult animals. It is not known if maternally administered GLP-2 or glucocorticosteroids have persistent effects on intestinal transport in the offspring. This study was undertaken to determine (1) the influence of maternal GLP-2, dexamethasone (DEX) and GLP-2+DEX on intestinal sugar uptake in postweaning offspring and (2) if alterations in uptake are due to variations in intestinal morphology, sugar transporter abundance or the abundance of selected signals. Nursing rat dams were treated during pregnancy and lactation with GLP-2 (0.1 mug/g per day sc), DEX (0.128 microg/g per day sc), GLP-2+DEX or placebo. The offspring were sacrificed 4 weeks after weaning, and glucose and fructose uptake was determined using an in vitro intestinal ring uptake technique. sodium-dependent glucose transporter, glucose transporter (GLUT) 5, GLUT2, sodium potassium adenosine triphosphatase and selected signals were assessed by immunohistochemistry. The treatments did not affect body weights or intestinal morphology. GLP-2 and GLP-2+DEX increased jejunal fructose uptake, and GLP-2+DEX increased the jejunal and ileal maximal transport rate for glucose uptake. Protein kinase B and mammalian target of rapamycin abundance were also increased, while transporter abundance was unchanged. We speculate that these alterations in sugar uptake may be due to changes in the intrinsic activity of the transporters mediated by the phosphatidylinositol-3-kinase pathway. These alterations in uptake may have nutritional implications for the offspring of mothers who may be treated with GLP-2 or glucocorticosteroids.