RESUMO
Advanced glycation end products (AGEs) prime macrophages for lipopolysaccharide (LPS)-induced inflammation. We investigated the persistence of cellular AGE-sensitization to LPS, considering the nuclear content of p50 and p65 nuclear factor kappa B (NFKB) subunits and the expression of inflammatory genes. Macrophages treated with control (C) or AGE-albumin were rested for varying intervals in medium alone before being incubated with LPS. Comparisons were made using one-way ANOVA or Student t-test (n = 6). AGE-albumin primed macrophages for increased responsiveness to LPS, resulting in elevated levels of TNF, IL-6, and IL-1beta (1.5%, 9.4%, and 5.6%, respectively), compared to C-albumin. TNF, IL-6, and IL-1 beta secretion persisted for up to 24 h even after the removal of AGE-albumin (area under the curve greater by 1.6, 16, and 5.2 times, respectively). The expressions of Il6 and RelA were higher 8 h after albumin removal, and Il6 and Abca1 were higher 24 h after albumin removal. The nuclear content of p50 remained similar, but p65 showed a sustained increase (2.9 times) for up to 24 h in AGE-albumin-treated cells. The prolonged activation of the p65 subunit of NFKB contributes to the persistent effect of AGEs on macrophage inflammatory priming, which could be targeted for therapies to prevent complications based on the AGE-RAGE-NFKB axis.
Assuntos
Interleucina-6 , NF-kappa B , NF-kappa B/metabolismo , Interleucina-6/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/metabolismo , Produtos Finais de Glicação Avançada/metabolismo , Albuminas/metabolismoRESUMO
A low-sodium (LS) diet has been shown to reduce blood pressure (BP) and the incidence of cardiovascular diseases. However, severe dietary sodium restriction promotes insulin resistance (IR) and dyslipidemia in animal models and humans. Thus, further clarification of the long-term consequences of LS is needed. Here, we investigated the effects of chronic LS on gastrocnemius gene and protein expression and lipidomics and its association with IR and plasma lipids in LDL receptor knockout mice. Three-month-old male mice were fed a normal sodium diet (NS; 0.5% Na; n = 12-19) or LS (0.06% Na; n = 14-20) over 90 days. Body mass (BM), BP, plasma total cholesterol, triacylglycerol (TG), glucose, hematocrit, and IR were evaluated. LS increased BM (9%), plasma TG (51%), blood glucose (19%), and IR (46%) when compared with the NS. RT-qPCR analysis revealed that genes involved in lipid uptake and oxidation were increased by the LS: Fabp3 (106%), Prkaa1 (46%), and Cpt1 (74%). Genes and proteins (assessed by Western blotting) involved in insulin signaling were not changed by the LS. Similarly, lipid species classically involved in muscle IR, such as diacylglycerols and ceramides detected by ultra-high-performance liquid chromatography coupled to mass spectrometry, were also unchanged by LS. Species of phosphatidylcholines (68%), phosphatidylinositol (90%), and free fatty acids (59%) increased while cardiolipins (41%) and acylcarnitines (9%) decreased in gastrocnemius in response to LS and were associated with glucose disposal rate. Together these results suggest that chronic LS alters glycerophospholipid and fatty acids species in gastrocnemius that may contribute to glucose and lipid homeostasis derangements in mice.
Assuntos
Dieta Hipossódica , Resistência à Insulina , Metabolismo dos Lipídeos , Músculo Esquelético/metabolismo , Animais , Lipidômica , Masculino , Camundongos , Sódio na Dieta/metabolismoRESUMO
Little is known about advanced glycation end products (AGEs) participation in glucose homeostasis, a process in which skeletal muscle glucose transporter GLUT4 (Scl2a4 gene) plays a key role. This study investigated (1) the in vivo and in vitro effects of AGEs on Slc2a4/GLUT4 expression in skeletal muscle of healthy rats, and (2) the potential involvement of endoplasmic reticulum and inflammatory stress in the observed regulations. For in vivo analysis, rats were treated with advanced glycated rat albumin (AGE-albumin) for 12 weeks; for in vitro analysis, soleus muscles from normal rats were incubated with bovine AGE-albumin for 2.5 to 7.5 hours. In vivo, AGE-albumin induced whole-body insulin resistance; decreased (~30%) Slc2a4 mRNA and GLUT4 protein content; and increased (~30%) the nuclear content of nuclear factor NF-kappa-B p50 subunit (NFKB1), and cellular content of 78 kDa glucose-regulated protein (GRP78). In vitro, incubation with AGE-albumin decreased (~50%) the Slc2a4/GLUT4 content; and increased cellular content of GRP78/94, phosphorylated-IKK-alpha/beta, nuclear content of NFKB1 and RELA, and the nuclear protein binding into Slc2a4 promoter NFKB-binding site. The data reveal that AGEs impair glucose homeostasis in non-diabetic states of increased AGEs concentration; an effect that involves activation of endoplasmic reticulum- and inflammatory-stress and repression of Slc2a4/GLUT4 expression.
Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Transportador de Glucose Tipo 4/genética , Produtos Finais de Glicação Avançada/farmacologia , Resistência à Insulina , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Animais , Biomarcadores/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Glucose/metabolismo , Homeostase/efeitos dos fármacos , Masculino , NF-kappa B/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos WistarRESUMO
Background: Advanced glycation endproducts elicit inflammation. However, their role in adipocyte macrophage infiltration and in the development of insulin resistance, especially in the absence of the deleterious biochemical pathways that coexist in diabetes mellitus, remains unknown. We investigated the effect of chronic administration of advanced glycated albumin (AGE-albumin) in healthy rats, associated or not with N-acetylcysteine (NAC) treatment, on insulin sensitivity, adipose tissue transcriptome and macrophage infiltration and polarization. Methods: Male Wistar rats were intraperitoneally injected with control (C) or AGE-albumin alone, or, together with NAC in the drinking water. Biochemical parameters, lipid peroxidation, gene expression and protein contents were, respectively, determined by enzymatic techniques, reactive thiobarbituric acid substances, RT-qPCR and immunohistochemistry or immunoblot. Carboxymethyllysine (CML) and pyrraline (PYR) were determined by LC/mass spectrometry (LC-MS/MS) and ELISA. Results: CML and PYR were higher in AGE-albumin as compared to C. Food consumption, body weight, systolic blood pressure, plasma lipids, glucose, hepatic and renal function, adipose tissue relative weight and adipocyte number were similar among groups. In AGE-treated animals, insulin resistance, adipose macrophage infiltration and Col12a1 mRNA were increased with no changes in M1 and M2 phenotypes as compared to C-albumin-treated rats. Total GLUT4 content was reduced by AGE-albumin as compared to C-albumin. NAC improved insulin sensitivity, reduced urine TBARS, adipose macrophage number and Itgam and Mrc mRNA and increased Slc2a4 and Ppara. CD11b, CD206, Ager, Ddost, Cd36, Nfkb1, Il6, Tnf, Adipoq, Retn, Arg, and Il12 expressions were similar among groups. Conclusions: AGE-albumin sensitizes adipose tissue to inflammation due to macrophage infiltration and reduces GLUT4, contributing to insulin resistance in healthy rats. NAC antagonizes AGE-albumin and prevents insulin resistance. Therefore, it may be a useful tool in the prevention of AGE action on insulin resistance and long-term complications of DM.
RESUMO
Because of the paucity of information regarding metabolic effects of advanced glycation end products (AGEs) on liver, we evaluated effects of AGEs chronic administration in (1) insulin sensitivity; (2) hepatic expression of genes involved in AGEs, glucose and fat metabolism, oxidative stress and inflammation and; (3) hepatic morphology and glycogen content. Rats received intraperitoneally albumin modified (AlbAGE) or not by advanced glycation for 12 weeks. AlbAGE induced whole-body insulin resistance concomitantly with increased hepatic insulin sensitivity, evidenced by activation of AKT, inactivation of GSK3, increased hepatic glycogen content, and decreased expression of gluconeogenesis genes. Additionally there was reduction in hepatic fat content, in expression of lipogenic, pro-inflamatory and pro-oxidative genes and increase in reactive oxygen species and in nuclear expression of NRF2, a transcription factor essential to cytoprotective response. Although considered toxic, AGEs become protective when administered chronically, stimulating AKT signaling, which is involved in cellular defense and insulin sensitivity.
Assuntos
Produtos Finais de Glicação Avançada/farmacologia , Hormese/efeitos dos fármacos , Resistência à Insulina , Fígado/metabolismo , Albuminas/farmacologia , Animais , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Gluconeogênese/efeitos dos fármacos , Gluconeogênese/genética , Produtos Finais de Glicação Avançada/administração & dosagem , Glicogênio/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Proteína HMGB1/metabolismo , Mediadores da Inflamação/metabolismo , Injeções Intraperitoneais , Lipogênese/efeitos dos fármacos , Lipogênese/genética , Fígado/efeitos dos fármacos , Masculino , Modelos Biológicos , Fator 2 Relacionado a NF-E2/metabolismo , Oxirredução , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismoRESUMO
AIMS: The fetal programming hypothesis suggests that intrauterine stimuli can induce metabolic changes in offspring, increasing the disease risk in adulthood. Periodontal disease may enhance serum cytokine levels. Cytokines such as tumor necrosis factor-alpha (TNF-α) have been associated with reduced glucose transporter type 4 (GLUT4) expression, decreased protein kinase B (Akt) phosphorylation, and insulin resistance. This study aimed to evaluate GLUT4 content, and Akt serine phosphorylation status in the gastrocnemius skeletal muscle (GSM), glycemia, insulinemia and change in body weight in offspring of rats with periodontal disease. MAIN METHODS: Female Wistar rats were distributed into a control group (CN) and an experimental periodontal disease group (PD), in which a ligature was placed around the mandibular first molars. Seven days after ligature placement, both groups were mated with normal male rats. The ligatures remained throughout pregnancy until weaning, after which the male offspring were distributed into groups: CN-o, control rat offspring; and PD-o, periodontal disease rat offspring. The body weight from 0 to 75days of age was measured. At 75days, the glycemia, insulinemia, TNF-α levels, Akt serine phosphorylation, and GLUT4 content in the GSM were measured in the offspring. KEY FINDINGS: The PD-o group showed a low birth weight (LBW), unchanged glycemia, increased insulinemia, insulin resistance, increased TNF-α levels, decreased Akt serine phosphorylation status, and reduced GLUT4 content in the plasma membrane and translocation index after insulin stimulation. SIGNIFICANCE: Maternal periodontal disease causes LBW, insulin resistance, and alterations in the final stage of insulin signaling in the GSM of adult offspring.
Assuntos
Membrana Celular/metabolismo , Transportador de Glucose Tipo 4/metabolismo , Músculo Esquelético/metabolismo , Complicações na Gravidez/metabolismo , Animais , Glicemia/metabolismo , Feminino , Resistência à Insulina/fisiologia , Masculino , Gravidez , Complicações na Gravidez/patologia , Ratos , Ratos WistarRESUMO
OBJECTIVES: Periapical lesion (PL) promotes insulin resistance; however, the mechanisms underlying this alteration are not fully understood. Therefore, in this study, we aimed to evaluate the Akt serine phosphorylation status and GLUT4 expression levels in the gastrocnemius muscle (GM) of rats with PL. MATERIALS AND METHODS: Male Wistar rats (n = 42) were distributed equally into control (CN) and PL groups. The pulpal tissue of the PL group rats was exposed to the oral environment for 30 days. Thereafter, glucose and insulin levels were assessed, followed by homeostasis model assessment of insulin resistance (HOMA-IR). The Akt serine phosphorylation and GLUT4 levels of microsomal (M) and plasma membrane (PM) fractions were evaluated by western blotting and analyzed statistically. RESULTS: Compared to CN group rats, PL group rats had lower insulin sensitivity (as observed by HOMA-IR), lower Akt serine phosphorylation status after insulin stimulus, and lower GLUT4 levels in the PM fraction. However, the M fraction in the PL group did not differ significantly from that of the CN group. CONCLUSIONS: PL decreases insulin sensitivity, Akt phosphorylation, and PM GLUT4 content. CLINICAL RELEVANCE: The present study indicates that preventing endodontic disease can thwart insulin resistance.
Assuntos
Polpa Dentária/lesões , Transportador de Glucose Tipo 4/metabolismo , Músculo Esquelético/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Western Blotting , Membrana Celular/metabolismo , Modelos Animais de Doenças , Resistência à Insulina , Masculino , Fosforilação , Ratos , Ratos WistarRESUMO
We characterized the metabolic profile of transgenic mice exhibiting enhanced muscle mass driven by increased mIGF-1 expression (MLC/mIGF-1). As expected, 6-month-old MLC/mIGF-1 mice were heavier than age-matched wild type (WT) mice (37.4 ± 0.3 versus 31.8 ± 0.6 g, resp.). MLC/mIGF-1 mice had higher respiratory quotient when compared to WT (0.9 ± 0.03 versus 0.74 ± 0.02, resp.) suggesting a preference for carbohydrate as the major fuel source. MLC/mIGF-1 mice had a higher rate of glucose disposal when compared to WT (3.25 ± 0.14 versus 2.39 ± 0.03%/min, resp.). The higher disposal rate correlated to â¼ 2-fold higher GLUT4 content in the extensor digitorum longus (EDL) muscle. Analysis of mRNA content for the glycolysis-related gene PFK-1 showed â¼ 3-fold upregulation in MLC/mIGF-1 animals. We also found a 50% downregulation of PGC1α mRNA levels in MLC/mIGF-1 mouse EDL muscle, suggesting less abundant mitochondria in this tissue. We found no difference in the expression of PPARα and PPARß/δ, suggesting no modulation of key elements in oxidative metabolism. These data together suggest a shift in metabolism towards higher carbohydrate utilization, and that could explain the increased insulin sensitivity of hypertrophied skeletal muscle in MLC/mIGF-1 mice.
Assuntos
Metabolismo dos Carboidratos/fisiologia , Hipertrofia/metabolismo , Resistência à Insulina/fisiologia , Fator de Crescimento Insulin-Like I/metabolismo , Insulina/metabolismo , Músculo Esquelético/metabolismo , Animais , Transportador de Glucose Tipo 4/metabolismo , Camundongos , Camundongos Transgênicos , Mitocôndrias/metabolismo , Proteínas Musculares/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , RNA Mensageiro/metabolismo , Fatores de Transcrição/metabolismoRESUMO
AIMS: Solute carrier 2a2 (Slc2a2) gene codifies the glucose transporter GLUT2, a key protein for glucose flux in hepatocytes and renal epithelial cells of proximal tubule. In diabetes mellitus, hepatic and tubular glucose output has been related to Slc2a2/GLUT2 overexpression; and controlling the expression of this gene may be an important adjuvant way to improve glycemic homeostasis. Thus, the present study investigated transcriptional mechanisms involved in the diabetes-induced overexpression of the Slc2a2 gene. MAIN METHODS: Hepatocyte nuclear factors 1α and 4α (HNF-1α and HNF-4α), forkhead box A2 (FOXA2), sterol regulatory element binding protein-1c (SREBP-1c) and the CCAAT-enhancer-binding protein (C/EBPß) mRNA expression (RT-PCR) and binding activity into the Slc2a2 promoter (electrophoretic mobility assay) were analyzed in the liver and kidney of diabetic and 6-day insulin-treated diabetic rats. KEY FINDINGS: Slc2a2/GLUT2 expression increased by more than 50% (P<0.001) in the liver and kidney of diabetic rats, and 6-day insulin treatment restores these values to those observed in non-diabetic animals. Similarly, the mRNA expression and the binding activity of HNF-1α, HNF-4α and FOXA2 increased by 50 to 100% (P<0.05 to P<0.001), also returning to values of non-diabetic rats after insulin treatment. Neither the Srebf1 and Cebpb mRNA expression, nor the SREBP-1c and C/EBP-ß binding activity was altered in diabetic rats. SIGNIFICANCE: HNF-1α, HNF-4α and FOXA2 transcriptional factors are involved in diabetes-induced overexpression of Slc2a2 gene in the liver and kidney. These data point out that these transcriptional factors are important targets to control GLUT2 expression in these tissues, which can contribute to glycemic homeostasis in diabetes.
Assuntos
Diabetes Mellitus Experimental/genética , Transportador de Glucose Tipo 2/genética , Fator 1-alfa Nuclear de Hepatócito/genética , Fator 3-beta Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/genética , Rim/metabolismo , Fígado/metabolismo , Animais , Sítios de Ligação , Proteína beta Intensificadora de Ligação a CCAAT/genética , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Transportador de Glucose Tipo 2/metabolismo , Fator 1-alfa Nuclear de Hepatócito/metabolismo , Fator 3-beta Nuclear de Hepatócito/metabolismo , Fator 4 Nuclear de Hepatócito/metabolismo , Insulina/farmacologia , Rim/efeitos dos fármacos , Fígado/efeitos dos fármacos , Masculino , Regiões Promotoras Genéticas , Ratos , Ratos Wistar , Proteína de Ligação a Elemento Regulador de Esterol 1/genéticaRESUMO
BACKGROUND: Oral health complications in diabetes and hypertension include decreased salivary secretion. The sodium-glucose cotransporter 1 (SGLT1) protein, which transports 1 glucose/2 Na+/264 H2O molecules, is described in salivary glands. We hypothesized that changes in SGLT1 expression in the luminal membrane of ductal cell may be related to an altered salivary flow. FINDINGS: By immunohistochemistry, we investigated SGLT1 expression in ductal cells of parotid and submandibular glands from Wistar Kyoto rats (WKY), diabetic WKY (WKY-D), spontaneously hypertensive rats (SHR) and diabetic SHR (SHR-D), as well as in parotid glands from WKY subjected to sympathetic stimulation, with or without previous propranolol blockade. Diabetes and hypertension decreased the salivary secretion and increased SGLT1 expression in the luminal membrane of ductal cells, and their association exacerbated the regulations observed. After 30 min of sympathetic stimulation, SGLT1 increased in the luminal membrane of ductal cells, and that was blocked by previous injection of propranolol. CONCLUSIONS: SGLT1 expression increases in the luminal membrane of salivary gland ductal cells and the salivary flow decreases in diabetic and hypertensive rats, which may be related to sympathetic activity. This study highlights the water transporter role of SGLT1 in salivary glands, which, by increasing ductal water reabsorption, may explain the hyposalivation of diabetic and hypertensive subjects.