Short-term low-protein diet during pregnancy alters islet area and protein content of phosphatidylinositol 3-kinase pathway in rats.

The phosphatidylinositol 3-kinase and mitogen-activated protein kinase pathways mediate ß cell growth, proliferation, survival and death. We investigated whether protein restriction during pregnancy alters islet morphometry or the expression and phosphorylation of several proteins involved in the phosphatidylinositol 3-kinase and mitogen-activated protein kinase pathways. As controls, adult pregnant and non-pregnant rats were fed a normal-protein diet (17%). Pregnant and non-pregnant rats in the experimental groups were fed a low-protein diet (6%) for 15 days. Low protein diet during pregnancy increased serum prolactin level, reduced serum corticosterone concentration and the expression of both protein kinase B/AKT1 (AKT1) and p70 ribosomal protein S6 kinase (p70S6K), as well as the islets area, but did not alter the insulin content of pancreatic islets. Pregnancy increased the expression of the Src homology/collagen (SHC) protein and the extracellular signal-regulated kinases 1/2 (ERK1/2) independent of diet. ERK1/2 phosphorylation (pERK1/2) was similar in islets from pregnant and non-pregnant rats fed a low-protein diet, and was higher in islets from pregnant rats than in islets from non-pregnant rats fed a normal-protein diet. Thus, a short-term, low-protein diet during pregnancy was sufficient to reduce the levels of proteins in the phosphatidylinositol 3-kinase pathway and affect islet morphometry.

Soybean diet alters the insulin-signaling pathway in the liver of rats recovering from early-life malnutrition.

OBJECTIVE: We investigated if alterations in the insulin-signaling pathway could contribute to reduced hepatic glycogen levels in adult rats subjected to a protein deficiency during intrauterine life and lactation and reared through to recovery on a soybean diet. METHODS: Rats from mothers fed with 17% or 6% protein (casein) during pregnancy and lactation were maintained with a 17% casein diet (offspring born to and suckled by mothers fed a control diet and subsequently fed the same diet after weaning [CC group] and offspring born to and suckled by mothers fed a control diet and subsequently fed a soybean flour diet with 17% protein after weaning [CS group]), a soybean diet (offspring of mothers fed a low-protein diet and a control diet after weaning [LC group] and offspring of mothers fed a low-protein diet and fed a soybean flour diet containing 17% protein after weaning [LS group]), or a 6% casein diet (offspring of mothers fed a low-protein diet and subsequently fed the same diet after weaning [LL group]) from weaning until 90 d of life. RESULTS: A soybean diet did not modify basal serum glucose and glucagon concentrations, but raised basal serum insulin and consequently increased the serum insulin/glucose ratio. Insulin receptor and insulin receptor substrate-1 levels were lower in rats fed a soybean diet compared with those maintained with a casein diet. In the LS group, the p85 levels were higher than in the LC group, whereas in CS rats its expression was lower than in CC rats. The expression of p110 was lower in the CS group compared with the CC group and similar in the LS and LC groups. Insulin receptor substrate-1 phosphorylation was similar in the LS, LC, and CS groups and lower compared with the CC group. The insulin receptor substrate-1-p85/phosphatidylinositol 3-kinase association was lower in LS than in LC rats and in CS than in CC rats. Akt phosphorylation was lower in the CS and LS groups than in the CC and LC groups. CONCLUSION: Adult rats maintained with a soybean diet exhibited insulin resistance due, at least in part, to alterations in the early steps of the insulin signal transduction pathway.

Soybean diet modulates acetyl-coenzyme A carboxylase expression in livers of rats recovering from early-life malnutrition.

OBJECTIVE: The present study evaluated the effect of nutritional recovery with a soybean diet on the gene and protein expressions and protein phosphorylation of several enzymes and transcription factors involved in hepatic lipid metabolism. METHODS: Rats from mothers fed with 17% or 6% protein (casein) during pregnancy and lactation were maintained with a 17% casein (CC and LC groups) or soybean (CS and LS groups) diet and with a 6% casein (LL group) diet until 90 d of life. RESULTS: The soybean diet enhanced serum insulin levels but decreased body and liver weights and hepatic lipid and glycogen concentrations. Liver peroxisome proliferator receptor-alpha mRNA abundance was higher in the LS and CS groups than in the LC and CC groups, but the protein content was similar in all groups. Hepatic acetyl-coenzyme A carboxylase (ACC)-alpha and ACCbeta mRNA expression was markedly lower in the LS and CS rats than in the LC and CC rats. ACC protein expression was lower in the CS group than in the CC, LC, and LS groups. Phospho-[Ser(79)]2-ACC content was similar in the CS, LC, and LS groups and lower than the CC group. In the CS rats this reduction paralleled the decrease in total ACC protein. Messenger RNA and protein expression of sterol regulatory element-binding protein 1c, adenosine monophosphate-activated protein kinase, and phospho-[Thr(172)]-adenosine monophosphate-activated protein kinase was not modified by the soybean diet. CONCLUSION: Thus, the soybean diet reduced the liver lipid concentration through downregulation of the ACC gene and protein expressions rather than by phosphorylation status, which possibly resulted in decreased lipogenesis and increased beta-oxidation.

Soybean diet improves insulin secretion through activation of cAMP/PKA pathway in rats.

Maternal malnutrition leads to permanent alterations in insulin secretion of offspring and the soybean diet contributes to improve insulin release. At least a soy component, genistein, seems to increase the insulin secretion by activating the cAMP/PKA and PLC/PKC pathways. Here, we investigated the effect of the soybean diet on the expression of PKAalpha and PKCalpha, and insulin secretion in response to glucose and activators of adenylate cyclase and PKC in adult pancreatic rat islets. Rats from mothers fed with 17% or 6% protein (casein) during pregnancy and lactation were maintained with 17% casein (CC and CR groups) or soybean (SC and SR groups) diet until 90 days of life. The soybean diet improved the insulin response to a physiological concentration of glucose in control islets, but only in the presence of supra-physiological concentrations of glucose in islets from CR and SR groups. PMA also improved the insulin response in islets of SC and SR groups. The expression of PKCalpha was similar in all groups. Forskolin increased the insulin secretion; however, the magnitude of the increment was lower in islets from CR and SR groups than in control animals and in those from rats maintained with soybean diet than in rats fed with casein diet. The PKAalpha expression was similar between SR and CR groups and lower in SC than in CC islets. Thus, soybean diet improved the secretory pattern of beta cells, at least in part, by activating the cAMP/PKA-signaling cascade.

Palmitic acid increase levels of pancreatic duodenal homeobox-1 and p38/stress-activated protein kinase in islets from rats maintained on a low protein diet.

A severe reduction in insulin release in response to glucose is consistently noticed in protein-deprived rats and is attributed partly to the chronic exposure to elevated levels of NEFA. Since the pancreatic and duodenal transcription factor homeobox 1 (PDX-1) is important for the maintenance of beta-cell physiology, and since PDX-1 expression is altered in the islets of rats fed a low protein (LP) diet and that rats show high NEFA levels, we assessed PDX-1 and insulin mRNA expression, as well as PDX-1 and p38/stress activated protein kinase 2 (SAPK2) protein expression, in islets from young rats fed low (6%) or normal (17%; control) protein diets and maintained for 48 h in culture medium containing 5.6 mmol/l glucose, with or without 0.6 mmol/l palmitic acid. We also measured glucose-induced insulin secretion and glucose metabolism. Insulin secretion by isolated islets in response to 16.7 mmol/l glucose was reduced in LP compared with control rats. In the presence of NEFA, there was an increase in insulin secretion in both groups. At 2.8 mmol/l glucose, the metabolism of this sugar was reduced in LP islets, regardless of the presence of this fatty acid. However, when challenged with 16.7 mmol/l glucose, LP and control islets showed a severe reduction in glucose oxidation in the presence of NEFA. The PDX-1 and insulin mRNA were significantly higher when NEFA was added to the culture medium in both groups of islets. The effect of palmitic acid on PDX-1 and p38/SAPK2 protein levels was similar in LP and control islets, but the increase was much more evident in LP islets. These results demonstrate the complex interrelationship between nutrients in the control of insulin release and support the view that fatty acids play an important role in glucose homeostasis by affecting molecular mechanisms and stimulus/secretion coupling pathways.

Expression of PDX-1 is reduced in pancreatic islets from pups of rat dams fed a low protein diet during gestation and lactation.

Intrauterine and early postnatal malnutrition has profound consequences on fetal and postnatal development in both humans and animals. In addition, low birth weight has been reported to be associated with impaired insulin secretion, insulin resistance and diminished area of pancreatic islets. Because the transcription factor pancreatic and duodenal homeobox 1 (PDX-1) is important for the maintenance of B-cell physiology, PDX-1 expression and islet area were assessed in neonatal rats of dams fed low (6%) or normal (17%) protein diets during pregnancy. PDX-1 protein and mRNA levels, as well as insulin secretion and islet area, were measured after 28 d of life in normal, low protein and recovered rats whose dams consumed a normal protein diet after delivery. Insulin secretion by isolated islets in response to 2.8 and 16.7 mmol glucose/L was reduced in 28-d-old low protein rats compared with the control (P < 0.05). At birth and after 28 d of life, the islet area and PDX-1 protein expression were also reduced (P < 0.05). In contrast, PDX-1 mRNA levels in islets from 28-d-old low protein rats were not different from control rats. PDX-1 protein expression in pancreatic islets, the area of islets and insulin secretion were restored in recovered rats, whereas PDX-1 mRNA levels were higher than in normal rats (P < 0.05). These results suggest a link among diminished PDX-1 protein expression, a reduction in islet area and impaired insulin secretion in low protein rats. The reintroduction of a normal diet early in life restored islet area and cell physiology.

Endurance training improves responsiveness to insulin and modulates insulin signal transduction through the phosphatidylinositol 3-kinase/Akt-1 pathway.

BACKGROUND: Endurance training increases insulin-stimulated muscle glucose transport and leads to improved metabolic control in diabetic patients. OBJECTIVE: To analyze the effects of endurance training on the early steps of insulin action in muscle of rats. DESIGN: Male rats submitted to daily swimming for 6 weeks were compared with sedentary controls. At the end of the training period, anesthetized animals received an intravenous (i.v.) injection of insulin and had a fragment of their gastrocnemius muscle excised for the experiments. METHODS: Associations between insulin receptor, insulin receptor substrates (IRS)-1 and -2 and phosphatidylinositol 3-kinase (PI3-kinase) were analyzed by immunoprecipitation and immunoblotting. Akt-1 serine phosphorylation and specific protein quantification were detected by immunoblotting of total extracts, and IRS-1/IRS-2-associated PI3-kinase activity were determined by thin-layer chromatography. RESULTS: Insulin-induced phosphorylation of IRS-1 and IRS-2 increased respectively by 1.8-fold (P<0.05) and 1.5-fold (P<0.05), whereas their association with PI3-kinase increased by 2.3-fold (P<0.05) and 1.9-fold (P<0.05) in trained rats as compared with sedentary controls, respectively. The activity of PI3-kinase associated with IRS-1 and IRS-2 increased by 1.8-fold (P<0.05) and 1.7-fold (P<0.05) respectively, in trained rats as compared with their untrained counterparts. Serine phosphorylation of Akt-1/PKB increased 1.7-fold (P<0.05) in trained rats in response to insulin. These findings were accompanied by increased responsiveness to insulin as demonstrated by a reduced area under the curve for insulin during an i.v. glucose tolerance test, by increased glucose disappearance rate during an insulin tolerance test, and by increased expression of glucose transporter-4. CONCLUSIONS: The increased responsiveness to insulin induced by chronic exercise in rat skeletal muscle may result, at least in part, from the modulation of the insulin signaling pathway at different molecular levels.

Deficiência nutricional e diabetes mellitus / Nutritional deficiency and diabetes mellitus

Distúrbios nutricionais estao relacionados ao desenvolvimento de diabetes (DM). Em países subdesenvolvidos milhares de indivíduos apresentam um tipo de DM relacionado à alimentaçao pobre em proteína e calorias. Existem indícios de que dietas hipoprotéicas e hipocalóicas induzam a uma perda funcional da célula beta pancreática e, conseqüentemente, levem a um desequilíbrio na homeostase da glicose. Outra possibilidade seria uma potencializaçao, pelo quadro de desnutriçao, de efeitos diabetogênicos ambientais, imunológicos ou genéticos preexistentes. Finalmente, a carência nutricional protéico-calórica poderia simplesmente modular o curso de um quadro preexistente de DM. Outro aspecto importante da influência do padrao nutricional sobre o controle dos níveis sangüíneos da glicose pode ser observado em indivíduos submetidos à carência nutricional durante as primeiras fases de vida, e que a seguir passam a se alimentar com dietas hipercalóricas. Tal mudança no padrao alimentar leva muitas vezes à obesidade e a uma maior incidência de diabetes. Em humanos e modelos animais de desnutriçao protéico-calórica observa-se uma reduzida secreçao de insulina em resposta a um estímulo de glicose. Aparentemente a carência de insulina é compensada, ao menos em parte, por uma maior sensibilidade periférica ao hormônio pancreático, a qual se deve a uma modulaçao nas etapas iniciais da sinalizaçao da insulina. Tal compensaçao garante a homeostase da glicose na maior parte dos indivíduos, entretanto diversos fatores podem romper este equilíbrio e desencadear o aparecimento de DM.