Sleep and probiotics in diabetes management.

In this manuscript, we call attention to the importance of investigating affordable strategies to improve the management of diabetes. Studies indicate that imbalances in the gut microbiota may lead to the development and progression of this disease. At the same time, poor sleep and sleep disorders, which are very prevalent in individuals with diabetes, can be related to a worse prognosis for the disease and can be impacted by changes in the intestinal microbiome. A suggested treatment that may be effective in controlling diabetes and improving sleep quality through increased metabolic regulation is probiotic supplementation. Scientific evidence has shown a relationship between the use of probiotics and improvements in sleep, in glucose concentrations, and in the levels of high density lipoprotein cholesterol. We suggest that probiotic supplementation can play an important role in the management of diabetes and sleep disorders in diabetic patients. Further randomized clinical studies should be undertaken to better understand the impact and effectiveness of the use of probiotics in improving sleep and controlling diabetes.

Effects of sleep restriction during pregnancy on lipids and glucose homeostasis of female offspring submitted to ovariectomy.

Sleep shortening during pregnancy may alter the mother's environment, affecting the offspring. Thus, the present study evaluated the metabolic profile of female offspring from sleep-restricted rats during the last week of pregnancy. Pregnant Wistar rats were distributed into two groups: control (C) and sleep restriction (SR). The SR was performed 20 h/day, from 14th to 20th day of pregnancy. At 2 months, half of the offspring were subjected to ovariectomy (OVX); the others, to sham surgery. Studied groups were Csham, Covx, SRsham and SRovx. Cholesterol (HDL, LDL and C-total), triglycerides (TG) and glucose and insulin tolerance tests (GTT-ITT) were evaluated at 8 months. RSsham presented higher values of TG, while SRovx presented higher TG, LDL and C-total. Basal glucose concentration was increased in SRsham and SRovx. These data suggest that SR during pregnancy may be a risk factor for the development of diseases in adult female offspring.

Fish Oil Supplementation Reduces Heart Levels of Interleukin-6 in Rats with Chronic Inflammation due to Epilepsy.

Sudden unexpected death in epilepsy (SUDEP) is a major cause of premature death related to epilepsy. The causes of SUDEP remain unknown, but cardiac arrhythmias and asphyxia have been suggested as a major mechanism of this event. Inflammation has been implicated in the pathogenesis of both epilepsy and ventricular arrhythmia, with interleukin-6 (IL-6) being recognized as a crucial orchestrator of inflammatory states. Our group previously reported that levels of IL-6 were increased in the hearts of epileptic rats. In this scenario, anti-inflammatory actions are among the beneficial effects of fish oil dietary supplementation. This investigation revealed that elevated levels of IL-6 in the heart were markedly reduced in epileptic rats that were treated in the long-term with fish oil, suggesting protective anti-inflammatory actions against dangerously high levels of IL-6. Based on these findings, our results suggest beneficial effects of long-term intake of fish oil in reducing the inflammation associated with chronic epilepsy.

L-Arginine supplementation improves insulin sensitivity and beta cell function in the offspring of diabetic rats through AKT and PDX-1 activation.

Maternal hyperglycemia can result in defects in glucose metabolism and pancreatic ß-cell function in offspring. The purpose of this study was to evaluate the impact of maternal diabetes mellitus on pancreatic islets, muscle and adipose tissue of the offspring, with or without oral l-Arginine supplementation. The induction of diabetes was performed using streptozotocin (60mg/kg). Animals were studied at 3 months of age and treatment (sucrose or l-Arginine) was administered from weaning. We observed that l-Arg improved insulin sensitivity in the offspring of diabetic mothers (DA), reflected by higher insulin-induced phosphorylation of Akt in muscle and adipose tissue. Insulin resistance is associated with increased oxidative stress and the NADPH oxidase enzyme plays an important role. Our results showed that the augmented interaction of p47PHOX with gp91PHOX subunits of the enzyme in skeletal muscle tissue in the offspring of diabetic rats (DV) was abolished after l-Arg treatment in DA rats. Maternal diabetes caused alterations in the islet functionality of the offspring leading to increased insulin secretion at both low (2.8mM) and high (16.7mM) concentrations of glucose. l-Arg reverses this effect, suggesting that it may be an important modulator in the insulin secretory process. In addition it is possible that l-Arg exerts its effects directly onto essential molecules for the maintenance and survival of pancreatic islets, decreasing protein expression of p47PHOX while increasing Akt phosphorylation and PDX-1 expression. The mechanism by which l-Arg exerts its beneficial effects may involve nitric oxide bioavailability since treatment restored NO levels in the pancreas.

Voltage-Gated Proton Channel in Human Glioblastoma Multiforme Cells.

Solid tumors tend to have a more glycolytic metabolism leading to an accumulation of acidic metabolites in their cytosol, and consequently, their intracellular pH (pHi) turns critically lower if the cells do not handle the acid excess. Recently, it was proposed that the voltage gated proton channels (HV1) can regulate the pHi in several cancers. Here we report the functional expression of voltage gated proton channels in a human glioblastoma multiforme (GBM) cell line, the most common and lethal brain tumor. T98G cells presented an outward, slow activating voltage-dependent proton current, which was also ΔpH-dependent and inhibited by ZnCl2, characterizing it as being conducted by HV1 channels. Furthermore, blocking HV1 channels with ZnCl2 significantly reduced the pHi, cell survival, and migration, indicating an important role for HV1 for tumor proliferation and progression in GBM. Overall, our results suggest that HV1 channels can be a new therapeutic target for GBM.

Myeloid-derived suppressor cells and associated events in urethane-induced lung cancer.

OBJECTIVES: Myeloid-derived suppressor cells contribute to the immunosuppressive microenvironment during tumor development and limit the efficacy of cancer immunotherapy. Identifying myeloid-derived suppressor cells and associated factors is the first step in creating strategies to reverse the suppressive effects of these cells on the immune system. METHODS: To induce lung cancer, we administered 2 doses of urethane to BALB/c mice and observed these animals for 120 days. After this period, we evaluated the percentage of myeloid-derived suppressor cells in the blood, lung and bone marrow. The expression of alpha-smooth muscle actin, transforming growth factor-ß, Toll-like receptor 2, Toll-like receptor 4, and interleukin-6 was also determined in the lung tissue. RESULTS: Myeloid-derived suppressor cells were increased in all evaluated tissues after lung cancer development in association with increased Toll-like receptor 4 expression and decreased interleukin-6 expression in the lung. We observed alpha-smooth muscle actin and transforming growth factor-ß expression in lung nodules. CONCLUSIONS: We believe that the early diagnosis of cancer through determining the blood levels of myeloid-derived suppressor cells followed by the depletion of these cells should be further investigated as a possible approach for cancer treatment.

Myeloid-derived suppressor cells and associated events in urethane-induced lung cancer

OBJECTIVES: Myeloid-derived suppressor cells contribute to the immunosuppressive microenvironment during tumor development and limit the efficacy of cancer immunotherapy. Identifying myeloid-derived suppressor cells and associated factors is the first step in creating strategies to reverse the suppressive effects of these cells on the immune system. METHODS: To induce lung cancer, we administered 2 doses of urethane to BALB/c mice and observed these animals for 120 days. After this period, we evaluated the percentage of myeloid-derived suppressor cells in the blood, lung and bone marrow. The expression of alpha-smooth muscle actin, transforming growth factor-β, Toll-like receptor 2, Toll-like receptor 4, and interleukin-6 was also determined in the lung tissue. RESULTS: Myeloid-derived suppressor cells were increased in all evaluated tissues after lung cancer development in association with increased Toll-like receptor 4 expression and decreased interleukin-6 expression in the lung. We observed alpha-smooth muscle actin and transforming growth factor-β expression in lung nodules. CONCLUSIONS: We believe that the early diagnosis of cancer through determining the blood levels of myeloid-derived suppressor cells followed by the depletion of these cells should be further investigated as a possible approach for cancer treatment. .

Oleic, linoleic and linolenic acids increase ros production by fibroblasts via NADPH oxidase activation.

The effect of oleic, linoleic and γ-linolenic acids on ROS production by 3T3 Swiss and Rat 1 fibroblasts was investigated. Using lucigenin-amplified chemiluminescence, a dose-dependent increase in extracellular superoxide levels was observed during the treatment of fibroblasts with oleic, linoleic and γ-linolenic acids. ROS production was dependent on the addition of ß-NADH or NADPH to the medium. Diphenyleneiodonium inhibited the effect of oleic, linoleic and γ-linolenic acids on fibroblast superoxide release by 79%, 92% and 82%, respectively. Increased levels of p47 (phox) phosphorylation due to fatty acid treatment were detected by Western blotting analyses of fibroblast proteins. Increased p47 (phox) mRNA expression was observed using real-time PCR. The rank order for the fatty acid stimulation of the fibroblast oxidative burst was as follows: γ-linolenic > linoleic > oleic. In conclusion, oleic, linoleic and γ-linolenic acids stimulated ROS production via activation of the NADPH oxidase enzyme complex in fibroblasts.

Increased renal sympathetic nerve activity leads to hypertension and renal dysfunction in offspring from diabetic mothers.

The exposure of the fetus to a hyperglycemic environment promotes the development of hypertension and renal dysfunction in the offspring at adult age. We evaluated the role of renal nerves in the hypertension and renal changes seen in offspring of diabetic rats. Diabetes was induced in female Wistar rats (streptozotocin, 60 mg/kg ip) before mating. Male offspring from control and diabetic dams were studied at an age of 3 mo. Systolic blood pressure measured by tail cuff was increased in offspring of diabetic dams (146 ± 1.6 mmHg, n = 19, compared with 117 ± 1.4 mmHg, n = 18, in controls). Renal function, baseline renal sympathetic nerve activity (rSNA), and arterial baroreceptor control of rSNA were analyzed in anesthetized animals. Glomerular filtration rate, fractional sodium excretion, and urine flow were significantly reduced in offspring of diabetic dams. Two weeks after renal denervation, blood pressure and renal function in offspring from diabetic dams were similar to control, suggesting that renal nerves contribute to sodium retention in offspring from diabetic dams. Moreover, basal rSNA was increased in offspring from diabetic dams, and baroreceptor control of rSNA was impaired, with blunted responses to infusion of nitroprusside and phenylephrine. Thus, data from this study indicate that in offspring from diabetic mothers, renal nerves have a clear role in the etiology of hypertension; however, other factors may also contribute to this condition.

Bradykinin inhibits hepatic gluconeogenesis in obese mice.

The kallikrein-kinin system (KKS) has been previously linked to glucose homeostasis. In isolated muscle or fat cells, acute bradykinin (BK) stimulation was shown to improve insulin action and increase glucose uptake by promoting glucose transporter 4 translocation to plasma membrane. However, the role for BK in the pathophysiology of obesity and type 2 diabetes remains largely unknown. To address this, we generated genetically obese mice (ob/ob) lacking the BK B2 receptor (obB2KO). Despite similar body weight or fat accumulation, obB2KO mice showed increased fasting glycemia (162.3 ± 28.2 mg/dl vs 85.3 ± 13.3 mg/dl), hyperinsulinemia (7.71 ± 1.75 ng/ml vs 4.09 ± 0.51 ng/ml) and impaired glucose tolerance when compared with ob/ob control mice (obWT), indicating insulin resistance and impaired glucose homeostasis. This was corroborated by increased glucose production in response to a pyruvate challenge. Increased gluconeogenesis was accompanied by increased hepatic mRNA expression of forkhead box protein O1 (FoxO1, four-fold), peroxisome proliferator-activated receptor gamma co-activator 1-alpha (seven-fold), phosphoenolpyruvate carboxykinase (PEPCK, three-fold) and glucose-6-phosphatase (eight-fold). FoxO1 nuclear exclusion was also impaired, as the obB2KO mice showed increased levels of this transcription factor in the nucleus fraction of liver homogenates during random feeding. Intraportal injection of BK in lean mice was able to decrease the hepatic mRNA expression of FoxO1 and PEPCK. In conclusion, BK modulates glucose homeostasis by affecting hepatic glucose production in obWT. These results point to a protective role of the KKS in the pathophysiology of type 2 diabetes mellitus.

Central but not peripheral glucoprivation is impaired in monosodium glutamate-treated rats.

In the present study, newborn male Wistar rats were injected, subcutaneously, five times, every other day, with monosodium glutamate (MSG, 4 g/kg bw) or saline (as control, C), during the neonatal period. MSG animals developed destruction of the arcuate nuclei (ARC) with absence of NPY-immunoreactive cell bodies, which impaired both the food intake (baseline) and the 2-deoxy-D-glucose (2DG) glucoprivic feeding response. Increases in the immunoreactivity of corticotropin-releasing hormone-cell bodies in the paraventricular nuclei might have developed to compensate for the atrophy of the pituitary in MSG-treated rats. After systemic 2DG injection, neither the C nor the MSG rats increased their food intake, but they showed similar hyperglycemic responses, whereas plasma free fatty acids (FFA) increased only in the C group. In other groups, 2DG, norepinephrine (NE), neostigmine (NEO) and saline were intracerebroventricularly (i.c.v.) administered. In this condition, impairment of the hyperglycemic and food intake responses, associated to a lower increase in plasma FFA levels, were observed. As opposed to this, the MSG treatment gives support to NE effects, enhancing food intake, as well as plasma glucose and FFA levels. After NEO, plasma glucose increased only in the MSG group, while plasma FFA levels were elevated in the C rats. Taken together, the results obtained after MSG treatment point to a separate neural control of the hyperglycemic response and of the lipid mobilization when stimulated by central 2DG, NE or NEO administration. It seems likely that the excitatory neural pathway that controls lipid metabolism and is present in C rats was destroyed by the MSG treatment.

Western diet modulates insulin signaling, c-Jun N-terminal kinase activity, and insulin receptor substrate-1ser307 phosphorylation in a tissue-specific fashion.

The mechanisms by which diet-induced obesity is associated with insulin resistance are not well established, and no study has until now integrated, in a temporal manner, functional insulin action data with insulin signaling in key insulin-sensitive tissues, including the hypothalamus. In this study, we evaluated the regulation of insulin sensitivity by hyperinsulinemic-euglycemic clamp procedures and insulin signaling, c-jun N-terminal kinase (JNK) activation and insulin receptor substrate (IRS)-1(ser307) phosphorylation in liver, muscle, adipose tissue, and hypothalamus, by immunoprecipitation and immunoblotting, in rats fed on a Western diet (WD) or control diet for 10 or 30 d. WD increased visceral adiposity, serum triacylglycerol, and insulin levels and reduced whole-body glucose use. After 10 d of WD (WD10) there was a decrease in IRS-1/phosphatidylinositol 3-kinase/protein kinase B pathway in hypothalamus and muscle, associated with an attenuation of the anorexigenic effect of insulin in the former and reduced glucose transport in the latter. In WD10, there was an increased glucose transport in adipose tissue in parallel to increased insulin signaling in this tissue. After 30 d of WD, insulin was less effective in suppressing hepatic glucose production, and this was associated with a decrease in insulin signaling in the liver. JNK activity and IRS-1(ser307) phosphorylation were higher in insulin-resistant tissues. In summary, the insulin resistance induced by WD is tissue specific and installs first in hypothalamus and muscle and later in liver, accompanied by activation of JNK and IRS-1(ser307) phosphorylation. The impairment of the insulin signaling in these tissues, but not in adipose tissue, may lead to increased adiposity and insulin resistance in the WD rats.

Regulation of insulin receptor substrate-2 tyrosine phosphorylation in animal models of insulin resistance.

Insulin induces a wide variety of growth and metabolic responses in many cell types. These actions are initiated by insulin binding to its receptor and involve a series of alternative and complementary pathways created by the multiple substrates of the insulin receptor (insulin receptor substrates [IRSs]). We investigated IRS-1 and IRS-2 tyrosine phosphorylation; their association with phosphatidylinositol-3-OH kinase (PI3-K); and the phosphorylation of Akt, a serine-threonine kinase situated downstream of PI3-K, in liver and muscle of two animal models of insulin resistance: epinephrine- or dexamethasone-treated rats. We used in vivo insulin infusion followed by tissue extraction, immunoprecipitation, and immunoblotting. IRS-1 and IRS-2 protein expression did not change in liver and muscle of the epinephrine- treated rats, but in dexamethasone-treated rats IRS-1 presented an increase in liver and a decrease in muscle tissue. PI3-K and Akt protein expression did not change in liver or muscle of the two animal models of insulin resistance. There was a downregulation in insulin- induced IRS-1 and IRS-2 tyrosine phosphorylation and association with PI3-K in both models of insulin resistance. In parallel, insulin-induced Akt phosphorylation was reduced in both tissues of epinephrine-treated rats, and in liver but not in muscle of dexamethasonetreated rats. The reduction in insulin-induced Akt phosphorylation may help to explain the insulin resistance in liver and muscle of epinephrine-treated rats and in the liver of dexamethasone-treated rats.

Modulation of IR/PTP1B interaction and downstream signaling in insulin sensitive tissues of MSG-rats.

PTP1B has been shown to be a negative regulator of the insulin signal transduction in insulin resistant states. Herein we investigated IR/PTP1B interaction and downstream signaling in insulin sensitive tissues of 10 and 28-week-old MSG-insulin resistant rats which represent different stages of insulin resistance. Our results demonstrated that the increase in PTP1B expression and/or association with IR in MSG animals may contribute to the impaired insulin signaling mainly in liver and muscle. Although, adipose tissue of 10-week-old MSG rats showed higher PTP1B expression and IR/PTP1B interaction, they were not sufficient to impair all insulin signaling since IRS-2 phosphorylation and association with PI3-kinase and Akt serine phosphorylation were increased, which may contribute for the increased adiposity of these animals. In 28-week-old-MSG rats there was an increase in IR/PTP1B interaction and reduced insulin signaling in liver, muscle and adipocytes, and a more pronounced insulin resistance.