CLK2 in GABAergic neurons is critical in regulating energy balance and anxiety-like behavior in a gender-specific fashion.

Introduction: Cdc2-like kinase (CLK2) is a member of CLK kinases expressed in hypothalamic neurons and is activated in response to refeeding, leptin, or insulin. Diet-induced obesity and leptin receptor-deficient db/db mice lack CLK2 signal in the hypothalamic neurons. The neurotransmiter gamma-aminobutyric acid (GABA) is among the most prevalent in the central nervous system (CNS), particularly in the hypothalamus. Given the abundance of GABA-expressing neurons and their potential influence on regulating energy and behavioral homeostasis, we aimed to explore whether the deletion of CLK2 in GABAergic neurons alters energy homeostasis and behavioral and cognitive functions in both genders of mice lacking CLK2 in Vgat-expressing neurons (Vgat-Cre; Clk2loxP/loxP) on chow diet. Methods: We generated mice lacking Clk2 in Vgat-expressing neurons (Vgat-Cre; Clk2loxP/loxP) by mating Clk2loxP/loxP mice with Vgat-IRES-Cre transgenic mice and employed behavior, and physiological tests, and molecular approaches to investigate energy metabolism and behavior phenotype of both genders. Results and discussion: We showed that deletion of CLK2 in GABAergic neurons increased adiposity and food intake in females. The mechanisms behind these effects were likely due, at least in part, to hypothalamic insulin resistance and upregulation of hypothalamic Npy and Agrp expression. Besides normal insulin and pyruvate sensitivity, Vgat-Cre; Clk2loxP/loxP females were glucose intolerant. Male Vgat-Cre; Clk2loxP/loxP mice showed an increased energy expenditure (EE). Risen EE may account for avoiding weight and fat mass gain in male Vgat-Cre; Clk2loxP/loxP mice. Vgat-Cre; Clk2loxP/loxP mice had no alteration in cognition or memory functions in both genders. Interestingly, deleting CLK2 in GABAergic neurons changed anxiety-like behavior only in females, not males. These findings suggest that CLK2 in GABAergic neurons is critical in regulating energy balance and anxiety-like behavior in a gender-specific fashion and could be a molecular therapeutic target for combating obesity associated with psychological disorders in females.

Acute exercise reduces feeding by activating IL-6/Tubby axis in the mouse hypothalamus.

Background: Acute exercise contributes to decreased feeding through leptin and interleukin/Janus kinase 2/signal transducers and activators of transcription 3 (IL-6/JAK2/STAT3) signaling. Considering the pleiotropic use of substrates by JAK2 and that JAK2 can phosphorylate the Tubby protein (TUB) in CHO-IR cells, we speculated that acute exercise can activate the IL-6/JAK2/TUB pathway to decrease food intake. Aims: We investigated whether acute exercise induced tyrosine phosphorylation and the association of TUB and JAK2 in the hypothalamus and if IL-6 is involved in this response, whether acute exercise increases the IL-6/TUB axis to regulate feeding, and if leptin has an additive effect over this mechanism. Methods: We applied a combination of genetic, pharmacological, and molecular approaches. Key findings: The in vivo experiments showed that acute exercise increased the tyrosine phosphorylation and association of JAK2/TUB in the hypothalamus, which reduced feeding. This response was dependent on IL-6. Leptin had no additive effect on this mechanism. Significance: The results of this study suggest a novel hypothalamic pathway by which IL-6 released by exercise regulates feeding and reinforces the beneficial effects of exercise.

Gut Microbiota Modifications and Weight Regain in Morbidly Obese Women After Roux-en-Y Gastric Bypass.

INTRODUCTION: The relationship between late post-bariatric surgery weight regain and gut microbiota is not completely understood. OBJECTIVE: To analyze the profile of gut microbiota among patients with and without late weight regain after post-Roux-en-Y gastric bypass (RYGB) and to compare it with a control group (CG) comprised of obese Brazilian individuals. METHODS: This is a cross-sectional study which enrolled 34 morbidly obese women divided into 3 groups: post-Roux-en-Y gastric bypass without (RYGB_non-regain), and with weight regain (RYGB_regain) at least 5 years after surgery, and a CG of preoperative individuals. Gut microbiota was determined by metagenomic analyses. RESULTS: The alpha diversity was higher in groups RYGB non-regain and RYGB regain when compared with CG (p < 0.05). Both RYGB non-regain and RYGB regain groups showed a lower abundance of the phylum Bacteroidetes when compared with CG (p < 0.01). The genera Bacteroides and SMB53 were increased in CG (p < 0.05). Group RYGB non-regain showed more abundance of the Akkermansia genus when compared with CG and group RYGB regain (p < 0.05). RYGB non-regain showed a greater abundance of the Phascolarctobacterium genus and lower of the SMB53 genus when compared with CG (p < 0.05). RYGB non-regain showed a greater abundance of the Phascolarctobacterium genus and a lower of the SMB53 genus when compared with CG (p < 0.05). CONCLUSION: The gut microbiota of individuals which presented late weight regain after RYGB was significantly different in comparison to individuals with a successful weight loss, a finding that points towards a significant role of gut microbiota on weight loss and maintenance after surgery.

Microbiota determines insulin sensitivity in TLR2-KO mice.

INTRODUCTION: Environmental factors have a key role in the control of gut microbiota and obesity. TLR2 knockout (TLR2-/-) mice in some housing conditions are protected from diet-induced insulin resistance. However, in our housing conditions these animals are not protected from diet-induced insulin-resistance. AIM: The aim of the present study was to investigate the influence of our animal housing conditions on the gut microbiota, glucose tolerance and insulin sensitivity in TLR2-/- mice. MATERIAL AND METHODS: The microbiota was investigated by metagenomics, associated with hyperinsulinemic euglycemic clamp and GTT associated with insulin signaling through immunoblotting. RESULTS: The results showed that TLR2-/- mice in our housing conditions presented a phenotype of metabolic syndrome characterized by insulin resistance, glucose intolerance and increase in body weight. This phenotype was associated with differences in microbiota in TLR2-/- mice that showed a decrease in the Proteobacteria and Bacteroidetes phyla and an increase in the Firmicutesphylum, associated with and in increase in the Oscillospira and Ruminococcus genera. Furthermore there is also an increase in circulating LPS and subclinical inflammation in TLR2-/-. The molecular mechanism that account for insulin resistance was an activation of TLR4, associated with ER stress and JNK activation. The phenotype and metabolic behavior was reversed by antibiotic treatment and reproduced in WT mice by microbiota transplantation. CONCLUSIONS: Our data show, for the first time, that the intestinal microbiota can induce insulin resistance and obesity in an animal model that is genetically protected from these processes.

miRNAs, target genes expression and morphological analysis on the heart in gestational protein-restricted offspring.

Gestational protein restriction was associated with low birth weight, hypertension and higher prevalence of cardiac disorders in adults. Several mechanisms, including epigenetics, could be related with the cardiovascular phenotype on protein-restricted offspring. Thus, we investigated the morphological cardiac effects of gestational protein restriction and left ventricle miRNAs and target genes expression pattern in both 12-day and 16-week old gestational protein-restricted male offspring. Pregnant Wistar rats were allocated into two groups, according to protein supply during pregnancy: NP (normal protein diet- 17%) or LP (low protein diet-6%). Dams on the gestational protein-restricted diet had lower body weight gain and higher food intake. Gestational protein-restricted offspring had low birth weight, followed by rapidly body weight recovery, hypertension, and increased myocytes cross-sectional area and collagen fraction at 16-week old age. At 12-days old, miR-184, miR-192, miR-376c, miR-380-3p, miR-380-5p, miR-451, and miR-582-3p had increased expression, and miR-547 and miR-743a had decreased expression in the gestational protein-restricted left ventricle. At 16-week old, let-7b, miR-125a-3p, miR-142-3p, miR-182 and miR-188-5p had increased expression and let-7g, miR-107, miR-127, miR-181a, miR-181c, miR-184, miR-324-5p, miR-383, miR-423-5p and miR-484 had decreased expression in gestational protein-restricted left ventricle. Target predicted gene expression analysis showed higher expression of Dnmt3a, Oxct1, Rictor and Trps1 and lower expression of Bbs1 and Calml3 in 12-day old protein-restricted offspring. 16-week old protein-restricted offspring had higher expression of Adrbk1, Bbs1, Dnmt3a, Gpr22, Inppl1, and Oxct1 genes. In conclusion, gestational protein restriction was related to offspring low birth weight, increased systolic blood pressure and morphological heart alterations that could be related to early heart miRNA expression changes that perpetuate into adulthood and which are associated with the regulation of essential genes involved in cardiovascular development, heart morphology, function, and metabolism.

Oral Glutamine Supplementation Reduces Obesity, Pro-Inflammatory Markers, and Improves Insulin Sensitivity in DIO Wistar Rats and Reduces Waist Circumference in Overweight and Obese Humans.

In the present study, we aimed to investigate whether chronic oral glutamine (Gln) supplementation may alter metabolic parameters and the inflammatory profile in overweight and obese humans as well as whether Gln may modulate molecular pathways in key tissues linked to the insulin action in rats. Thirty-nine overweight/obese volunteers received 30 g of Gln or alanine (Ala-control) for 14 days. Body weight (BW), waist circumference (WC), hormones, and pro-inflammatory markers were evaluated. To investigate molecular mechanisms, Gln or Ala was given to Wistar rats on a high-fat diet (HFD), and metabolic parameters, euglycemic hyperinsulinemic clamp with tracers, and Western blot were done. Gln reduced WC and serum lipopolysaccharide (LPS) in overweight volunteers. In the obese group, Gln diminished WC and serum insulin. There was a positive correlation between the reduction on WC and LPS. In rats on HFD, Gln reduced adiposity, improved insulin action and signaling, and reversed both defects in glucose metabolism in the liver and muscle. Gln supplementation increased muscle glucose uptake and reversed the increased hepatic glucose production, in parallel with a reduced glucose uptake in adipose tissue. This insulin resistance in AT was accompanied by enhanced IRS1 O-linked-glycosamine association in this tissue, but not in the liver and muscle. These data suggest that Gln supplementation leads to insulin resistance specifically in adipose tissue via the hexosamine pathway and reduces adipose mass, which is associated with improvement in the systemic insulin action. Thus, further investigation with Gln supplementation should be performed for longer periods in humans before prescribing as a beneficial therapeutic approach for individuals who are overweight and obese.

Impact of the length of vitamin D deficiency on cardiac remodeling.

BACKGROUND: This study was aimed to evaluate the influence of vitamin D (VD) deficiency on cardiac metabolism, morphology, and function. Thus, we investigated the relationship of these changes with the length of the nutrient restriction. METHODS AND RESULTS: Male weanling Wistar rats were allocated into 4 groups: C2 (n=24), animals were fed an AIN-93G diet with 1000 IU VD/kg of chow and were kept under fluorescent light for 2 months; D2 (n=22), animals were fed a VD-deficient AIN-93G diet and were kept under incandescent light for 2 months; C4 (n=21) animals were kept in the same conditions of C2 for 4 months; and D4 (n=23) animals were kept in the same conditions of D2 for 4 months. Biochemical analyses showed lower ß-hydroxyacyl coenzyme-A dehydrogenase activity and higher lactate dehydrogenase activity in VD-deficient animals. Furthermore, VD deficiency was related to increased cytokines release, oxidative stress, apoptosis, and fibrosis. Echocardiographic data showed left ventricular hypertrophy and lower fractional shortening and ejection fraction in VD-deficient animals. Difference became evident in the lactate dehydrogenase activity, left ventricular weight, right ventricle weight, and left ventricular mass after 4 months of VD deficiency. CONCLUSIONS: Our data indicate that VD deficiency is associated with energetic metabolic changes, cardiac inflammation, oxidative stress, fibrosis and apoptosis, cardiac hypertrophy, left chambers alterations, and systolic dysfunction. Furthermore, length of the restriction influenced these cardiac changes.

N-acetylcysteine in high-sucrose diet-induced obesity: energy expenditure and metabolic shifting for cardiac health.

To study the effects of N-acetylcysteine (NAC, C(5)H(9)-NO(3)S) on high-sucrose diet-induced obesity and its effects on energy metabolism and cardiac oxidative stress, male Wistar 24 rats were divided into four groups (n=6): (C) given standard chow and water; (N) receiving standard chow and 2g/l N-acetylcysteine in its drinking water; (HS) given standard chow and 30% sucrose in its drinking water, and (HS-N) receiving standard chow, 30% sucrose and N-acetylcysteine in its drinking water. After 30 days of the treatment, obesity was evidenced in HS rats from enhanced body weight, respiratory quotient, hypertriglyceridemia. As well depressed resting metabolic rate, and oxygen consumption per surface area. HS rats had triacylglycerol accumulation, oxidative stress and metabolic shifting in cardiac tissue. NAC enhanced fat oxidation and energy expenditure, normalizing these adverse effects, comparing HS-N and HS rats. The beta-hydroxyacyl coenzymne-A dehydrogenase activity was higher in HS-N animals, indicating higher heart fatty acid degradation than in HS. NAC normalized myocardial glycogen and lactate dehydrogenase activity, comparing HS-N and HS rats, but had no effects on calorimetric and biochemical parameters in standard-fed rats, comparing N and C groups. In conclusion, N-acetylcysteine offers promising therapeutic value in prevention of high-sucrose induced-obesity and its effect on cardiac tissue. N-acetylcysteine reduced the oxidative stress and prevented the metabolic shifting in cardiac tissue, enhancing fatty acid oxidation and reducing anaerobic metabolism in high-sucrose-fed conditions. The application of this agent in food system via exogenous addition may be feasible and beneficial for antioxidant protection and energy metabolism in cardiac tissue.