Early weaning induces short- and long-term effects on pancreatic islets in Wistar rats of both sexes.

KEY POINTS: The World Health Organization recommends exclusive breastfeeding until 6 months of age as an important strategy to reduce child morbidity and mortality. Studies have associated early weaning with the development of obesity and type 2 diabetes in adulthood. In our model, we demonstrated that early weaning leads to increased insulin secretion in adolescent males and reduced insulin secretion in adult offspring. Early weaned males exhibit insulin resistance in skeletal muscle. Early weaning did not change insulin signalling in the muscle of female offspring. Taking into account that insulin resistance is one of the primary factors for the development of type 2 diabetes mellitus, this work demonstrates the importance of breastfeeding in the fight against this disease. ABSTRACT: Early weaning (EW) leads to short- and long-term obesity and diabetes. This phenotype is also observed in experimental models, in which early-weaned males exhibit abnormal insulinaemia in adulthood. However, studies regarding the effect of EW on pancreatic islets are rare. We investigated the mechanisms by which glycaemic homeostasis is altered in EW models through evaluations of insulin secretion and its signalling pathway in offspring. Lactating Wistar rats and their pups were divided into the following groups: non-pharmacological EW (NPEW): mothers were wrapped with an adhesive bandage on the last 3 days of lactation; pharmacological EW (PEW): mothers received bromocriptine to inhibit prolactin (1 mg/kg body mass/day) on the last 3 days of lactation; and control (C): pups underwent standard weaning at PN21. Offspring of both sexes were euthanized at PN45 and PN180. At PN45, EW males showed higher insulin secretion (vs. C). At PN170, PEW males exhibited hyperglycaemia in an oral glucose tolerance test (vs. C and NPEW). At PN180, EW male offspring were heavier; however, both sexes showed higher visceral fat. Insulin secretion was lower in EW offspring of both sexes. Males from both EW groups had lower glucokinase in islets, but unexpectedly, PEW males showed higher GLUT2, than did C. EW males exhibited lower insulin signalling in muscle. EW females exhibited no changes in these parameters compared with C. We demonstrated distinct alterations in the insulin secretion of EW rats at different ages. Despite the sex dimorphism in insulin secretion in adolescence, both sexes showed impaired insulin secretion in adulthood due to EW.

Body Adiposity and Endocrine Profile of Female Wistar Rats of Distinct Ages that were Early Weaned.

Early weaning (EW) is a risk factor for metabolic syndrome. Male rats that were precociously weaned present neonatal malnutrition and, in adulthood, developed overweight, accumulation of body fat, dyslipidemia, changes in glycemic homeostasis, hyperleptinemia, and increase of vitamin D. As metabolic profile of early-weaned females is not known, we investigated the endocrine-metabolic parameters in adolescence and adult female rats of 2 different EW models. Wistar lactating rats and pups from both sexes were separated into 3 groups: non-pharmacological EW (NPEW), dams were involved with a bandage interrupting suckling in the last 3 days of lactation; pharmacological EW (PEW), dams were bromocriptine-treated (0.5 mg/twice a day via intraperitoneal injection) for 3 days before weaning; and control, dams whose pups ate milk throughout lactation. At 21 days-old, NPEW and PEW females had lower body weight. At 180 days-old, NPEW and PEW females showed higher feed efficiency, weight gain, body fat percentage, and greater accumulation of gonadal and retroperitoneal fat depots associated with adipocyte hypertrophy. NPEW females also showed hyperphagia. Only NPEW females presented hyperleptinemia. Plasma thyroid hormones and vitamin D were unchanged among EW females. Regarding sex hormones, at 45 days-old, no change was found in EW females, while at 180 days-old, PEW females had hypoestrogenemia. EW increases the risk for obesity in female rats in adulthood, as already demonstrated for males, although through distinct mechanisms involving some hormones.

Maternal Roux-en-Y gastric bypass impairs insulin action and endocrine pancreatic function in male F1 offspring.

PURPOSE: Obesity is predominant in women of reproductive age. Roux-en-Y gastric bypass (RYGB) is the most common bariatric procedure that is performed in obese women for weight loss and metabolic improvement. However, some studies suggest that this procedure negatively affects offspring. Herein, using Western diet (WD)-obese female rats, we investigated the effects of maternal RYGB on postnatal body development, glucose tolerance, insulin secretion and action in their adult male F1 offspring. METHODS: Female Wistar rats consumed a Western diet (WD) for 18 weeks, before being submitted to RYGB (WD-RYGB) or SHAM (WD-SHAM) operations. After 5 weeks, WD-RYGB and WD-SHAM females were mated with control male breeders, and the F1 offspring were identified as: WD-RYGB-F1 and WD-SHAM-F1. RESULTS: The male F1 offspring of WD-RYGB dams exhibited decreased BW, but enhanced total nasoanal length gain. At 120 days of age, WD-RYGB-F1 rats displayed normal fasting glycemia and glucose tolerance but demonstrated reduced insulinemia and higher glucose disappearance after insulin stimulus. In addition, these rodents presented insulin resistance in the gastrocnemius muscle and retroperitoneal fat, as judged by lower Akt phosphorylation after insulin administration, but an increase in this protein in the liver. Finally, the islets from WD-RYGB-F1 rats secreted less insulin in response to glucose and displayed increased ß-cell area and mass. CONCLUSIONS: RYGB in WD dams negatively affected their F1 offspring, leading to catch-up growth, insulin resistance in skeletal muscle and white fat, and ß-cell dysfunction. Therefore, our data are the first to demonstrate that the RYGB in female rats may aggravate the metabolic imprinting induced by maternal WD consumption, in their male F1 descendants. However, since we only used male F1 rats, further studies are necessary to demonstrate if such effect may also occur in female F1 offspring from dams that underwent RYGB operation.

Maternal exposure to tributyltin alters the breast milk, hormonal profile, and thyroid morphology of dams and induces sex-specific changes in neonate rat offspring.

Tributyltin (TBT) is the chemical substance commonly used worldwide to prevent biofouling of vessels. Due to its ability to bioaccumulate and biomagnify, even after being banned, significant concentrations of TBT can be detected in sediment, affecting marine and human life. Although studies have shown that direct exposure to TBT alters physiological parameters in mammals, the relationship between exposure to TBT during pregnancy and lactation, considered critical windows for metabolic programming, has not been fully elucidated. Our hypothesis is that offspring whose mothers were exposed to TBT during critical stages of development may exhibit dysfunctions in endocrine-metabolic parameters. We used pregnant Wistar rats that were divided into groups and received the following treatments from gestational day 7 until the end of lactation by intragastric gavage: vehicle (ethanol 0.01%; Control), low TBT dose (100 ng/kg of body weight (bw)/day; TBT100ng) and high TBT dose (1000 ng/kg bw/day; TBT1000ng). Dams and offspring at birth and weaning (21 days old) were studied. Maternal exposure to TBT promoted dose-dependent changes in dams. The findings for adiposity, milk composition and lipid profile were more pronounced in TBT100 ng dam; however, thyroid morphology was altered in TBT1000 ng dam. Female offspring were differentially affected by the dose of exposure. At birth, females in the TBT100ng group had low body weight, lower naso-anal length (NAL), and higher plasma T4, and at weaning, females in the TBT100ng group had lower insulin and leptin levels. Females in the TBT1000ng group had lower NAL at birth and lower leptinemia and weight of white adipose tissue at weaning. Male offspring from TBT groups showed high T3 at birth, without biometric alterations at birth or weaning. Despite these findings, both sexes exhibited dose-dependent morphological changes in the thyroid gland. Thus, maternal exposure to TBT constitutes an important route of contamination for both dams and offspring.

Maternal low-dose caffeine intake during the perinatal period promotes short- and long-term sex-dependent hormonal and behavior changes in the offspring.

AIM: Maternal caffeine crosses the placenta and mammary barriers, reaching the baby and, because his/her caffeine metabolism is immature, our hypothesis is that even a low caffeine intake (250 mg/day), lower than the dose limit recommended by the World Health Organization, can promote caffeine overexposure in the offspring, leading to short- and long-term changes. MAIN METHODS: Pregnant Wistar rats received intragastric caffeine (CAF) (25 mg/Kg/day) or vehicle during the gestation and lactation periods. We evaluated morphometrical, metabolic, hormonal, and behavioral parameters of male and female offspring at different ages. KEY FINDINGS: Even a low caffeine intake promoted lower maternal body mass and adiposity, higher plasma cholesterol and lower plasma T3, without changes in plasma corticosterone. Female CAF offspring exhibited lower birth weight, body mass gain and food intake throughout life, and hyperinsulinemia at weaning, while male CAF offspring showed reduced food intake and lower plasma T3 at weaning. At puberty and adulthood, male CAF showed higher preference for palatable food, aversion to caffeine intake and higher locomotor activity, while female CAF only showed lower preference for high fat diet (HFD) and lower anxiety-like behavior. At adulthood, both male and female offspring showed higher plasma T3. Male CAF showed hypertestosteronemia, while female CAF showed hypoinsulinemia without effect on glucose tolerance. SIGNIFICANCE: A low caffeine intake during the perinatal period affects rat's offspring development, promoting sex-dependent hormonal and behavior changes. Current data suggest the need to review caffeine recommendations during the perinatal period.

Liraglutide prevents body and fat mass gain in ovariectomized Wistar rats.

Estrogens exert beneficial metabolic effects by reducing food intake and enhancing energy expenditure through both central and peripheral mechanisms. The decrease of estrogen, as occurs in ovariectomy (OVX), leads to metabolic disturbances, such as increased body weight, adipose tissue mass, basal blood glucose, and impaired glucose tolerance. These effects can be reversed by reintroducing estrogen. GLP-1 and its receptor agonists, known for their antihyperglycemic properties, also exhibit anorexigenic effects. Besides that, research indicates that GLP-1 analogs can induce metabolic changes peripherally, such as increased fatty acid oxidation and inhibited lipogenesis. Given the shared metabolic actions of GLP-1 and estrogens, we explored whether liraglutide, a GLP-1 agonist, could mitigate the metabolic effects of estrogen deficiency. We tested this hypothesis using ovariectomized rats, a model that simulates menopausal estrogen deficiency, and treated them with either liraglutide or 17ß-Estradiol benzoate for 21 days. Ovariectomy resulted in elevated DPP-IV activity in both plasma and inguinal white adipose tissue (iWAT). While estrogen replacement effectively countered the DPP-IV increase in both plasma and iWAT, liraglutide only prevented the rise in iWAT DPP-IV activity. Liraglutide prevented body weight and fat mass gain after ovariectomy to the same extent as estradiol treatment. This can be explained by the lower food intake and food efficiency caused by estradiol and liraglutide. However, liraglutide was associated with increased pro-inflammatory cytokines and inflammatory cells in white adipose tissue. Further research is crucial to fully understand the potential benefits and risks of using GLP-1 receptor agonists in the context of menopause.

Citrus aurantium L. and synephrine improve brown adipose tissue function in adolescent mice programmed by early postnatal overfeeding.

Introduction and aims: Obesity is a multifactorial condition with high health risk, associated with important chronic disorders such as diabetes, dyslipidemia, and cardiovascular dysfunction. Citrus aurantium L. (C. aurantium) is a medicinal plant, and its active component, synephrine, a ß-3 adrenergic agonist, can be used for weight loss. We investigated the effects of C. aurantium and synephrine in obese adolescent mice programmed by early postnatal overfeeding. Methods: Three days after birth, male Swiss mice were divided into a small litter (SL) group (3 pups) and a normal litter (NL) group (9 pups). At 30 days old, SL and NL mice were treated with C. aurantium standardized to 6% synephrine, C. aurantium with 30% synephrine, isolated synephrine, or vehicle for 19 days. Results: The SL group had a higher body weight than the NL group. Heart rate and blood pressure were not elevated. The SL group had hyperleptinemia and central obesity that were normalized by C. aurantium and synephrine. In brown adipose tissue, the SL group showed a higher lipid droplet sectional area, less nuclei, a reduction in thermogenesis markers related to thermogenesis (UCP-1, PRDM16, PGC-1α and PPARg), and mitochondrial disfunction. C. aurantium and synephrine treatment normalized these parameters. Conclusion: Our data indicates that the treatment with C. aurantium and synephrine could be a promising alternative for the control of some obesity dysfunction, such as improvement of brown adipose tissue dysfunction and leptinemia.

Low protein diet during lactation programs hepatic metabolism in adult male and female rats.

The liver is an essential regulator of energy metabolism, and its function can be disrupted by nutritional alterations. Since liver development continues during breastfeeding nutritional challenges during this period predispose patients to diseases throughout life. A maternal protein-restricted (PR) diet during lactation promotes reductions in the body weight, adiposity, and plasma glucose and insulin, leptin resistance and an increase in corticosterone and catecholamines in adult male rat offspring. Here, we investigated hepatic metabolism in the offspring (both sexes) of PR (8% protein diet during lactation) and control (23% protein diet) dams. Both male and female offspring were evaluated at 6 months of age. PR males had no liver steatosis and manifested a reduction in lipids in hepatocytes adjacent to the vasculature. These animals had lower levels of esterified cholesterol in hepatocytes, suggesting higher biliary excretion, unchanged glycolysis and gluconeogenesis, and lower contents of the markers of mitochondrial redox balance and endoplasmic reticulum (ER) stress response and estrogen receptor alpha. PR females showed normal hepatic morphology associated with higher uptake of cholesterol esters, normal glycolysis and gluconeogenesis, and lower ER stress parameters without changes in the key markers of the redox balance. Additionally, these animals had lower content of estrogen receptor alpha and higher content of androgen receptor. The maternal PR diet during lactation did not program hepatic lipid accumulation in the adult progeny. However, several repair homeostasis pathways were altered in males and females, possibly compromising maintenance of normal liver function.

Late effects of early weaning on food preference and the dopaminergic and endocannabinoid systems in male and female rats.

Early weaning (EW) is associated with obesity later in life. Here, using an EW model in rats, we investigated changes in feeding behavior and the dopaminergic and endocannabinoid systems (ECS) in the adult offspring. Lactating Wistar rats were divided into two groups: EW, dams were wrapped with a bandage to interrupt suckling during the last 3 days of breastfeeding; CONT; dams fed the pups throughout the period without hindrances. EW animals were compared with CONT animals of the same sex. At PN175, male and female offspring of both groups could freely self-select between high-fat and high-sugar diets (food challenge test). EW males preferred the high-fat diet at 30 min and more of the high-sugar diet after 12 h compared to CONT males. EW females did not show differences in their preference for the palatable diets compared to CONT females. Total intake of standard diet from PN30-PN180 was higher in both male and female EW animals, indicating hyperphagia. At PN180, EW males showed lower type 2 dopamine receptor (D2r) in the nucleus accumbens (NAc) and dorsal striatum, while EW females had lower tyrosine hydroxylase in the ventral tegmental area and NAc, D1r in the NAc, and D2r in the prefrontal cortex. In the lateral hypothalamus, EW males had lower fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase, whereas EW females showed lower N-arachidonoyl-phosphatidylethanolamine phospholipase-D and increased FAAH. Early weaning altered both the dopaminergic and ECS parameters at adulthood, contributing to the eating behavior changes of the progeny in a sex-dependent manner.

Pregnancy and lactation after Roux-en-Y gastric bypass worsen nonalcoholic fatty liver disease in obese rats and lead to differential programming of hepatic de novo lipogenesis in offspring.

Maternal obesity increases the risk of nonalcoholic fatty liver disease (NAFLD) in offspring. The Roux-en-Y gastric bypass (RYBG) is effective for achieving weight loss and ameliorates NAFLD. To determine whether these benefits are maintained after pregnancy and/or lactation, and whether they modulate hepatic morphofunction in the next generation, we evaluated hepatic lipid metabolism in Western diet (WD)-obese female rats that underwent RYGB and in their F1 offspring at adulthood. Female Wistar rats consumed a WD from 21 to 130 days of age, before being submitted to RYGB (WD-RYGB-F0) or SHAM (WD-SHAM-F0) operations. After 5 weeks, these females were mated with control male breeders, and the male and female F1 offspring were identified as WD-RYGB-F1 and WD-SHAM-F1. WD-RYGB-F0 dams exhibited lower serum lipids levels, but severe hepatic steatosis and pathological features of advanced liver injury. The hepatic proteins involved in lipogenesis were reduced in WD-RYGB-F0, as were the genes related to ß-oxidation and bile acids (BAs). Although the female and male WD-RYGB-F1 groups did not exhibit hepatic steatosis, the livers of female WD-RYGB-F1 demonstrated higher amounts of lipogenic genes and proteins, while male WD-RYGB-F1 presented a similar downregulation of lipogenic factors to that seen in WD-RYGB-F0 dams. In contrast, maternal and offspring groups of both sexes displayed reductions in the expressions of genes involved in BAs physiology and gluconeogenesis. As such, RYGB aggravates NAFLD after pregnancy and lactation and induces a gender-dependent differential expression of the hepatic lipogenesis pathway in offspring, indicating that female WD-RYGB-F1 may be an increased risk of developing NAFLD.