Natália da Silva Lima

Resveratrol attenuates oxidative stress and prevents steatosis and hypertension in obese rats programmed by early weaning.

We hypothesized that resveratrol, a natural phytoalexin found in grapes, can prevent oxidative stress, obesity and its related disturbances in obese rats programmed by early weaning. Lactating Wistar rats were separated into two groups: early weaning (EW) - dams who were wrapped with a bandage to interrupt the lactation in the last 3 days of lactation; control - dams whose pups had free access to milk during all lactation. At the 150th day, EW offspring were randomly subdivided into EW+resveratrol (EW+Res) - resveratrol (30 mg/kg/day); EW+vehicle (EW) - rats that received 0.5% (w/v) aqueous methylcellulose. The control group received vehicle. Rats were treated by gavage daily for 30 days. EW offspring developed hyperphagia, higher body weight, visceral obesity, higher systolic (SBP) and diastolic blood pressure (DBP) (+15% and +20%, respectively; P<.05) and higher serum triglycerides (TG) and low-density lipoprotein but lower high-density lipoprotein (+55%, +33% and -13%, respectively; P<.05). Resveratrol normalized food intake, SBP and DBP and prevented obesity and dyslipidemia in EW+Res. EW rats had higher plasma and liver thiobarbituric-acid-reactive substances (TBARS) and lower plasma superoxide dismutase (SOD) and liver glutathione peroxidase activities (+51%, +18%, -58%, -31%, respectively; P<.05), and resveratrol normalized both plasma and liver TBARS and increased the activity of SOD and catalase in plasma. EW rats presented liver steatosis and higher liver TG, and resveratrol prevented these hepatic alterations. In conclusion, this study demonstrated a potential therapeutic use of resveratrol in preventing obesity and oxidative stress and reducing the risk of hypertension, dyslipidemia and steatosis in adult rats programmed by early weaning.

Early weaning causes undernutrition for a short period and programmes some metabolic syndrome components and leptin resistance in adult rat offspring

Maternal malnutrition during lactation programmes for overweight and central leptin resistance in adulthood. The inhibition of lactation by maternal treatment with bromocriptine (a prolactin inhibitor) programmes for obesity, hyperleptinaemia and leptin resistance. Here, we evaluated the short- and long-term effects of early weaning (EW) on body-weight regulation, leptin signalling, and hormone and lipid profiles in rats offspring. Lactating rats were separated into two groups: EW--dams were wrapped with a bandage to interrupt the lactation in the last 3 d of lactation; control--dams whose pups had free access to milk during all lactation (21 d). Data were significant at P < 0·05. At weaning, EW pups presented lower body weight (-10%), length (-4%), visceral fat (-40%), total fat (-30%), serum leptin (-73%), glycaemia (-10%), serum insulin (-20%) and insulin resistance index (IRI; -30 %), but higher total body protein content (+40%). At 180 d, EW offspring showed hyperphagia, higher length (+3%), body weight (+8%), visceral and total fat (+36 and 84%), serum TAG (+96%), glycaemia (+15%), leptinaemia (+185%) and IRI (+29%); however, they showed lower total protein content (-23%), leptin:body fat ratio (41%), prolactinaemia (-38%) and adiponectinaemia (-59%). Despite unchanged leptin receptor (OB-R) and signal transducer and activator of transcription 3 (STAT3), they displayed lower hypothalamic janus tyrosine kinase 2, phosphorylated STAT3 and a higher suppressor of cytokine signalling 3 levels, suggesting a central leptin resistance. Adult rats that were early weaned displayed higher adiposity, insulin resistance and dyslipidaemia, which are related to metabolic syndrome development. Our model reinforces the idea that neonatal malnutrition caused by shortening of the lactation period is important for metabolic programming of future diseases.

Neonatal nicotine exposure causes insulin and leptin resistance and inhibits hypothalamic leptin signaling in adult rat offspring

Maternal nicotine (NIC) exposure during lactation leads to overweight, hyperleptinemia, and hypothyroidism in adult rat offspring. In this model, we analyzed adipocyte morphology, glucose homeostasis (serum insulin and adiponectin; liver and muscle glycogen), serum lipid, and the leptin signaling pathway. After birth, osmotic minipumps were implanted in lactating rats, which were divided into the groups NIC (6 mg/kg per day s.c. for 14 days) and control (C, saline). NIC and C offspring were killed at the age of 180 days. Adult NIC rats showed higher total body fat (+10%, P<0.05), visceral fat mass (+12%, P<0.05), and cross-sectional area of adipocytes (epididymal: +12% and inguinal: +43%, P<0.05). Serum lipid profile showed no alteration except for apolipoprotein AI, which was lower. We detected a lower adiponectin:fat mass ratio (-24%, P<0.05) and higher insulinemia (+56%, P<0.05), insulin resistance index (+43%, P<0.05), leptinemia (+113%, P<0.05), and leptin:adiponectin ratio (+98%, P<0.05) in the adult NIC group. These rats presented lower hypothalamic contents of the proteins of the leptin signaling pathway (leptin receptor (OB-R): -61%, janus tyrosine kinase 2: -41%, and p-signal transducer and activator of transcription 3: -56%, P<0.05), but higher suppressor of cytokine signaling 3 (+81%, P<0.05). Therefore, NIC exposure only during lactation programs rats for adipocyte hypertrophy in adult life, as well as for leptin and insulin resistance. Through the effects of NIC, perinatal maternal cigarette smoking may be responsible for the future development of some components of the metabolic syndrome in the offspring.

Calcium supplementation prevents obesity, hyperleptinaemia and hyperglycaemia in adult rats programmed by early weaning

It is known that Ca therapy may have anti-obesity effects. Since early weaning leads to obesity, hyperleptinaemia and insulin resistance, we studied the effect of dietary Ca supplementation in a rat model. Lactating rats were separated into two groups: early weaning (EW) - dams were wrapped with a bandage to interrupt lactation in the last 3 d of lactation and control (C) - dams whose pups had free access to milk during the entire lactation period (21 d). At 120 d, EW and C offspring were subdivided into four groups: (1) C, received standard diet; (2) CCa, received Ca supplementation (10 g of calcium carbonate/kg of rat chow); (3) EW, received standard diet; (4) EWCa, received Ca supplementation similar to CCa. The rats were killed at 180 d. The significance level was at P < 0·05. Adult EW offspring displayed hyperphagia (28 %), higher body weight (9 %) and adiposity (77 %), hyperleptinaemia (twofold increase), hypertriacylglycerolaemia (64 %), hyperglycaemia (16 %), higher insulin resistance index (38 %) and higher serum 25-hydroxyvitamin D₃ (fourfold increase), but lower adiponectinaemia:adipose tissue ratio (44 %). In addition, they showed Janus tyrosine kinase 2 and phosphorylated signal transducer and activator of transcription 3 underexpression in hypothalamus (36 and 34 %, respectively), suggesting leptin resistance. Supplementation of Ca for 2 months normalised these disorders. The EW group had no change in serum insulin, thyroxine or triiodothyronine, and Ca treatment did not alter these hormones. In conclusion, we reinforced that early weaning leads to late development of some components of the metabolic syndrome and leptin resistance. Dietary Ca supplementation seems to protect against the development of endocrine and metabolic disorders in EW offspring, maybe through vitamin D inhibition.

Calcium supplementation reverts central adiposity, leptin, and insulin resistance in adult offspring programed by neonatal nicotine exposure

Obesity is a worldwide epidemic. Calcium influences energy metabolism regulation, causing body weight loss. Because maternal nicotine exposure during lactation programs for obesity, hyperleptinemia, insulin resistance (IR), and hypothyroidism, we decided to evaluate the possible effect of dietary calcium supplementation on these endocrine dysfunctions in this experimental model. Osmotic minipumps containing nicotine solution (N: 6 mg/kg per day for 14 days) or saline (C) were s.c. implanted in lactating rats 2 days after giving birth (P2). At P120, N and C offspring were subdivided into four groups: 1) C - standard diet; 2) C with calcium supplementation (CCa, 10 g calcium carbonate/kg rat chow); 3) N - standard diet; and 4) N with calcium supplementation (NCa). Rats were killed at P180. As expected, N offspring showed higher visceral and total body fat, hyperleptinemia, lower hypothalamus leptin receptor (OB-R) content, hyperinsulinemia, and higher IR index. Also, higher tyrosine hydroxylase (TH) expression (+51%), catecholamine content (+37%), and serum 25-hydroxyvitamin D(3) (+76%) were observed in N offspring. Dietary calcium supplementation reversed adiposity, hyperleptinemia, OB-R underexpression, IR, TH overexpression, and vitamin D. However, this supplementation did not reverse hypothyroidism. In NCa offspring, Sirt1 mRNA was lower in visceral fat (-37%) and higher in liver (+42%). In conclusion, dietary calcium supplementation seems to revert most of the metabolic syndrome parameters observed in adult offspring programed by maternal nicotine exposure during lactation. It is conceivable that the reduction in fat mass per se, induced by calcium therapy, is the main mechanism that leads to the increment of insulin action.

Resveratrol reduces lipid peroxidation and increases sirtuin 1 expression in adult animals programed by neonatal protein restriction

Resveratrol (Res) has been associated with protective effects against oxidative stress. This study evaluated the effect of Res over lipid peroxidation, antioxidant defense, hepatic sirtuin 1 (SIRT1), which up-regulates antioxidant enzymes, and copper/zinc superoxide dismutase (Cu/Zn SOD) in adult offspring whose mothers were protein restricted during lactation. Lactating Wistar rats were divided into control (C) group, which were fed a normal diet (23% protein), and low-protein and high-carbohydrate (LPHC) group, which were fed a diet containing 8% protein. After weaning (21 days), C and LPHC offspring were fed a normal diet until they were 180 days old. At the 160th day, animals were separated into four groups as follows: control, control+Res, LPHC, and LPHC+Res. Resveratrol was given for 20 days (30  mg/kg per day by gavage). LPHC animals showed a higher total antioxidant capacity (TAC) without change in lipid peroxidation and SIRT1 expression. The treatment with Res increased TAC only in the control group without effect on lipid peroxidation and SIRT1. LPHC animals treated with Res had lower lipid peroxidation and higher protein and mRNA expression of SIRT1 without any further increase in TAC. No significant difference in liver Cu/Zn SOD expression was observed among the groups. In conclusion, maternal protein restriction during lactation programs the offspring for a higher antioxidant capacity, and these animals seem to respond to Res treatment with a lower lipid peroxidation and higher hepatic SIRT1 expression that we did not observe in the Res-treated controls. It is probable that the protective effect can be attributed to Res activating SIRT1, only in the LPHC-programmed group.

Effects of Ilex paraguariensis (yerba mate) treatment on leptin resistance and inflammatory parameters in obese rats primed by early weaning.

AIMS We evaluated the effects of yerba mate treatment over 30 days on body weight, food intake, hypothalamic leptin action and inflammatory profile in adult rats that were weaned early. MAIN METHODS To induce early weaning, the teats of lactating rats were blocked with a bandage to interrupt milk access for the last 3 days of lactation (EW group). Control offspring had free access to milk throughout lactation. On postnatal day (PN) 150, EW offspring were subdivided into: EW and M groups were treated with water and mate aqueous solution (1g/kg BW/day, gavage), respectively, for 30 days. Control offspring received water by gavage. On PN180, offspring were killed. KEY FINDINGS EW group presented hyperphagia; higher adiposity; higher NPY and TNF-α expression in the ARC nucleus; higher TNF-α and IL-1β levels in the adipose tissue; and lower IL-10 levels in the adipose tissue. These characteristics were normal in M group. As expected, the leptin injection in control offspring caused lower food intake. However, EW group exhibited no change in food intake after the leptin injection, indicating leptin resistance. In contrast, M group had a normal response to the leptin injection. SIGNIFICANCE Thirty days of mate treatment prevented the development of hyperphagia, overweight, visceral obesity and central leptin resistance. This beneficial effect on the satiety of M offspring most likely occurred after the improvement of inflammatory markers in the hypothalamus and adipocytes, which suggests that Ilex paraguariensis plays an important role in the management of obesity by acting on the inflammatory profile.

Anxiety-like, novelty-seeking and memory/learning behavioral traits in male Wistar rats submitted to early weaning

The most frequently used animal models of early weaning (EW) in rodents, maternal deprivation and pharmacological inhibition of lactation, present confounding factors, such as high stress or drug side effects, that can mask or interact with the effects of milk deprivation per se. Given these limitations, the development of new models of EW may provide useful information regarding the impact of a shortened period of breastfeeding on the endocrine and nervous systems, both during development and at adulthood. Using a model of EW in which lactating Wistar rat dams are wrapped with a bandage to block access to milk during the last three days of lactation, we have recently shown that the adult offspring presented higher body mass, hyperphagia, hyperleptinemia, leptin as well as insulin resistance, and higher adrenal catecholamine content at adulthood. Here, we used this EW model, which involves no pharmacological treatment or maternal separation, to analyze anxiety-like, novelty-seeking and memory/learning behavioral traits in the adult male offspring. To that end, animals were tested in the elevated plus maze, in the hole board arena and in the radial arm water maze. Except for an increased number of rearing events (a measure of vertical activity), no other behavioral differences were observed between EW and control animals. The contrasting behavioral results between the three EW models may be associated with differences in HPA axis function in the offspring at weaning, since it has been observed that bandaging does not affect corticosteronemia while maternal separation and pharmacological EW increase it.

Developmental plasticity of endocrine disorders in obesity model primed by early weaning in dams.

Early weaning is associated with changes in the developmental plasticity. Here, we studied the adipocytes morphology, adipokines expression or content in adipose tissue as well as adrenal and thyroid function of neonate and adult offspring primed by early weaning. After birth, lactating rats were divided into 2 groups: EW (early weaning)--dams were wrapped with a bandage to block access to milk during the last 3 days of lactation, and Control--dams whose pups had free access to milk throughout lactation (21 days). At postnatal day (PN) 21, EW pups had lower visceral and subcutaneous adipocyte area (-67.7% and -62%, respectively), body fat mass (-26%), and leptin expression in visceral adipocyte (-64%) but higher leptin expression in subcutaneous adipocyte (2.9-fold increase). Adrenal evaluations were normal, but neonate EW pups presented lower serum T3 (-55%) and TSH (-44%). At PN 180, EW offspring showed higher food intake, higher body fat mass (+21.6%), visceral and subcutaneous adipocyte area (both 3-fold increase), higher leptin (+95%) and ADRβ3 (2-fold increase) content in visceral adipose tissue, and higher adiponectin expression in subcutaneous adipose tissue (+47%) but lower in visceral adipose tissue (-40%). Adult EW offspring presented higher adrenal catecholamine content (+31%), but no changes in serum corticosterone or thyroid status. Thus, early weaning primed for hypothyroidism at weaning, which can be associated with the adipocyte hypertrophy at adulthood. The marked changes in catecholamine adrenal content and visceral adipocyte ADRB3 are generally found in obesity, contributing to the development of other cardiovascular and metabolic disturbances.

Bone metabolism in obese rats programmed by early weaning

OBJECTIVE Obesity and osteoporosis seem to have a common pathogenesis, especially because bone and adipose tissue have common origins. Since early weaning (EW) decreases adipogenesis and osteogenesis in neonate, further programming for obesity and hyperleptinemia, we hypothesized that these changes in adipogenesis could affect bone metabolism. MATERIALS/METHODS Lactating rats were separated into 3 groups: control - dams whose pups ate milk throughout lactation; mechanical EW (MEW) - dams were involved with a bandage interrupting suckling in the last 3days of lactation; pharmacological EW (PEW) - dams were bromocriptine-treated (0.5mg/twice a day via intraperitoneal injection) 3days before weaning. The adult offspring was subjected to dual-energy X-ray absorptiometry and bone tissue was also evaluated by computed tomography, microcomputed tomography and biomechanical tests, beyond serum analyses. RESULTS MEW and PEW presented higher total bone mineral density (BMD), total bone mineral content, spine BMD and bone area in postnatal day 150 (PN150). In PN180, both groups also presented increase of these parameters and higher femur BMD and fourth lumbar vertebra (LV4) BMD, femoral head radiodensity and LV4 vertebral body radiodensity, trabecular number, stiffness and break load; lower trabecular separation, maximal deformation and break deformation, and also hyperleptinemia and higher visceral fat mass and 25-hydroxivitamin D, whereas parathyroid hormone was unchanged. Serum C-terminal cross-linked telopeptide of type I collagen was lower for both groups. CONCLUSIONS Since both models program for obesity and increased bone mass, and leptin increases plasma vitamin D levels, probably leptin is the link between obesity and higher bone mass.