Next Generation Sequencing Transcriptome Quantitative Analysis of Livers of Adult Male Offspring of Obese Female Mice
ABSTRACT: Obesity during pregnancy is associated with increased risk of non-alcoholic fatty liver disease in the offspring. We used RNA next generation sequencing analysis (HiSeq2000) to have a snapshot of the liver transcriptome in twelve week-old male offspring exposed to maternal obesity and weaned onto an obesogenic diet. Overall design: Thirty-five female C57BL/6J mice were fed either a standard control diet, RM1 [7% simple carbohydrates/3% lipids (w/w)] or an energy-rich obesogenic diet [10% simple carbohydrates/20% lipids (w/w)] plus sweetened condensed milk [55% simple carbohydrates, 8% lipids, 8% protein (w/w), supplemented with mineral and vitamin mix AIN93G]. Female mice were mated, and maintained on their diets until weaning. On post-natal day 21, male offspring were weaned onto either standard chow (RM1) or the obesogenic diet, generating 4 experimental groups: maternal and offspring control diet (CC), maternal control diet and offspring obesogenic diet (CO), maternal obesogenic diet and offspring control diet (OC), maternal and offspring obesogenic diet (OO). At 12 weeks of age, mice were killed by raising on a fasted state. Liver were dissected, and snap frozen. Three high quality RNA samples from each group were randomly selected and subjected to a cDNA library preparation for further mRNA sequencing using TruSeq RNA Library Preparation kit
Project description:Obesity during pregnancy is associated with increased risk of non-alcoholic fatty liver disease in the offspring. We used RNA next generation sequencing analysis (HiSeq2000) to have a glance at the miRNome in twelve week-old male offspring exposed to maternal obesity and weaned onto an obesogenic diet. Overall design: Thirty-five female C57BL/6J mice were fed either a standard control diet, RM1 [7% simple carbohydrates/3% lipids (w/w)] or an energy-rich obesogenic diet [10% simple carbohydrates/20% lipids (w/w)] plus sweetened condensed milk [55% simple carbohydrates, 8% lipids, 8% protein (w/w), supplemented with mineral and vitamin mix AIN93G]. Female mice were mated, and maintained on their diets until weaning. On post-natal day 21, male offspring were weaned onto either standard chow (RM1) or the obesogenic diet, generating 4 experimental groups: maternal and offspring control diet (CC), maternal control diet and offspring obesogenic diet (CO), maternal obesogenic diet and offspring control diet (OC), maternal and offspring obesogenic diet (OO). At 12 weeks of age, mice were killed by raising on a fasted state. Liver were dissected, and snap frozen. Three high quality RNA samples from each group were randomly selected and subjected to a cDNA library preparation for further sRNA sequencing using TruSeq RNA Library Preparation kit
Project description:Obesity during pregnancy is associated with fetal growth restriction in the offspring. We used RNA next generation sequencing analysis (HiSeq2000) to have a snapshot of the placenta transcriptome at embryonic day 19 to examine whether mice exposed to maternal obesity had significant changes in their transcriptome which may lead to growth restriction in the fetus. Overall design: C57BL/6J mice were fed either a standard control diet, RM1 [7% simple carbohydrates/3% lipids (w/w)] or an energy-rich obesogenic diet [10% simple carbohydrates/20% lipids (w/w)] plus sweetened condensed milk [55% simple carbohydrates, 8% lipids, 8% protein (w/w), Nestle (Nestle, UK), supplemented with mineral and vitamin mix AIN93G]. Female mice were mated, and Pregnant mice were housed individually and maintained on their respective diets until sacrafice. A subset of the mice on the Obesegenic diet were exercised until E17 of pregnancy. Placenta tissue was collected at embryonic day 19.
Project description:Diet of mothers during gestation may impact offspring phenotype. This study evaluated the consequences of a maternal High-Protein (HP) diet during gestation on food preferences and phenotypic characteristics in adult rat offspring. Dams were fed a HP or a Normal-Protein (NP) isocaloric diet during gestation only. Weaned female pups were divided into 3 diet groups: NP control or one of two dietary self-selection (DSS) conditions. In DSS1, offspring had a free choice between proteins (100%) or a mix of carbohydrates (88%) and lipids (12%). In DSS2, the choice was between proteins (100%), carbohydrate (100%) or lipids (100%). DSS2 groups consumed more of their energy from protein and lipids, with a decreased carbohydrate intake (p < 0.0001) compared to NP groups, regardless of the maternal diet. Offspring from HP gestation dams fed the DSS2 diet (HPDSS2) had a 41.2% increase of total adiposity compared to NPDSS2 (p < 0.03). Liver Insulin receptor and Insulin substrate receptor 1 expression was decreased in offspring from HP compared to NP gestation dams. These results showed the specific effects of DSS and maternal diet and data suggested that adult, female offspring exposed to a maternal HP diet during foetal life were more prone to adiposity development, in response to postweaning food conditions.
Project description:The developmental origins of health and disease hypothesis proposes that an adverse early life environment, including in utero exposure to a maternal obesogenic environment, can lead to an increased long-term risk of obesity and related metabolic complications in offspring. We assessed whether maternal supplementation with conjugated linoleic acid (CLA) could prevent some of these adverse effects in offspring exposed to a maternal high fat diet. Sprague-Dawley dams consumed either a: control (CD), control with CLA (CLA), high fat (HF) or high fat with CLA (HFCLA) diet 10 days prior to mating and throughout pregnancy/lactation. Male offspring were weaned onto a standard chow diet. Body composition was quantified by DXA and oral glucose tolerance tests conducted on adult offspring. Gene/protein expression and histological analysis were conducted in adipose tissue. Offspring from HF dams had increased body weight, body fat deposition, impaired insulin sensitivity and adipocyte hypertrophy; all of which were rescued in HFCLA offspring. Molecular and histological analyses of the adipose tissue suggest that disturbances in adipogenesis may mediate the metabolic dysfunction observed in HF offspring. Therefore, CLA supplementation to a maternal obesogenic diet may be a promising strategy to prevent adverse programming outcomes.
Project description:Mice were weaned onto standard RM1 diet or onto a highly palatable obesogenic diet (824018 – ‘45% AFE fat) supplemented with condensed milk. After 12 weeks, mice were killed, and liver NK cells (Lin- NK1.1+ CD49a- CD49b+) or ILC1 (Lin- NK1.1+ CD49a+ CD49b-) were sorted. Total RNA was extracted from sorted cells, cDNA generated and RNASeq performed. Overall design: Cells from the livers or 3 lean and 3 obese mice were sorted, with liver NK cells and ILC1 isolated from each mouse, for a total of 12 samples
Project description:Background: It is well established that maternal exercise during pregnancy improves metabolic outcomes associated with obesity in mothers and offspring, however, its effects on the gut microbiota of both mother and offspring, are unknown. Here, we investigated whether wheel running exercise prior to and during pregnancy and prolonged feeding of an obesogenic diet were associated with changes in the gut microbiomes of Sprague-Dawley rat dams and their offspring. Female rats were fed either chow or obesogenic diet, and half of each diet group were given access to a running wheel 10 days before mating until delivery, while others remained sedentary. 16S rRNA gene amplicon sequencing was used to assess gut microbial communities in dams and their male and female offspring around the time of weaning. Results: Statistical analyses at the operational taxonomic unit (OTU) level revealed that maternal obesogenic diet decreased gut microbial alpha diversity and altered abundances of bacterial taxa previously associated with obesity such as Bacteroides and Blautia in dams, and their offspring of both sexes. Distance based linear modeling revealed that the relative abundances of Bacteroides OTUs were associated with adiposity measures in both dams and offspring. We identified no marked effects of maternal exercise on the gut microbiota of obesogenic diet dams or their offspring. In contrast, maternal exercise decreased gut microbial alpha diversity and altered the abundance of 88 microbial taxa in offspring of control dams. Thirty of these taxa were altered in a similar direction in offspring of sedentary obesogenic vs. control diet dams. In particular, the relative abundances of Oscillibacter OTUs were decreased in offspring of both exercised control dams and sedentary obesogenic diet dams, and associated with blood glucose concentrations and adiposity measures. Analyses of predicted bacterial metabolic pathways inferred decreased indole alkaloid biosynthesis in offspring of both obesogenic diet and exercised control dams. Conclusions: Our data suggest that maternal exercise prior to and during pregnancy resulted in gut dysbiosis in offspring of control dams. Importantly, alterations in the maternal gut microbiota by obesogenic diet or obesity were transferred to their offspring.
Project description:Emerging evidence suggests that maternal obesity (MO) predisposes offspring to obesity and the recently described non-alcoholic fatty pancreas disease (NAFPD) but involved mechanisms remain unclear. Using a pathophysiologically relevant murine model, we here investigated a role for the biological clock--molecular core circadian genes (CCG) in the generation of NAFPD.Female C57BL6 mice were fed an obesogenic diet (OD) or standard chow (SC) for 6 weeks, prior to pregnancy and throughout gestation and lactation: resulting offspring were subsequently weaned onto either OD (Ob_Ob and Con_Ob) or standard chow (Ob_Con and Con_Con) for 6 months. Biochemical, pro-inflammatory and pro-fibrogenic markers associated with NAFPD were then evaluated and CCG mRNA expression in the pancreas determined.Offspring of obese dams weaned on to OD (Ob_Ob) had significantly increased (p?0.05): bodyweight, pancreatic triglycerides, macrovesicular pancreatic fatty-infiltration, and pancreatic mRNA expression of TNF-?, IL-6, ?-SMA, TGF-? and increased collagen compared to offspring of control dams weaned on to control chow (Con_Con). Analyses of CCG expression demonstrated a phase shift in CLOCK (-4.818, p<0.01), REV-ERB-? (-1.4,p<0.05) and Per2 (3.27,p<0.05) in association with decreased amplitude in BMAL-1 (-0.914,p<0.05) and PER2 (1.18,p<0.005) in Ob_Ob compared to Con_Con. 2-way ANOVA revealed significant interaction between MO and post-weaning OD in expression of CLOCK (p<0.005), PER1 (p<0.005) and PER2 (p<0.05) whilst MO alone influenced the observed rhythmic variance in expression of all 5 measured CCG.Fetal and neonatal exposure to a maternal obesogenic environment interacts with a post-natal hyper-calorific environment to induce offspring NAFPD through mechanisms involving perturbations in CCG expression.
Project description:Maternal diet and gestational hyperglycaemia have implications for offspring health. Leptin (LEP) and fat mass and obesity-associated (FTO) alleles are known to influence body fat mass in humans, potentially via effects on appetite. We hypothesized that expression of Fto, Lep, and other appetite-related genes (Argp, Npy, Pomc, Cart, Lepr) in the offspring of female mice are influenced by the glycaemic index (GI) of carbohydrates in the maternal diet. C57BL/6 mice were randomly assigned to low or high GI diets and mated with chow-fed males at eight weeks of age. Male pups were weaned at four weeks and randomly divided into two groups, one group following their mother's diet (LL and HH), and one following the standard chow diet (LC and HC) to 20 weeks. Fto expression was 3.8-fold higher in the placenta of mothers fed the high GI diet (p = 0.0001) and 2.5-fold higher in the hypothalamus of 20-week old offspring fed the high GI (HH vs. LL, p < 0.0001). By contrast, leptin gene (Lep) expression in visceral adipose tissue was 4.4-fold higher in four-week old offspring of low GI mothers (LC vs. HC, p < 0.0001) and 3.3-fold higher in visceral adipose tissue of 20-week old animals (LL vs. HH, p < 0.0001). Plasma ghrelin and leptin levels, and hypothalamic appetite genes were also differentially regulated by maternal and offspring diet. These findings provide the first evidence in an animal model that maternal high GI dietary carbohydrates that are digested and absorbed faster may contribute to programming of appetite in offspring.
Project description:Previous studies have shown that maternal diet-induced obesity leads to increased risk of type 2 diabetes in offspring. The current study investigated if weaning onto an obesogenic diet exaggerated the detrimental effects of maternal diet-induced obesity in adipose tissue. Maternal obesity and offspring obesity led to reduced expression of key insulin signalling proteins, including insulin receptor substrate-1 (IRS-1). The effects of maternal obesity and offspring obesity were, generally, independent and additive. Irs1 mRNA levels were similar between all four groups of offspring, suggesting that in both cases post-transcriptional regulation was involved. Maternal diet-induced obesity increased miR-126 expression however levels of this miR were not influenced by a post-weaning obesogenic diet. In contrast, a post-weaning obesogenic diet was associated with increased levels of suppressor of cytokine signaling-1, implicating increased degradation of IRS-1 as an underlying mechanism. Our results suggest that whilst programmed reductions in IRS-1 are associated with increased levels of miR-126 and consequently reduced translation of Irs1 mRNA, the effects of a post-weaning obesogenic diet on IRS-1 are mediated by miR-126 independent mechanisms, including increased IRS-1 protein degradation. These divergent mechanisms explain why the combination of maternal obesity and offspring obesity leads to the most pronounced effects on offspring metabolism.
Project description:Maternal obesity is associated with obesity and metabolic disorders in offspring. However, intervention strategies to reverse or ameliorate the effects of maternal obesity on offspring health are limited. Following maternal undernutrition, taurine supplementation can improve outcomes in offspring, possibly via effects on glucose homeostasis and insulin secretion. The effects of taurine in mediating inflammatory processes as a protective mechanism has not been investigated. Further, the efficacy of taurine supplementation in the setting of maternal obesity is not known. Using a model of maternal obesity, we examined the effects of maternal taurine supplementation on outcomes related to inflammation and lipid metabolism in mothers and neonates. Time-mated Wistar rats were randomised to either: 1) control : control diet during pregnancy and lactation (CON); 2) CON supplemented with 1.5% taurine in drinking water (CT); 3) maternal obesogenic diet (high fat, high fructose) during pregnancy and lactation (MO); or 4) MO supplemented with taurine (MOT). Maternal and neonatal weights, plasma cytokines and hepatic gene expression were analysed. A MO diet resulted in maternal hyperinsulinemia and hyperleptinemia and increased plasma glucose, glutamate and TNF-? concentrations. Taurine normalised maternal plasma TNF-? and glutamate concentrations in MOT animals. Both MO and MOT mothers displayed evidence of fatty liver accompanied by alterations in key markers of hepatic lipid metabolism. MO neonates displayed a pro-inflammatory hepatic profile which was partially rescued in MOT offspring. Conversely, a pro-inflammatory phenotype was observed in MOT mothers suggesting a possible maternal trade-off to protect the neonate. Despite protective effects of taurine in MOT offspring, neonatal mortality was increased in CT neonates, indicating possible adverse effects of taurine in the setting of normal pregnancy. These data suggest that maternal taurine supplementation may ameliorate the adverse effects observed in offspring following a maternal obesogenic diet but these effects are dependent upon prior maternal nutritional background.