Congenital Zika virus infection impacts on male mouse offspring's reproductive biology.

Effects of developmental exposure to indole-3-carbinol or2,3,7,8-tetrachlorodibenzo-p-dioxin on reproductive potential of male rat offspring

Obstructive sleep apnoea (OSA) is characterized by intermittent hypoxia, which causes oxidative stress and inflammation and increases the risk of cardiovascular disease. OSA during pregnancy causes adverse maternal and fetal outcomes. The effects of pre-existing OSA in pregnant women on cardiometabolic outcomes in the offspring are unknown. We evaluated basic metabolic parameters, as well as aortic vascular and perivascular adipose tissue (PVAT) function in response to adiponectin, and examined DNA methylation of adiponectin gene promoter in PVAT in 16-week-old adult offspring exposed to gestational intermittent hypoxia (GIH). GIH decreased body weights at week 1 in both male and female offspring, and caused subsequent increases in body weight and food consumption in male offspring only. Adult female offspring had normal levels of lipids, glucose and insulin, with no endothelial dysfunction. Adult male offspring exhibited dyslipidaemia, insulin resistance and hyperleptinaemia. Decreased endothelial-dependent vasodilatation, loss of anti-contractile activity of PVAT and low circulating PVAT adiponectin levels, as well as increased pro-inflammatory gene expression and DNA methylation of adiponectin gene promoter, occurred in adult male offspring. Our results suggest that male offspring of women with OSA could be at risk of developing cardiometabolic disease during adulthood. Perturbations during pregnancy can program the offspring to develop cardiometabolic diseases later in life. Obstructive sleep apnoea (OSA) is a chronic condition that frequently affects pregnancies and leads to adverse fetal outcomes. We assessed the offspring of female mice experiencing gestational intermittent hypoxia (GIH), a hallmark of OSA, for changes in metabolic profiles, aortic nitric oxide (NO)-dependent relaxations, perivascular adipose tissue (PVAT) anti-contractile activities and the responses to adiponectin, and DNA methylation of the adiponectin gene promoter in PVAT tissue. Pregnant mouse dams were exposed to intermittent hypoxic cycles ( 21-12%) for 18 days. GIH resulted in lower body weights of pups at week 1, followed by significant weight gain by week 16 of age in male but not female offspring. Plasma lipids, leptin and insulin resistance were higher in GIH male adult offspring. Endothelium-dependent relaxation in response to ACh and the anti-contractile activity of PVAT in the abdominal aorta was reduced in GIH adult male offspring. Incubation of arteries from GIH adult male offspring with adiponectin restored the anti-contractile activity of PVAT. Both circulating and PVAT tissue homogenate levels of adiponectin, as well as gene expression of adiponectin in PVAT, were lower in GIH male offspring, along with an increased gene expression of inflammatory cytokines. Pyrosequencing of adiponectin gene promoter in PVAT showed increased DNA methylation in GIH male offspring. Our results indicate that GIH leads to vascular disease in adult male offspring through PVAT dysfunction, which was associated with low adiponectin levels and epigenetic modifications on the adiponectin gene promoter.

Epidemiologic studies suggest that prenatal exposure to maternal cigarette smoking is associated with an increased risk of elevated blood pressure in postnatal life. The present study was designed to test the hypothesis that fetal and neonatal nicotine exposure increased vascular contractility in adult offspring. Nicotine was administered to pregnant rats via s.c. osmotic minipumps throughout gestation and up to 10 days after delivery. Aortas were isolated from adult male and female offspring at the age of 3 months old. Nicotine significantly increased KCl- and norepinephrine-induced contractions of the aorta in male, but not female, offspring. Inhibition of endothelial nitric oxide synthase (eNOS) with N(G)-nitro-L-arginine (L-NNA) significantly increased norepinephrine-induced contractions in control male offspring but showed no effect in nicotine-treated male offspring. In the presence of L-NNA, there was no significant difference in norepinephrine-induced contractions between control and nicotine-treated males. In contrast, nicotine caused a significant increase in L-NNA-mediated potentiation of norepinephrine-induced contractions in female offspring. Nicotine had no effect on sodium nitroprusside-induced endothelium-independent relaxations of aortas from either male or female offspring. However, it decreased endothelium-dependent relaxations induced by acetylcholine in male offspring but increased them in females. There were no differences in eNOS protein levels in aortas between the control and nicotine-treated animals in either male or female offspring. The results suggest that fetal and neonatal nicotine exposure alters vascular functions in adult offspring in a gender-specific manner, which may lead to an increased risk of cardiovascular dysfunction in later life.

Bisphenol A (BPA) is an endocrine-disrupting chemical (EDC) associated with non-alcoholic fatty liver disease (NAFLD). The effects of gestational BPA exposure on hepatic lipid accumulation in offspring are not fully understood. Here, we investigate the sex-dependent effects of gestational BPA exposure on hepatic lipid and glucose metabolism in the offspring of mice to reveal the mechanisms underlying gestational BPA exposure-associated NAFLD. Pregnant mice were administered gavage with or without 1 μg kg−1 day−1 BPA at embryonic day 7.5 (E7.5)–E16.5. Hepatic glucose and lipid metabolism were evaluated in these models. Both male and female offspring mice exhibited hepatic fatty liver after BPA treatment. Lipid accumulation and dysfunction of glucose metabolism were observed in male offspring. We revealed abnormal expression of lipid regulators in the liver and that inhibition of peroxisome proliferator-activated receptor γ (PPARγ) repressed hepatic lipid accumulation induced by gestational BPA exposure. We also found a sex-dependent decrease of hepatocyte nuclear factor 1b (HNF1b) expression in male offspring. The transcriptional repression of PPARγ by HNF1b was confirmed in L02 cells. Downregulation of HNF1b, upregulation of PPARγ, and subsequent upregulation of hepatic lipid accumulation were essential for NAFLD development in male offspring gestationally exposed to BPA as well as BPA-exposed adult male mice. Dysregulation of the HNF1b/PPARγ pathway may be involved in gestational BPA exposure-induced NAFLD in male offspring. These data provide new insights into the mechanism of gestational BPA exposure-associated sex-dependent glucose and lipid metabolic dysfunction.Graphical abstractSchematic of the mechanism of gestational BPA exposure-induced glucose and lipid metabolic dysfunction.

In vitro fertilization (IVF) is a noncoital method of conception used to treat human infertility. Although IVF is viewed as largely safe, it is associated with adverse outcomes in the fetus, placenta, and adult offspring. Because studies focusing on the effect of IVF on the male reproductive system are limited, we used a mouse model to assess the morphological and molecular effects of IVF on male offspring. We evaluated 3 developmental stages: 18.5-day fetuses and 12- and 39-week-old adults. Regardless of age, we observed changes in testicular-to-body weight ratios, serum testosterone levels, testicular morphology, gene expression, and DNA methylation. Also, sperm showed changes in morphology and DNA methylation. To assess multigenerational phenotypes, we mated IVF-conceived and naturally conceived males with wild-type females. Offspring from IVF males exhibited decreased fetal-to-placental weight ratios and changes in placenta gene expression and morphology regardless of sex. At 12 weeks of age, offspring showed higher body weights and differences in glucose, triglyceride, insulin, total cholesterol, HDL-C, and LDL/VLDL-C levels. Both sexes showed changes in gene expression in liver, testes, and ovaries and decreased global DNA methylation. Collectively, our findings demonstrate that male IVF offspring exhibit abnormal testicular and sperm morphology and molecular alterations with a multigenerational impact.

Effects of developmental exposure to indole-3-carbinol or 2,3,7,8-tetrachlorodibenzo-p-dioxin on reproductive potential of male rat offspring.

Maternal immune activation in late gestation enhances locomotor response to acute but not chronic amphetamine treatment in male mice offspring: Role of the D1 receptor

Research suggests that maternal exercise in pregnancy may have beneficial effects on the brain function of offspring. This study sought to determine if voluntary wheel running during pregnancy improves depression-like behavior, temporal order memory, and hippocampal neurogenesis in both female and male offspring mice. Pregnant mice were allowed to run voluntarily by introducing running wheels into the housing cages throughout the gestational period. Male and female mice offspring at the age of 8- to 9-week-old were then tested on the temporal order task and forced swim test, then euthanized for immunostaining for examining adult hippocampal cell proliferation and neuronal differentiation. Results showed that both male and female pups had reduced depression-like behavior, while only male offspring demonstrated improvement in temporal order memory. Immunostaining revealed that male offspring showed an increase in the number of immature neurons in the ventral hippocampus, whereas female offspring showed enhanced cell proliferation in the dorsal hippocampus. These findings indicate that maternal voluntary wheel running benefits both female and male offspring on reducing depression-like behavior, but with gender effect on promoting hippocampal cell proliferation, neuronal differentiation, and temporal order memory.

Nutritional conditions during fetal life can influence the risk to develop the metabolic syndrome in adult life (‘metabolic programming’). We aimed to establish a mouse model of metabolic programming focusing on the effects of a maternal low protein diet during gestation on glucose and lipid metabolism in the adult offspring. Pregnant C57BL/6J mice received a control or a low protein diet throughout gestation. Offspring received a low fat diet or a high fat diet from 6–22 weeks of age. Glucose metabolism was studied with a whole‐body‐glucose test using [6,6‐2H]‐glucose. Hepatic gene expression was characterized by microarray. Maternal low‐protein‐diet did not affect glucose metabolism in male offspring. Female offspring from normal‐protein fed dams was relatively resistant to diet‐induced metabolic dysregulation. Maternal low‐protein‐diet during gestation led to deteriorated insulin sensitivity upon high‐fat feeding in female offspring. Microarray analyses showed a sex‐specific effect of the maternal diet. We conclude that, in mice, maternal protein restriction during gestation does not change the glucose response to a high fat diet in male offspring. However, it changes fatty acid and glucose metabolism in female offspring in such a way that it resembles male metabolism. Supported by the Netherlands Heart Foundation, grant 2004T048.

Late gestation during pregnancy has been associated with a relatively high prevalence of obstructive sleep apnoea (OSA). Intermittent hypoxia, a hallmark of OSA, could impose significant long-term effects on somatic growth, energy homeostasis and metabolic function in offspring. Here we show that late gestation intermittent hypoxia induces metabolic dysfunction as reflected by increased body weight and adiposity index in adult male offspring that is paralleled by epigenomic alterations and inflammation in visceral white adipose tissue. Fetal perturbations by OSA during pregnancy impose long-term detrimental effects manifesting as metabolic dysfunction in adult male offspring. Pregnancy, particularly late gestation (LG), has been associated with a relatively high prevalence of obstructive sleep apnoea (OSA). Intermittent hypoxia (IH), a hallmark of OSA, could impose significant long-term effects on somatic growth, energy homeostasis, and metabolic function in offspring. We hypothesized that IH during late pregnancy (LG-IH) may increase the propensity for metabolic dysregulation and obesity in adult offspring via epigenetic modifications. Time-pregnant female C57BL/6 mice were exposed to LG-IH or room air (LG-RA) during days 13-18 of gestation. At 24weeks, blood samples were collected from offspring mice for lipid profiles and insulin resistance, indirect calorimetry was performed and visceral white adipose tissues (VWAT) were assessed for inflammatory cells as well as for differentially methylated gene regions (DMRs) using a methylated DNA immunoprecipitation on chip (MeDIP-chip). Body weight, food intake, adiposity index, fasting insulin, triglycerides and cholesterol levels were all significantly higher in LG-IH male but not female offspring. LG-IH also altered metabolic expenditure and locomotor activities in male offspring, and increased number of pro-inflammatory macrophages emerged in VWAT along with 1520DMRs (P<0.0001), associated with 693genes. Pathway analyses showed that genes affected by LG-IH were mainly associated with molecular processes related to metabolic regulation and inflammation. LG-IH induces metabolic dysfunction as reflected by increased body weight and adiposity index in adult male offspring that is paralleled by epigenomic alterations and inflammation in VWAT. Thus, perturbations to fetal environment by OSA during pregnancy can have long-term detrimental effects on the fetus, and lead to persistent metabolic dysfunction in adulthood.

Paternal retrievals increase testosterone levels in both male and female California mouse (Peromyscus californicus) offspring

Linoleic acid (LA), an n-6 polyunsaturated fatty acid (PUFA), is essential for fetal growth and development. The consumption of LA has increased over the last decades due to availability in foods. In this study, we aimed to investigate the effect of maternal and postnatal diet high in LA (HLA) diet on plasma fatty acid (FA) composition, plasma and hepatic lipids, and genes involved in lipid metabolism in the liver of adult offspring, and any benefit of postnatal diet low in LA. Female rats were fed with low LA (LLA; 1.44% energy from LA: recommended daily intake) or HLA (6.21% energy from LA: what Australians are consuming) diets for 10 weeks before pregnancy, and during gestation/lactation. Offspring were weaned at postnatal day 25 (PN25), fed LLA or HLA diets and sacrificed at PN180 (6 months old). Postnatal HLA diet increased the brain weight as a percentage of body weight in adult male offspring. Postnatal HLA diet decreased circulating total n-3 PUFA and alpha-linolenic acid (ALA), while increased circulating total n-6 PUFA, LA and arachidonic acid (AA) in both male and female offspring. There were no differences in fat mass, systolic blood pressure, glucose tolerance and insulin sensitivity among the groups. Maternal HLA diet increased circulating leptin in female offspring, but not in male offspring. Interestingly, maternal HLA diet decreased adiponectin concentration in the plasma of male offspring, but not in females. Postnatal HLA diet significantly decreased aspartate transaminase (AST), a circulating liver enzyme, in female offspring, however, other liver enzymes such as alanine transaminase (ALT) and alkaline phosphatase (ALP) remained unchanged. Furthermore, postnatal HLA diet downregulated total cholesterol, high density lipoprotein (HDL) cholesterol and triglyceride in the plasma of male offspring, but not in females. There were no differences in hepatic cholesterol and triglycerides among the groups. Interestingly, maternal HLA diet downregulated the mRNA expression of Hmgcr (3-hydroxy-3- methyl-glutaryl-coenzyme A reductase) in the liver of both male and female offspring. Maternal HLA diet decreased the mRNA expression of Cpt1a and Acox1 in the liver of female offspring. In conclusion, maternal and postnatal HLA diet affected the plasma FA composition, circulating adiponectin, leptin and lipids, and hepatic expression of genes related to lipid metabolism in the adult offspring in a sex-specific manner. This may contribute the altered lipid metabolism in the liver and development of lipid dysfunction associated with chronic conditions like non-alcoholic fatty liver disease in the adult offspring.

There is a paucity of studies on the multigenerational reproductive toxicity of fine particle matter (PM2.5) exposure during pregnancy on male offspring and the underlying mechanisms. This study explored the effects of PM2.5 exposure during pregnancy on the spermatogenesis of three consecutive generations of male mouse offspring. We randomized pregnant C57BL/6 mice into the control group, the Quartz Fiber Membrane control group, and two experimental groups exposed to different concentrations of PM2.5 (4.8 and 43.2 mg/kg B.Wt.). Pregnant mice from experimental groups received intratracheal instillation of PM2.5 of different doses on a three-day basis until birth. F1 mature male offspring from PM2.5-exposed pregnant mice were mated with normal female C57BL/6 mice. Likewise, their F2 mature male followed the same to produce the F3 generation. The results showed that PM2.5 exposure during pregnancy led to decreased body and tail length, body weight, and survival rates, decreased sperm concentration and sperm motility, and increased sperm abnormality rates significantly in F1 male offspring. We barely observed significant impacts of PM2.5 on the birth number, survival rates, and index of testes in the F2 and F3 offspring. Further exploration showed that PM2.5 exposure during pregnancy caused the morphological abnormality of Sertoli cells, downregulated androgen receptor (AR) and connexin43, upregulated anti-Müllerian hormone (AMH), cytokeratin-18 (CK-18), caspase-3, and cleaved caspase-3, decreased thyroid-stimulating hormone (TSH) and testosterone (T), and increased triiodothyronine (T3) in F1 male mouse offspring. Overall, we hypothesize that PM2.5 exposure during pregnancy mainly negatively impacts spermatogenesis in the F1 offspring. The possible mechanism could be that PM2.5 exposure during pregnancy disrupts endocrine hormone release in the F1 generation, thereby influencing the maturation and proliferation of their Sertoli cells and hindering spermatogenesis. This study for the first time investigates the role of Sertoli cells in the reproductive toxicity of PM2.5 on offspring.

Perinatal programming of depressive-like behavior by inflammation in adult offspring mice whose mothers were fed polluted eels: Gender selective effects

P.1.h.034 Epigenetic reversal of early-life stress-induced visceral hypersensitivity and anxiety behaviour