Uteroplacental insufficiency in rats induces renal apoptosis and delays nephrogenesis completion.

AIM: Uteroplacental insufficiency in rats reduces nephron endowment, leptin concentrations and programmes cardiorenal disease in offspring. Cross-fostering growth-restricted (Restricted) offspring onto a mother with normal lactation restores leptin concentrations and nephron endowment. This study aimed to determine whether the reduced nephron endowment in Restricted offspring is due to delayed glomerular formation and dysregulation of renal genes regulating branching morphogenesis, apoptosis or leptin signalling. Furthermore, we aimed to investigate whether cross-fostering Restricted offspring onto Control mothers could improve glomerular maturation and restore renal gene abundance. METHODS: Uteroplacental insufficiency was induced by bilateral uterine vessel ligation (Restricted) or sham (Control) surgery on gestation day 18 (E18). Kidneys were collected at E20, postnatal day 1 (PN1) and PN7. An additional cohort was cross-fostered onto separate mothers at birth and kidneys collected at PN7. RESULTS: Kidneys were lighter in the Restricted group, but weight was restored with cross-fostering. At E20, abundance of Bax, Flt1 and Vegfa was increased in Restricted offspring, while Ret and Bcl2 transcripts were increased only in Restricted females. At PN7, abundance of Gdnf and Ret was higher in Restricted offspring, as was Casp3. Restricted offspring had a wider nephrogenic zone with more immature glomeruli suggesting a delayed or extended nephrogenic period. Cross-fostering had subtle effects on gene abundance and glomerular maturity. CONCLUSION: Uteroplacental insufficiency induced apoptosis in the developing kidney and delayed and extended nephrogenesis. Cross-fostering Restricted offspring onto Control mothers had beneficial effects on kidney growth and renal maturity, which may contribute to the restoration of nephron endowment.

The current state of reproductive biology research in Australia and New Zealand: core themes from the Society for Reproductive Biology Annual Meeting, 2016.

Because reproduction is essential for all life, it is central to our understanding of all aspects of biology. The Society for Reproductive Biology (SRB) 2016 conference held on the Gold Coast (Qld, Australia) displayed the current breadth of reproductive research in Australia and New Zealand, with additional insights from world leaders in the field. This conference review provides a focused summary of the key questions, emerging ideas and novel technologies that were presented in the symposia. Presented research demonstrated key advances in how stem cell biology may allow us to better understand pluripotency, as well as how environmental and lifestyle factors, such as circadian disruption, smoking, alcohol and diet, affect gametogenesis, embryo implantation, placental function and reproductive capacity. Sessions also highlighted the role of reproductive biology in providing insight into the mechanisms and processes governing a wide range of biological science disciplines, including cancer research and therapies, oncofertility, conservation of native species and chronic non-communicable diseases. Recurring themes included the importance of male and female gamete quality for reproductive potential and the critical and varied roles of the placenta in the maintenance of a healthy pregnancy. Dysregulation of reproductive processes can contribute to a variety of pathological states that affect future health, fertility and fecundity. Research being conducted by the SRB has the potential to shape not only the fertility of the current generation, but also the health and reproductive viability of future generations.

Prenatal corticosterone exposure programs sex-specific adrenal adaptations in mouse offspring.

Maternal stress can impair foetal development and program sex-specific disease outcomes in offspring through the actions of maternally produced glucocorticoids, predominantly corticosterone (Cort) in rodents. We have demonstrated in mice that male but not female offspring prenatally exposed to Cort (33 µg/kg/h for 60 h beginning at E12.5) develop cardiovascular/renal dysfunction at 12 months. At 6 months of age, renal function was normal but male offspring had increased plasma aldosterone concentrations, suggesting that altered adrenal function may precede disease. This study investigated the long-term impact of prenatal exposure to Cort on adrenal growth, morphology and steroidogenic capacity as well as plasma Cort concentrations in offspring at postnatal day 30 (PN30), 6 months and 12 months of age. Prenatal Cort exposure decreased adrenal volume, particularly of the zona fasciculata, in male offspring at PN30 but increased both relative and absolute adrenal weight at 6 months of age. By 12 months of age, male Cort-exposed offspring had reduced absolute adrenal weight in association with increased adrenal plaque deposition (lipogenic pigmentation). Plasma Cort concentrations were elevated in male 6-month offspring but not at other ages. mRNA expression of Mc2r (ACTH receptor) was increased in males at PN30, and Cyp11a1 expression was decreased at 6 and 12 months of age. There were no changes in the adrenals of female Cort-exposed offspring. This study demonstrates that prenatal Cort exposure induces offspring adrenal gland dysfunction in an age- and sex-specific manner, which may contribute to long-term programmed disease in male offspring after maternal stress.

Maternal hypomagnesemia causes placental abnormalities and fetal and postnatal mortality.

INTRODUCTION: Magnesium (Mg(2+)) is essential for cellular growth and the maintenance of normal cellular processes. However, little is known about how maternal hypomagnesemia during pregnancy affects fetal growth and development. This study investigated the effects of maternal hypomagnesemia on the late gestation placenta and fetus, and postnatal outcomes until weaning. METHODS: Female CD1 mice consumed a control (0.2% w/w Mg(2+)), moderately Mg(2+) deficient (MMD; 0.02% w/w Mg(2+)) or severely Mg(2+) deficient (SMD; 0.005% w/w Mg(2+)) diet for 4 weeks prior to mating and throughout pregnancy. Dams were killed at E18.5 for embryonic studies or allowed to litter naturally and the offspring studied up to postnatal day 21. RESULTS: At E18.5, both Mg(2+) deficient diets decreased maternal plasma and bone Mg(2+) but only the SMD diet decreased fetal plasma Mg(2+). Maternal hypomagnesemia led to fetal loss and fetal growth restriction. Maternal Mg(2+) deficiency increased placental glycogen cell area and decreased spongiotrophoblast cell area while upregulating mRNA expression of the MagT1 Mg(2+) transporter in spongiotrophoblast cells. The SMD animals also displayed instances of gross placental abnormalities. After birth, pups in the SMD group had increased early postnatal mortality and failed to thrive. Pups in the MMD group underwent catch-up growth but remained shorter than controls at PN21 and were hypomagnesemic and hypoglycemic. CONCLUSIONS: These changes suggest that maternal Mg(2+) deficiency during pregnancy impairs placental development and fetal growth, which may have long-term health consequences for offspring. Collectively, these results have important implications for women who are Mg(2+) deficient during pregnancy.

The effects of gestational age and maternal hypoxia on the placental renin angiotensin system in the mouse.

INTRODUCTION: The renin angiotensin system (RAS) is an important mediator of placental development. However, a comprehensive expression profile for 8 key components of the placental RAS throughout murine gestation has not been performed. Furthermore, maternal hypoxia induces dysregulation of RAS expression in fetal tissues but the effects on the murine placental RAS are less well known. METHODS: Placentas were collected from male and female CD1 mouse fetuses at seven gestational ages for qPCR analysis of Agt, Ren1, Atp6ap2, Ace, Ace2, Agtr1a, Agtr2 and Mas1. mRNA localisation of Agtr1 and Mas1 and protein localisation of ACE and ACE2 was determined at E18.5. To determine the effects of maternal hypoxia on the placental RAS, mice were housed in 12% oxygen from E14.5-E18.5 and placentas examined at E18.5. RESULTS: All RAS genes were expressed in the placenta throughout pregnancy and expression varied with fetal sex and age. Agtr1 was expressed within the labyrinth while Mas1 was expressed within the intraplacental yolk sac. ACE and ACE2 were localised to both labyrinth and junctional zones. In response to maternal hypoxia the expression of Agt, Ace and Ace2 was decreased but expression of Agtr1a was increased. Ace and Agtr1a mRNA levels were affected to a greater extent in females compared to males. DISCUSSION: Collectively, the location within the placenta as well as the expression profiles identified, support a role for the placental RAS in labyrinth development. The placental RAS is disturbed by maternal hypoxia in a sexually dimorphic manner and may contribute to impairment of placental vascular development.

Mid- to late term hypoxia in the mouse alters placental morphology, glucocorticoid regulatory pathways and nutrient transporters in a sex-specific manner.

Maternal hypoxia is a common perturbation that can disrupt placental and thus fetal development, contributing to neonatal impairments. Recently, evidence has suggested that physiological outcomes are dependent upon the sex of the fetus, with males more susceptible to hypoxic insults than females. This study investigated the effects of maternal hypoxia during mid- to late gestation on fetal growth and placental development and determined if responses were sex specific. CD1 mice were housed under 21% or 12% oxygen from embryonic day (E) 14.5 until tissue collection at E18.5. Fetuses and placentas were weighed before collection for gene and protein expression and morphological analysis. Hypoxia reduced fetal weight in both sexes at E18.5 by 7% but did not affect placental weight. Hypoxia reduced placental mRNA levels of the mineralocorticoid and glucocorticoid receptors and reduced the gene and protein expression of the glucocorticoid metabolizing enzyme HSD11B2. However, placentas of female fetuses responded differently to maternal hypoxia than did placentas of male fetuses. Notably, morphology was significantly altered in placentas from hypoxic female fetuses, with a reduction in placental labyrinth blood spaces. In addition mRNA expression of Glut1, Igf2 and Igf1r were reduced in placentas of female fetuses only. In summary, maternal hypoxia altered placental formation in a sex specific manner through mechanisms involving placental vascular development, growth factor and nutrient transporter expression and placental glucocorticoid signalling. This study provides insight into how sex differences in offspring disease development may be due to sex specific placental adaptations to maternal insults.

Periconceptional alcohol consumption causes fetal growth restriction and increases glycogen accumulation in the late gestation rat placenta.

INTRODUCTION: Alcohol consumption is a common social practice among women of childbearing age. With 50% of pregnancies being unplanned, many embryos are exposed to alcohol prior to pregnancy recognition and formation of the placenta. The effects of periconceptional (PC) alcohol exposure on the placenta are unknown. METHODS: Sprague-Dawley rats were exposed to alcohol (12.5% v/v ad libitum) from 4 days prior to 4 days after conception and effects on placental growth, morphology and gene/protein expression examined at embryonic day (E) 20. RESULTS: PC ethanol (EtOH)-exposed fetuses were growth restricted and their placental/body weight ratio and placental cross-sectional area were increased. This was associated with an increase in cross-sectional area of the junctional zone and glycogen cells, especially in PC EtOH-exposed placentas from female fetuses. Junctional Glut1 and Igf2 mRNA levels were increased. Labyrinth Igf1 mRNA levels were decreased in placentas from both sexes, but protein IGF1R levels were decreased in placentas from male fetuses only. Labyrinth mRNA levels of Slc38a2 were decreased and Vegfa were increased in placentas following PC EtOH-exposure but only placentas from female fetuses exhibited increased Kdr expression. Augmented expression of the protective enzyme 11ßHsd2 was found in PC EtOH-exposed labyrinth. DISCUSSION: These observations are consistent with a stress response, apparent well beyond the period of EtOH-exposure and demonstrate that PC EtOH alters placental development in a sex specific manner. CONCLUSION: Public awareness should be increased to educate women about how excessive drinking even before falling pregnant may impact on placental development and fetal health.

The effects of low-moderate dose prenatal ethanol exposure on the fetal and postnatal rat lung.

Little is known about whether exposure of the fetus to alcohol alters pulmonary development or function. This study aimed to determine whether low-moderate ethanol (EtOH) exposure throughout gestation alters structural and non-respiratory functional aspects of the fetal and postnatal lung. Sprague-Dawley rats were fed an ad libitum liquid diet ±6% v/v EtOH daily throughout pregnancy, achieving a plasma ethanol (EtOH) concentration of 0.03%. Gene and protein expression was determined in pulmonary tissue collected from fetuses at embryonic day (E) 20 and adult offspring. The percentage of airspace and alveolar size was measured in pulmonary tissue collected at postnatal day (PN) 1. At E20, EtOH-exposed fetuses had decreased aquaporin 5 mRNA levels and a non-significant trend for decreased epithelial sodium channel type α; expression of other pulmonary fluid homeostatic and development genes and surfactant protein genes were not different between groups. At PN1, there was no difference between EtOH-exposed and control offspring in the distal airspace percentage or diameter. At 8 months, collagen type III α1 gene expression was upregulated in EtOH-exposed male offspring; this was associated with increased collagen deposition at 10 months. At 19 months, male EtOH-exposed offspring had a 25% reduction in the protein levels of surfactant protein B. The alterations observed in male EtOH-exposed offspring suggest chronic low-moderate prenatal EtOH-exposure during development may result in increased pulmonary fibrosis. Such an alteration would decrease the respiratory capacity of the lung.

Maternal corticosterone exposure in the mouse has sex-specific effects on placental growth and mRNA expression.

Maternal exposure to increased synthetic glucocorticoids (GC) during pregnancy is known to disturb fetal development and increase the risk of long-term disease. Maternal exposure to elevated levels of natural GC is likely to be common yet is relatively understudied. The placenta plays an important role in regulating fetal exposure to maternal GC but is itself vulnerable to maternal insults. This study uses a mouse model of maternal corticosterone (Cort) exposure to investigate its effects on the developing placenta. Mice were treated with Cort (33 µg/kg·h) for 60 h starting at embryonic d 12.5 (E12.5) before collection of placentas at E14.5 and E17.5. Although Cort exposure did not affect fetal size, placentas of male fetuses were larger at E17.5 in association with changes in placental Igf2. This increase in size was associated with an increase in placental thickness and an increase in placental junctional zone volume. Placentas from female fetuses were of normal size and had no changes in growth factor mRNA levels. The expression of the protective enzyme 11ß-hydroxysteroid dehydrogenase type 2 was increased at E14.5 but was decreased in males at E17.5. In contrast, the expression of Nr3c1 (which encodes the GC receptor) was increased during the Cort exposure and remained elevated at E17.5 in the placentas of male fetuses. Our study has shown that maternal Cort exposure infers a sex-specific alteration to normal placental growth and growth factor expression, thus further adding to our understanding of the mechanisms of male dominance of programmed disease.

Sex specific changes in placental growth and MAPK following short term maternal dexamethasone exposure in the mouse.

OBJECTIVES: Maternal glucocorticoid (GC) exposure during pregnancy can alter fetal development and program the onset of disease in adult offspring. The placenta helps protect the fetus from excess GC exposure but is itself susceptible to maternal insults and may be involved in sex dependant regulation of fetal programming. This study aimed to investigate the effects of maternal GC exposure on the developing placenta. STUDY DESIGN AND MAIN OUTCOME MEASURES: Pregnant mice were treated with dexamethasone (DEX-1 µg/kg/h) or saline (SAL) for 60 h via minipump beginning at E12.5. Placentas were collected at E14.5 and E17.5 and the expression of growth factors and placental transporters examined by real-time PCR and/or Western blot. Histological analysis was performed to assess for morphological changes. RESULTS: At E14.5, DEX exposed male and female fetuses had a lower weight compared to SAL animals but placental weight was lower in females only. Hsd11b2 and Vegfa gene expression was increased and MAPK1 protein expression decreased in the placentas of females only. At E17.5 placental and fetal body weights were similar and differences in MAPK were no longer present although HSD11B2 protein was elevated in placentas of DEX females. Levels of glucose or amino acid transporters were unaffected. CONCLUSIONS: Results suggest sex specific responses to maternal GCs within the placenta. Decreased levels of MAPK protein in placentas of female fetuses suggest alterations in the MAPK pathway may contribute to the lower placental weights in this sex. This may contribute towards sex specific fetal programming of adult disease.

Review: Sex specific programming: a critical role for the renal renin-angiotensin system.

The "Developmental Origins of Health and Disease" hypothesis has caused resurgence of interest in understanding the factors regulating fetal development. A multitude of prenatal perturbations may contribute to the onset of diseases in adulthood including cardiovascular and renal diseases. Using animal models such as maternal glucocorticoid exposure, maternal calorie or protein restriction and uteroplacental insufficiency, studies have identified alterations in kidney development as being a common feature. The formation of a low nephron endowment may result in impaired renal function and in turn may contribute to disease. An interesting feature in many animal models of developmental programming is the disparity between males and females in the timing of onset and severity of disease outcomes. The same prenatal insult does not always affect males and females in the same way or to the same degree. Recently, our studies have focused on changes induced in the kidney of both the fetus and the offspring, following a perturbation during pregnancy. We have shown that changes in the renin-angiotensin system (RAS) occur in the kidney. The changes are often sex specific which may in part explain the observed sex differences in disease outcomes and severity. This review explores the evidence suggesting a critical role for the RAS in sex specific developmental programming of disease with particular reference to the immediate and long term changes in the local RAS within the kidney.