Female Intrauterine Growth Restriction Research Articles

Abstract P210: Sex Specific Changes In Cerebral Proinflammatory Cytokines In Intrauterine Growth Restricted Offspring

Background: Intrauterine growth restriction (IUGR) often times results from placental ischemia, which restricts the distribution of nutrients and oxygen to meet the metabolic demand of the growing fetus. IUGR offspring (OS) have an increased risk for developing cerebrovascular diseases (CVD) later in life. Factors for development of CVDs are HTN and inflammation. Our lab has shown 17 week IUGR male rats from preeclamptic pregnancies have HTN and elevated inflammation via systemic IL-17, while our females show no change. The role of inflammation to cause HTN in IUGR offspring in a sex specific manner is unknown and the major focus of this study which examines the changes in cerebral inflammation IUGR offspring at 12 weeks. We hypothesize that cerebral inflammation in IUGR males will be elevated, while no change in female IUGR offspring will be observed. Methods: IUGR offspring originate from preeclamptic Sprague Dawley dams. OS weaned for 3 weeks and were separated by sex and IUGR status. At 12 weeks, the rats underwent carotid catheterization to measure BP. Brains were collected to measure pro-inflammatory cytokines, such as TNF-α and IL-17 via ELISAs. Results: At 12 weeks, all OS were normotensive. Brain TNF-α increased in female IUGR (17,366± 1,500 vs 8,116± 1,341 pg/mL, p<0.05) and was elevated in male IUGR (20,845±7571 vs 8,516±1712 pg/mL, ns). Brain IL-17 increased in male IUGR (13,580± 945 vs 9,620± 945 pg/mL, p<0.05) and was elevated in female IUGR (13,143± 1613 vs 10,930± 696 pg/mL, ns). Systemic IL-17 was increased in IUGR male OS (112±16.8vs100±3.9 IU/Protein/CON %, ns). Conclusion: At 12 weeks, brain TNF-α and IL-17 were elevated in IUGR males and females. The differences in expression of each cytokine between the sexes suggest that IL-17 contributes to the increase in BP, especially in males. Understanding the timing of these developments could help to elucidate the role of inflammation in the development of CVD with or without HTN. This study is clinically relevant as it provides a sex specific inflammatory target for therapeutics, to help prevent hypertension and CVD in IUGR offspring. In the future, we will evaluate the role of inflammation in IUGR offspring in a sex specific manner.

Abstract P202: Changes In Cerebral Mitochondrial Function In Intrauterine Growth-Restricted Offspring Before The Development Of Hypertension And Cerebrovascular Dysfunction

Background: Preeclampsia (PE), new-onset gestational hypertension (HTN) affects 3-8% of all births in the USA. PE is the leading cause for intrauterine growth restriction (IUGR). IUGR children have increased risks of HTN and cerebrovascular disease (CVD) later in life. Prior studies in our lab show adult male IUGR rat offspring (OS) to develop HTN, CVD, and cerebral mitochondrial dysfunction (mtDys) at 17 weeks while female IUGR OS exhibit CVD. The role of mtDys in HTN is unknown and is the focus of our study. We hypothesize that IUGR OS will have mtDys prior to HTN. Methods: Our surgical PE animal model utilized the reduced-uterine-perfusion pressure (RUPP) surgery at GD 14. OS from NP served as controls (CON) while OS from RUPP mothers were IUGR. At 12 weeks, OS BP was measured alongside cerebral mtDys through mitochondrial fusion and fission, electron-transport chain (ETC) proteins, and respiration. Results: BP did not change between sexes or among IUGR and CON offspring. Male IUGR OS showed elevated mtRes (State 2: 199 ± 1.08 v. 76 ± 1.13 pmol/mg, p < .05; State 3: 555 ± 3.02 v. 245 ± 1.77 pmol/mg, p < .05; State 4: 346 ± 1.66 v. 85 ± 1.12 pmol/mg, p < .05). Female IUGR and CON OS had no changes in BP or mtRes. Male IUGR OS had increased mito-fusion proteins MFN-1 (176 ± 1.68 v. 100 ± .57 IU/protein/CON %, p < .05), MFN-2 (155 ± 1.36 v. 100 ± .74 IU/protein/CON %, p < .05) and OPA-1 (166 ± 1.99 v. 100 ± .73 IU/protein/CON %, p < .05). DRP-1 had no changes in males; females had no changes in fusion or fission. Male IUGR OS had changes in ETC proteins (CI: 8 ± 9.34 v. 100 ± .82 IU/protein/CON %, p < .05; CII: 23 ± .53 v. 100 ± .52 IU/protein/CON %, p < .05; CIII: 26 ± .23 v. 100 ± .41 IU/protein/CON %, p < .05), as did female IUGR OS (CI: 12 ± .24 v. 100 ± .86 IU/protein/CON %, p < .05; CII: 29 ± .39 v. 100 ± .61 IU/protein/CON %, p < .05; CIII: 39 ± .56 v. 100 ± .62 IU/protein/CON %, p < .05; CIV: 76 ± .74 v. 100 ± .73 IU/protein/CON %, p < .05) vs. CON OS. Summary: Male IUGR OS have elevated mtRes, changed ETC proteins, and fusion proteins that may lead to mtDys and HTN development. This study is clinically relevant because specific cerebral therapy for mtDys for IUGR OS may prevent HTN and CVD development. Future studies will examine changes in mtDys causing male HTN and protection from HTN in females.

Abstract P420: Changes In Cardiac Oxidative Stress Is Protected In Intrauterine Growth Restricted Females Vs Males Before The Development Of Hypertension And Cardiovascular Diseases

Preeclampsia (PE) is a pregnancy complication characterized by hypertension (HTN) typically during the third trimester. PE is the leading cause for intrauterine growth restricted (IUGR) offspring (OFS). Women with PE and their OFS (IUGR) are at a higher risk for developing cardiovascular diseases (CVD) later in life. Two major factors that contribute to the development of CVDs are HTN and oxidative stress (OS). Studies show that human and animals adult IUGR OFS have a higher risk of developing HTN and CVDs compared to females later in life. The rationale for these sex differences is unknown; and we hypothesize that male IUGR rats will have elevated cardiac oxidative stress, while female IUGR will have lower cardiac OS .To test, we will examine sex differences between 12 week IUGR and control (CON) OFS that are not hypertensive, to determine if there are any change in cardiac OS before the development of HTN and CVD. Methods: Sprague Dawley rats were divided into 4 groups: IUGR male, IUGR female, and control (CON) male and female. IUGR rats were derived from the OFS reduced uterine perfusion pressure (RUPP) PE rat model. CON OFS were from normal pregnant rats. At 12wk, BP was measured via carotid catheterization. Heart tissue was collected to assess oxidative stress, via examining reactive oxygen species (ROS) Heat shock protein 1 (HSP-1) and hydrogen peroxide (H2O2). Antioxidants were measured via manganese superoxide dismutase (MnSOD), copper zinc superoxide dismutase (CuZnSOD) and antioxidant capacity (AC). There were no significant changes in HSP-1, CuZnSOD, and H2O2 between male and female OFS. However, female IUGR OFS had a 20% increase in antioxidant MnSOD (119.9 ± 6.133 vs 93.98 ± 5.197 %MnSOD U/protein/CON, p <0.05) compared to CON in the heart. Male OFS had no significant changes for MnSOD. IUGR female AC had a 2-fold increase in activity (22.13 ± 1.019 vs 14.15 ± 1.852 mM Trolox/mg protein, p <0.05) compared to its CON. Furthermore, IUGR male OFS had a slight decrease in AC compared to CON male (11.00 ± 1.420 vs 13.24 ± 0.4676 mM Trolox/mg protein, ns). The significant increase of AC and MnSOD in IUGR female OFS suggests that females may be more protected from OS, via mitochondria-specific antioxidants compared to males.

Abstract 028: Sex Differences In Hypertension And Cerebral Oxidative Stress Of Preeclamptic Rat Offspring

Intrauterine growth restriction (IUGR) occurs when the fetus doesn’t receive adequate nutrients. The leading cause of IUGR is placental ischemia, often seen in preeclampsia (PE). Adult IUGR offspring have an increased risk of HTN and oxidative stress (OS), with sex differences. Studies have shown that IUGR males have HTN, cerebrovascular dysfunction (CVD), and cerebral OS at 17 wks; IUGR females are NT with limited CVD and no significant OS. Our study aims to characterize the sex differences in timing and changes of cerebral OS in IUGR offspring. HTN and OS are both present in adult IUGR males, however, the order and timing is unknown. We hypothesize that OS leads to HTN. To test this, we examined the changes in brain OS in 12 wk IUGR and control (CON) males (before HTN), and 17 wk IUGR males (which have HTN) vs CON males. We investigated the same time points in NT females. We divided pups from normal pregnant and PE Sprague-Dawley rats into groups based on pregnancy status and sex. Pups were aged to 12 and 17 wks. We collected brains to measure OS via western blot (Manganese superoxide dismutase) and colorimetric assays: hydrogen peroxide (H 2 O 2 ) and total antioxidant capacity (TAC). 12 wk IUGR and CON offspring were NT with no changes between groups or sex. There were no changes in cerebral H 2 O 2 , TAC, and MnSOD for IUGR offspring compared to their CON. At 17 wks, IUGR males had elevated mean arterial pressure (MAP; 143.0 ± 1.7 vs 133.0 ± 2.7 mmHg; p < 0.01). IUGR females were NT. Cerebral H2O2 levels in IUGR males were elevated (20.5 ± 8.0 vs 3.7 ± 1.0 uM/ mg Protein; p < 0.05) and unchanged in IUGR females. TAC in IUGR males was decreased (157.7 ±21.9 vs 234.9 ± 22.0 mM Trolox/mg Protein; p < 0.05) and unchanged in IUGR females. MnSOD was elevated in females with no differences between groups (150.4 ± 3.8 vs 110.9 ±7.1 IU/Protein/CON%; p < 0.002). All 12 wk rats were NT with no changes in cerebral OS. At 17 wks, IUGR males were HTN with elevated cerebral OS, with no changes in females. The lack of increase in OS at 12 wks, suggest that it’s not a precursor to HTN in males, but rather OS is a downstream effect from HTN. Future studies will look at factors that contribute to HTN and CVD in in IUGR offspring. Studies will also examine why 17 wk female IUGR are protected from HTN and cerebral OS.

S-43-3: INTRAUTERINE GROWTH RESTRICTION CAUSES AN ALTERATION IN KIDNEY GENE EXPRESSION

High blood pressure (BP) and kidney disease are the leading cause of death in the world population. Experimental and epidemiological studies indicate that the fetal environment is critical in determining adult cardiovascular (CV)-renal outcome. It is known that intrauterine growth restriction (IUGR) or low birth weight (LBW) induces an increased risk of hypertension and kidney disease in later life. Placental insufficiency is the common cause of IUGR. Low birth weight is a significant public health problem globally and is associated with short- and long-term consequences. About 15–20% of all births worldwide are LBW, representing more than 20 million births a year. Clinical studies indicate that birth weight is positively associated with glomerular filtration rate (GFR) and negatively associated with BP. Also, animal models of IUGR exhibit a higher susceptibility to ANGII induced high BP and kidney damage in response to acute ischemic reperfusion injury. Thus, these data suggest that IUGR programs greater risk of hypertension and renal dysfunction. However, the exact mechanisms linking an adverse fetal environment with a later increased risk of these diseases are unclear. Objective: This study aims to evaluate the global genetic change in the kidney of the animal model of IUGR. Design: female C57Bl/6J were mated with male C57bl/6J. Upon confirming the pregnancy by sperm plug (day 0 of gestation), the pregnant animal will be undergone reduced uterine perfusion surgery (RUP) or shame surgery at day 13 of gestation. The offspring of RUP mom will be labeled as IUGR, and from the sham will be labeled as control. The offspring will be used for blood pressure measurement using radio-telemetry and kidney microarray analysis when they reach 24 weeks. Results: Th IUGR offspring significantly has an increase in blood pressure compared to the control. The kidneys of male IUGR have 6529 gene upregulation compared to male control. The make IUGR also has 5713 genes dow-regulation as compared to same-sex control. The female IUGR has 1635 genes upregulated and 1513 genes downregulated compared to the kidney from the female control group. Conclusion: the complication during pregnancy, such as reduced uterine perfusion, which is commonly found in preeclampsia programs, alters gene expression in the kidneys of their offspring. This finding suggests that IUGR causes the gene alteration in the kidneys, which is the primary organ in controlling the blood pressure of the offspring, and may lead to the development of hypertension and kidney disease.

Sex differences in cerebrovascular dysfunction and hypertension in offspring of hypertensive pregnant rats

BACKGROUNDOffspring from preeclamptic women have a greater risk of hypertension and cerebrovascular dysfunction later in life. One of the major contributors of cerebrovascular dysfunction is impaired cerebral blood flow (CBF) autoregulation. In the USA, 1 in 25 pregnancies are preeclamptic. Preeclampsia, as defined as new‐onset hypertension during pregnancy, is an inflammatory condition characterized by elevated interleukin 17 (IL‐17), mitochondrial reactive oxygen species (mtROS), mitochondrial dysfunction (mt‐Dys), and intrauterine growth restriction (IUGR). Preliminary work from our laboratory demonstrates that offspring from preeclamptic rats have sex differences. Males have hypertension and a greater impairment of CBF autoregulation, while females have no change in blood pressure with impaired CBF autoregulation. The exact mechanisms for the sex differences in cerebrovascular dysfunction and hypertension is unknown and is the focus of this study. We hypothesize that changes in cerebral mt‐Dys, cerebral mtROS, and circulating IL‐17 contributes to the sex differences in hypertension and cerebrovascular dysfunction in male and female offspring from preeclamptic rats.METHODSIn this study, we compared male and female offspring from normal pregnant and preeclamptic Sprague Dawley rats. All offspring were divided into controls (CON) and IUGR by sex. Hypertensive male and non‐hypertensive female offspring’s with impaired CBF autoregulation were examined at 17 weeks of age for changes in cerebral mitochondrial electron transport chain (ETC) protein complexes, cerebral manganese superoxide dismutase (MnSOD), and circulating IL‐17.RESULTSFemale CON (151±5% IU/Protein/male CON) and IUGR (149±8 %IU/Protein/male CON) offspring have increased MnSOD compared to CON (100±7 %IU/Protein/male CON) and IUGR (122±4 %IU/Protein/male CON) males (p<0.05). No changes in female ETC protein complexes between IUGR and CON. Male IUGR have a decrease in complex II (71±5 vs. 100±8 %IU/Protein/male CON, p<0.05) and V (57±2 vs. 100±10 %IU/Protein/male CON, p<0.05) ETC proteins, and elevated IL‐17 (944±370 vs. 412±115 pg/mL, ns) compared to CON males.CONCLUSIONMale IUGR offspring have mt‐Dys and elevated IL‐17, which may contribute to hypertension and a greater impairment in CBF autoregulation. Female IUGR offspring may be protected due to an increase in mitochondrial antioxidants. In summary, studying the dimorphic sex differences in the mechanisms of hypertension and cerebrovascular dysfunction in offspring of preeclamptic women, may improve the offspring’s risk of hypertension, cardiovascular disease, stroke, and cognitive dysfunction later in life.

No evidence of the unfolded protein response in the placenta of two rodent models of preeclampsia and intrauterine growth restriction.

In humans, intrauterine growth restriction (IUGR) and preeclampsia (PE) are associated with induction of the unfolded protein response (UPR) and increased placental endoplasmic reticulum (ER) stress. Especially in PE, oxidative stress occurs relative to the severity of maternal vascular underperfusion (MVU) of the placental bed. On the premise that understanding the mechanisms of placental dysfunction could lead to targeted therapeutic options for human IUGR and PE, we investigated the roles of the placental UPR and oxidative stress in two rodent models of these human gestational pathologies. We employed a rat IUGR model of gestational maternal protein restriction, as well as an endothelial nitric oxide synthase knockout mouse model (eNOS-/-) of PE/IUGR. Placental expression of UPR members was analyzed via qRT-PCR (Grp78, Calnexin, Perk, Chop, Atf6, and Ern1), immunohistochemistry, and Western blotting (Calnexin, ATF6, GRP78, CHOP, phospho-eIF2α, and phospho-IRE1). Oxidative stress was determined via Western blotting (3-nitrotyrosine and 4-hydroxy-2-nonenal). Both animal models showed a significant reduction of fetal and placental weight. These effects did not induce placental UPR. In contrast to human data, results from our rodent models suggest retention of placental plasticity in the setting of ER stress under an adverse gestational environment. Oxidative stress was significantly increased only in female IUGR rat placentas, suggesting a sexually dimorphic response to maternal malnutrition. Our study advances understanding of the involvement of the placental UPR in IUGR and PE. Moreover, it emphasizes the appropriate choice of animal models researching various aspects of these pregnancy complications.

Liver transcriptome profiling and functional analysis of intrauterine growth restriction (IUGR) piglets reveals a genetic correction and sexual-dimorphic gene expression during postnatal development

BackgroundIntrauterine growth restriction (IUGR) remains a major problem associated with swine production. Thus, understanding the physiological changes of postnatal IUGR piglets would aid in improving growth performance. Moreover, liver metabolism plays an important role in the growth and survival of neonatal piglets.ResultsBy profiling the transcriptome of liver samples on postnatal Days 1, 7, and 28, our study focused on characterizing the growth, function, and metabolism in the liver of IUGR neonatal piglets. Our study demonstrates that the livers of IUGR piglets were associated with a series of complications, including inflammatory stress and immune dysregulation; cytoskeleton and membrane structure disorganization; dysregulated transcription events; and abnormal glucocorticoid metabolism. In addition, the abnormal liver function index in the serum [alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total protein (TP)], coupled with hepatic pathological and ultrastructural morphological changes are indicative of liver damage and dysfunction in IUGR piglets. Moreover, these results reveal the sex-biased developmental dynamics between male and female IUGR piglets, and that male IUGR piglets may be more sensitive to disrupted metabolic homeostasis.ConclusionsThese observations provide a detailed reference for understanding the mechanisms and characterizations of IUGR liver functions, and suggest that the potential strategies for improving the survival and growth performance of IUGR offspring should consider the balance between postnatal catch-up growth and adverse metabolic consequences. In particular, sex-specific intervention strategies should be considered for both female and male IUGR piglets.

Sex Differences in Fetal Programming of Blood Pressure and Kidney Gene Expression of Intrauterine Growth Restricted Mouse Model

The fetal environment is critical in determining future cardiovascular‐renal function in adulthood. Intrauterine growth restriction (IUGR) results in an unfavorable intrauterine milieu, and can lead to hypertension, renal, and cardiovascular disease of the offspring later in life. However, the underlying mechanisms linking IUGR to cardiovascular and renal afflictions remain unclear. IUGR offspring exhibit impaired kidney development. Furthermore, IUGR reprograms kidney function, resulting in an increase in blood pressure (BP) and susceptibility to kidney injury in the adult. While some renal factors have been postulated, others remain to be elucidated. Here, we have utilized a relevant preclinical mouse model of IUGR induced by reduced uterine perfusion pressure (RUP) to explore the effect of IUGR on renal gene expression patterns as well as sex differences. Pregnant C57bl/6J mice underwent sham or reduced uterine perfusion (RUP) at day 13 of gestation with delivery at full term. IUGR offspring (from RUP dam) had a lower birth weight than control (p<0.05). At 24 weeks of age, control and IUGR mice were subjected to BP measurement. Kidney tissues from control and IUGR mice were isolated and mRNA from these tissues was subjected to microarray analysis to determine differential gene expression. We found that male IUGR offspring had a significantly higher BP compared to male controls (p<0.05). No significant differences in BP were found between female IUGR and female controls, suggesting that sex differences in IUGR programs increased BP in mice. Microarray analysis, at a statistical cutoff of p < 0.05, demonstrates clear sex differences between IUGR and control mice. Of the total number of genetic elements (n = 39,724), 8,356 were significantly different due to IUGR alone, with the majority of these differences occurring in the males (8,319). Network analysis indicated a strong effect on several pathways such as protein processing in the endoplasmic reticulum, oxidative phosphorylation, and mTOR signaling pathways in male IUGR compared to male controls. Interestingly, two pathways (PPAR signaling and AMPK signaling) were significantly altered by IUGR in the female group. In conclusion, there is a clear sex difference in the effect of IUGR on BP and kidney gene expression. The functional implications of these novel factors need to be further investigated.Support or Funding InformationDr. Intapad was supported by funding from the AHA 16SDG27770041, ASN Kidney Research Career Development grant, and grant P20GM109036 K. J. Maeda was supported by National Institutes of Health Grants T32HL‐105324 and 20PRE352000021. Dr. Garrett was supported by P20GM‐103476, P20GM‐104357 and P20GM‐121334

Uteroplacental Insufficiency Impairs Cholesterol Elimination in Adult Female Growth-Restricted Rat Offspring Fed a High-Fat Diet.

Uteroplacental insufficiency (UPI) causes intrauterine growth restriction (IUGR) and increases the risk of hypercholesterolemia and cardiovascular disease, which are leading causes of morbidity and mortality worldwide. Little is known about the mechanism through which UPI increases cholesterol. Hepatic Cholesterol 7 alpha-hydroxylase (Cyp7a1) is the rate-limiting and most highly regulated step of cholesterol catabolism to bile acids. Cholesterol 7 alpha-hydroxylase is regulated by transcription factor liver X receptor α (Lxrα) and by microRNA-122. We previously showed that microRNA-122 inhibition of Cyp7a1 translation decreased cholesterol catabolism to bile acids in female IUGR rats at the time of weaning. We hypothesized that UPI would increase cholesterol and microRNA-122 and decrease Cyp7a1 protein and hepatic bile acids in young adult female IUGR rats. To test our hypothesis, we used a rat model of IUGR induced by bilateral uterine artery ligation. Both control and IUGR offspring were exposed to a maternal high-fat diet from before conception through lactation, and all offspring were weaned to a high-fat diet on postnatal day 21. At postnatal day 60, IUGR female rats had increased total and low-density lipoprotein serum cholesterol and hepatic cholesterol, decreased Lxrα and Cyp7a1 protein, and decreased hepatic bile acids. Hepatic microRNA-122 was not changed by UPI. Our findings suggest that UPI decreased cholesterol catabolism to bile acids in young adult female rats through a mechanism independent of microRNA-122.

Placental ESRRG-CYP19A1 Expressions and Circulating 17-Beta Estradiol in IUGR Pregnancies.

Introduction: Sex steroids are regulating factors for intrauterine growth. 17-β Estradiol (E2) is particularly critical to a physiological pregnancy, as increased maternal E2 was correlated to lower fetal weight at delivery. The placenta itself is a primary source of estrogens, synthetized from cholesterol precursors. Cytochrome P450 aromatase (encoded by CYP19A1 gene) is a rate-limiting enzyme for E2 biosynthesis. CYP19A1 transcription is supported by Estrogen Related-Receptor Gamma (ERRγ- ESRRG gene), which thus has an indirect role in placental steroidogenesis. Here we investigated maternal E2 levels and placental CYP19A1 and ESRRG expressions in pregnancies with IntraUterine Growth Restriction (IUGR).Methods: Singleton pregnancies were studied. E2 was measured in maternal plasma by electrochemiluminescence in 16 term controls and 11 IUGR (classified by umbilical artery doppler pulsatility index) at elective cesarean section, and also in 13 controls during pregnancy at a gestational age comparable to IUGR. CYP19A1 and ESRRG expressions were analyzed in placental tissue. Maternal/fetal characteristics, placental and molecular data were compared among study groups and tested for correlations.Results: Maternal E2 plasma concentrations were significantly decreased in IUGR compared to controls at delivery. When analyzing normal pregnancies at a gestational age similar to IUGR, E2 levels were not different to pathological cases. However, E2 levels at delivery positively correlated with placental efficiency. Placental CYP19A1 levels were significantly higher in IUGR placental tissue vs. controls, and specifically increased in female IUGR placentas. ESRRG expression was not different among groups.Discussion: We report a positive correlation between 17-β Estradiol levels and placental efficiency, that might indicate a disrupted steroidogenesis in IUGR pregnancies. Moreover, we show alterations of CYP19A1 expression in IUGR placentas, possibly indicating a compensatory effect to the adverse IUGR intrauterine environment, also depending on fetal sex. Further studies are needed to deeper investigate IUGR alterations in the complex interaction among molecules involved in placental steroidogenesis.

Activation of G Protein‐Coupled Estrogen Receptor 1, with a GPER Agonist, Reduces Blood Pressure in Female Growth‐Restricted Rats in Later Life

The prevalence of hypertension (HTN) is two‐fold higher in low birth weight (LBW) women at age 60 relative to their normal birth weight (NBW) counterparts. Yet, the mechanisms that contribute to greater cardiovascular risk in LBW women are not known. In our model of intrauterine growth restriction (IUGR), induced by placental insufficiency in the pregnant rat, female IUGR offspring have an elevated blood pressure (BP) at 12 months of age that is associated with early reproductive senescence. Although estrogen levels do not differ between control and IUGR groups, there is a significant increase in testosterone levels in female IUGR that is not seen in female control. Elevated testosterone levels, due to an imbalance in the hormonal milieu, is suggested to contribute to the development of hypertension in women in later life. We previously reported that chronic administration of estradiol is able to abolish the increase in blood pressure in the female IUGR rat at 12 months of age. However, this was not accomplished through alterations in expression of the nuclear estrogen receptor alpha (ER‐alpha). In the current study, we tested the hypothesis that activation of G protein‐coupled estrogen receptor 1 (GPER) would reduce blood pressure in the female IUGR rat. The selective GPER agonist, G1 (400 g/kg/day), or vehicle was administered via osmotic pumps for 2 weeks beginning at 14 months of age. BP was measured in conscious, chronically catheterized rats following treatment (2way ANOVA; Tukey's multiple comparison). As shown previously, BP was significantly elevated in vehicle‐treated IUGR relative to vehicle‐treated control (138.3± 4.01 vs. 124.3 ± 0.70mmHG, respectively). BP was significantly decreased in G1‐treated IUGR relative to vehicle‐treated IUGR (125.14±4.03 vs. 138.3 ± 4.01mmHG, respectively), whereas BP was not significantly altered in G1‐treated control relative to vehicle‐treated control (124.5 ± 0.66 vs. 124.3 ± 0.70mmHG, respectively). Uterine weight did not differ between groups. Therefore, this data suggests that activation of GPER was able to abolish the increase in blood pressure in the female IUGR rat in later life. This study suggests that GPER may serve as a therapeutic target for ameliorating increases in blood pressure seen in LBW women in later life.Support or Funding InformationNIH: R56HL143459, P20GM121334, P20GM104357, P01‐HL51971, T32HL105324, F30DK112718. AHA: GRNT19900004, PRE34060010.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Placental insufficiency contributes to fatty acid metabolism alterations in aged female mouse offspring.

Intrauterine growth restriction (IUGR) is an accepted risk factor for metabolic disorders in later life, including obesity and type 2 diabetes. The level of metabolic dysregulation can vary between subjects and is dependent on the severity and the type of IUGR insult. Classical IUGR animal models involve nutritional deprivation of the mother or uterine artery ligation. The latter aims to mimic a placental insufficiency, which is the most frequent cause of IUGR. In this study, we investigated whether IUGR attributable to placental insufficiency impacts the glucose and lipid homeostasis at advanced age. Placental insufficiency was achieved by deletion of the transcription factor AP-2y ( Tfap2c), which serves as one of the major trophoblast differentiation regulators. TdelT-IUGR mice were obtained by crossing mice with a floxed Tfap2c allele and mice with Cre recombinase under the control of the Tpbpa promoter. In advanced adulthood (9-12 mo), female and male IUGR mice are respectively 20% and 12% leaner compared with controls. At this age, IUGR mice have unaffected glucose clearance and lipid parameters (cholesterol, triglycerides, and phospholipids) in the liver. However, female IUGR mice have increased plasma free fatty acids (+87%) compared with controls. This is accompanied by increased mRNA levels of fatty acid synthase and endoplasmic reticulum stress markers in white adipose tissue. Taken together, our results suggest that IUGR by placental insufficiency may lead to higher lipogenesis in female mice in advanced adulthood, at least indicated by greater Fasn expression. This effect was sex specific for the aged IUGR females.

Nonhuman primate breath volatile organic compounds associate with developmental programming and cardio-metabolic status

Rodent and nonhuman primate studies indicate that developmental programming by reduced perinatal nutrition negatively impacts life course cardio-metabolic health. We have developed a baboon model in which we feed control mothers (CON) ad libitum while nutrient restricted mothers are fed 70% of ad libitum global feed in pregnancy and lactation. Offspring of nutrient restricted mothers are intrauterine growth restricted (IUGR) at term. By 3.5 years IUGR baboons showed signs of insulin resistance, indicating a pre-diabetic phenotype, in contrast to healthy CON offspring. We hypothesized that a novel breath analysis approach would provide markers of the altered cardio-metabolic state in a non-invasive manner. Here we assess whether exhaled breath volatile organic compounds (VOCs) collected from this unique cohort of juvenile baboons with documented cardio-metabolic dysfunction resulting from in utero programming can be detected from their breath signatures. Breath was collected from male and female CON and IUGR baboons at 4.8 ± 0.2 years (human equivalent ∼13 years). Breath VOCs were quantified using a two-dimensional gas chromatography mass spectrometer. Two-way ANOVA, on 76 biologically relevant VOCs identified 27 VOCs (p < 0.05) with altered abundances between groups (sex, birthweight, and sex x birthweight). The 27 VOCs included 2-pentanone, 2-octanone, 2,2,7,7-tetramethyloctane and 3-methyl-1-heptene, which have not previously been associated with cardio-metabolic disease. Unsupervised principal component analysis of these VOCs could discriminate the four clusters defining males, females, CON and IUGR. This study, which is the first to assess quantifiable breath signatures associated with cardio-metabolic programing for any model of IUGR, demonstrates the translational value of this unique model to identify metabolites of programmed cardio-metabolic dysfunction in breath signatures. Future studies are required to validate the translatability of these findings to humans.

Sex Differences in Distal Tubular Injury and Renal Inflammation in Intrauterine Growth‐ Restricted Offspring After Uni‐nephrectomy in Late Adulthood

Low birth weight (LBW) and aging are well‐established risk factors for kidney disease, but little is known about the compounded effect of aging in LBW individuals. Our laboratory uses a model of intrauterine growth restriction (IUGR) induced by placental insufficiency in the rat to study the association between LBW and chronic disease. We previously reported that male IUGR offspring have elevated blood pressure (BP) at 3 months of age, while female IUGR are protected from increased BP until 12 months of age. Whether sex alters the risk of renal disease in LBW individuals is unknown. Although hypertension does not completely account for the increased risk of kidney disease in LBW individuals, it is a likely contributor. Therefore, we hypothesize that male IUGR offspring will exhibit greater evidence of kidney injury compared to female IUGR due to the longer exposure to high BP. We tested the hypothesis that male IUGR offspring are programmed for enhanced susceptibility to a chronic renal insult during late adulthood while female IUGR offspring remain protected. At 18 months of age, rats underwent uni‐nephrectomy (Uni‐X) or a sham procedure (Sham). After one month, glomerular filtration rate (GFR) was measured by FITC‐sinistrin clearance, urinary markers of renal injury and expression of renal inflammatory markers were quantified, and histological injury was scored by two blinded observers. Male offspring (control Sham, IUGR Sham, control Uni‐X, and IUGR Uni‐X) had greater renal mass determined by kidney weight and kidney surface area, increased renal injury determined by proteinuria and glomerular injury scores, and increased glomerular area compared to their female counterparts at this age, but these parameters did not differ between Sham versus Uni‐X groups. When normalized to body weight, the remnant kidney was larger in male Uni‐X IUGR (4.76±0.35 mg/g; P&lt;0.05) compared to male Sham IUGR (3.03±0.21), female Sham IUGR (3.05±0.10), and female Uni‐X IUGR (3.79±0.13), n=7/group, demonstrating greater renal hypertrophy in male IUGR offspring. GFR was significantly lower in male Uni‐X IUGR compared to female Uni‐X IUGR offspring at 19 months of age (0.49±0.10 versus 0.81±0.06 mL/min/g KW, males versus females respectively; P&lt;0.05, n=9/group). Neutrophil gelatinase‐associated lipocalin, a urinary marker of distal tubular injury, was elevated in male Uni‐X IUGR offspring (59,871±18,697 ng/day; P&lt;0.05) compared to male Sham IUGR (20,603±4,078) and female Uni‐X IUGR offspring (14,823±2,798), n=7/group. In addition, cortical mRNA expression of TNF‐α and TGF‐β was greater in male Uni‐X IUGR offspring compared to male Sham IUGR and male Uni‐X controls. These results demonstrate that IUGR leads to greater renal hypertrophy, distal tubular injury, and renal inflammation in response to a chronic renal insult during late adulthood in males. In summary, in addition to differences in kidney structure and injury that exist between men and women in the general population with aging, IUGR programs an additional sex difference in the renal response to a chronic renal insult in later life.Support or Funding InformationRO1HL074927, HL51971, P20GM104357, AHAGRNT19900004, T32HL105324, F30DK112718This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Effects of Intrauterine Growth Restriction (IUGR) on the Surfactant System in Response to Sepsis

IntroductionAcute respiratory distress syndrome (ARDS) is defined as severe lung dysfunction. The lung impairments in ARDS result, in part, from alterations to pulmonary surfactant; a lipid‐protein mixture coating the inside of the lung that increases the lung compliance during respiration. The most common cause of ARDS is sepsis and due to a lack of effective pharmacological therapies the mortality associated with this condition is over 30%. Our research focuses on risk factors that indicate a susceptibility to the disease, which could provide new and early therapeutic options. One such potential risk factor is intrauterine growth restriction (IUGR). IUGR is defined by a low birth weight and contributes to a variety of adult‐onset diseases with different severities between males and females. There are underlying shared mechanisms between these IUGR‐induced diseases and ARDS, such as systemic inflammation. This study investigates the IUGR effect on surfactant system in response to sepsis in adult males and females. It was hypothesized that IUGR is a risk factor for developing ARDS through surfactant alterations with different outcomes between adult males and females.MethodsIUGR was induced in pregnant rat model via a low protein diet. At postnatal day 130, control and IUGR rats were randomly assigned into sham and septic groups. Rats were anesthetized and received either an intraperitoneal injection (IP) of saline or fecal slurry solution (1m/100g at 0.5g/ml). Following injection, respiratory rate and the animal activity were monitored for 6hr. After 6hr, animals were sacrificed (sodium pentobarbital IP injection) and arterial blood and the surfactant were taken for cytokine and surfactant analysis. Some non‐lavaged lungs were fixed for histological assessments.ResultsIUGR was confirmed by significant decrease in birth weight in both male and female offspring. Septicemia was confirmed by culturing bacteria from the blood samples, while cultures from sham animals were negative. After 6hr, respiratory rates significantly decreased in septic male and female IUGRs compare to sham male and female IUGRs at time zero (before IP) (p&lt;0.01). Surfactant analysis showed significant reduction in the total surfactant phospholipid content in septic male IUGRs compared to shams (p&lt;0.05) with no changes in surfactant phospholipid content between female septic IUGRs and shams. Control groups demonstrated no changes in surfactant content between septics and shams in either sexes.DiscussionChanges in surfactant pools occurring between male IUGR septic animals compared to shams without changes in control animals, support our hypothesis that IUGR is a risk factor for developing surfactant impairments. Also, the results indicate that the effect is different between males and females. Reduction of surfactant content in septic IUGR males could potentially cause lower lung compliance, alter lung function and therefore, developing or exacerbating lung injury during the development of ARDS. To further study the IUGR effect on the surfactant and evaluate its role as an ARDS risk factor, future experiments will assess the surfactant biophysical properties and lung histology.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Role of fetal nutrient restriction and postnatal catch-up growth on structural and mechanical alterations of rat aorta.

Intrauterine growth restriction (IUGR), induced by maternal undernutrition, leads to impaired aortic development. This is followed by hypertrophic remodelling associated with accelerated growth during lactation. Fetal nutrient restriction is associated with increased aortic compliance at birth and at weaning, but not in adult animals. This mechanical alteration may be related to a decreased perinatal collagen deposition. Aortic elastin scaffolds purified from young male and female IUGR animals also exhibit increased compliance, only maintained in adult IUGR females. These mechanical alterations may be related to differences in elastin deposition and remodelling. Fetal undernutrition induces similar aortic structural and mechanical alterations in young male and female rats. Our data argue against an early mechanical cause for the sex differences in hypertension development induced by maternal undernutrition. However, the larger compliance of elastin in adult IUGR females may contribute to the maintenance of a normal blood pressure level. Fetal undernutrition programmes hypertension development, males being more susceptible. Deficient fetal elastogenesis and vascular growth is a possible mechanism. We investigated the role of aortic mechanical alterations in a rat model of hypertension programming, evaluating changes at birth, weaning and adulthood. Dams were fed ad libitum (Control) or 50% of control intake during the second half of gestation (maternal undernutrition, MUN). Offspring aged 3days, 21days and 6months were studied. Blood pressure was evaluated in vivo. In the thoracic aorta we assessed gross structure, mechanical properties (intact and purified elastin), collagen and elastin content and internal elastic lamina (IEL) organization. Only adult MUN males developed hypertension (systolic blood pressure: MUNmales =176.6±5.6mmHg; Controlmales =136.1±4.9mmHg). At birth MUN rats were lighter, with smaller aortic cross-sectional area (MUNmales =(1.51±0.08)×105 μm2 , Controlmales =(2.8±0.04)×105 μm2 ); during lactation MUN males and females exhibited catch-up growth and aortic hypertrophy (MUNmales =(14.5±0.5)×105 μm2 , Controlmales =(10.4±0.9)×105 μm2 ), maintained until adulthood. MUN aortas were more compliant until weaning (functional stiffness: MUNmales =1.0±0.04; Controlmales =1.3±0.03), containing less collagen with larger IEL fenestrae, returning to normal in adulthood. Purified elastin from young MUN offspring was more compliant in both sexes; only MUN adult females maintained larger elastin compliance (slope: MUNfemales =24.1±1.9; Controlfemales =33.3±2.8). Fetal undernutrition induces deficient aortic development followed by hypertrophic remodelling and larger aortic compliance in the perinatal period, with similar alterations in collagen and elastin in both sexes. The observed alterations argue against an initial mechanical cause for sex differences in hypertension development. However, the maintenance of high elastin compliance in adult females might protect them against blood pressure rise.

Abstract 046: Chronic Estrogen Replacement Prevents the Increase in Blood Pressure in Female IUGR Offspring at 12 Months of Age

Low birth weight (LBW ) is associated with an earlier age at menopause and a greater prevalence of hypertension in women in later life. Using a rodent model of LBW induced by placental insufficiency, previous studies from our lab indicate that intrauterine growth restriction ( IUGR ) programs a significant increase in blood pressure ( BP ) by 12 months of age in female IUGR offspring. Female IUGR are also acyclic by 12 months of age, or 6 months prior to control indicative of early reproductive senescence. The mechanisms involved in menopause related cardiovascular risk are not clear. Furthermore, the link between birth weight and the greater prevalence for hypertension in LBW women in later life is not known. Female IUGR exhibit a shift in the testosterone to estradiol ratio at 12 months of age. Thus, this study tested the hypothesis that a loss of estradiol ( E2 ) contributes to the increase in BP in female IUGR offspring that develops by 12 months of age. Female rats received vehicle or 17β-Estradiol valerate minipellets (1.5mg for 60-day release) for 6 weeks starting at 12 months of age with BP measured in conscious, chronically catheterized rats following treatment (2way ANOVA; Tukey’s multiple comparison). BP was significantly increased in vehicle-treated IUGR relative to vehicle-treated Control (*, p&lt;0.05); an increase that was abolished by chronic E2 in IUGR (**, p&lt;0.05). Uterine weight, a crude marker of E2, was significantly increased by chronic E2 in both Control and IUGR E2-treated offspring (**, p&lt;0.05). Therefore, this study indicates that E2 withdrawal after early reproductive senescence contributes to the increase in BP that develops in later life in female IUGR offspring.

Sexual dimorphism in the fetal cardiac response to maternal nutrient restriction

Poor maternal nutrition causes intrauterine growth restriction (IUGR); however, its effects on fetal cardiac development are unclear. We have developed a baboon model of moderate maternal undernutrition, leading to IUGR. We hypothesized that the IUGR affects fetal cardiac structure and metabolism. Six control pregnant baboons ate ad-libitum (CTRL)) or 70% CTRL from 0.16 of gestation (G). Fetuses were euthanized at C-section at 0.9G under general anesthesia. Male but not female IUGR fetuses showed left ventricular fibrosis inversely correlated with birth weight. Expression of extracellular matrix protein TSP-1 was increased (p<0.05) in male IUGR. Expression of cardiac fibrotic markers TGFβ, SMAD3 and ALK-1 were downregulated in male IUGRs with no difference in females. Autophagy was present in male IUGR evidenced by upregulation of ATG7 expression and lipidation LC3B. Global miRNA expression profiling revealed 56 annotated and novel cardiac miRNAs exclusively dysregulated in female IUGR, and 38 cardiac miRNAs were exclusively dysregulated in males (p<0.05). Fifteen (CTRL) and 23 (IUGR) miRNAs, were differentially expressed between males and females (p<0.05) suggesting sexual dimorphism, which can be at least partially explained by differential expression of upstream transcription factors (e.g. HNF4α, and NFκB p50). Lipidomics analysis of fetal cardiac tissue exhibited a net increase in diacylglycerol and plasmalogens and a decrease in triglycerides and phosphatidylcholines. In summary, IUGR resulting from decreased maternal nutrition is associated with sex-dependent dysregulations in cardiac structure, miRNA expression, and lipid metabolism. If these changes persist postnatally, they may program offspring for higher later life cardiac risk.

Intrauterine Growth Restriction Programs Greater Susceptibility to Chronic Kidney Injury in Male and Female Aged Rats

Although the correlation between low birth weight (LBW) and increased risk of chronic kidney disease (CKD) is well‐established, the mechanisms involved remain unknown. Our laboratory uses a rodent model of intrauterine growth restriction (IUGR) to study the etiology of the developmental programming of enhanced cardio‐renal risk. Previous studies by our laboratory report that blood pressure (BP) is increased in male IUGR offspring relative to male controls at 4 months of age whereas female IUGR remain normotensive. However, BP is elevated by 12 months of age in female IUGR indicating that female IUGR do not remain protected. Male IUGR exhibit an enhanced susceptibility to acute renal injury at 4 months of age, but whether female IUGR exhibit a similar response is unknown. Epidemiological data related to sex differences in this phenomenon are limited, especially with regard to the effect of aging, but suggest that males progress to renal failure more rapidly than females. Thus, this study tested the hypothesis that IUGR offspring exhibit a sex difference in enhanced susceptibility to a chronic renal insult in later life. At 16 ± 1 months of age, control and IUGR rats underwent sham or uni‐nephrectomy (UNI‐X) to reduce renal mass. 24‐hour metabolic studies were performed before and one‐month post‐surgery to compare urinary markers of kidney injury. After one month of recovery, rats underwent carotid and jugular catheterization with BP and glomerular filtration rate (GFR) measured in conscious rats. Measurement of GFR involved the transcutaneous clearance method using intravenous injection of FITC‐sinistrin. Data were analyzed by 2‐way ANOVA with Tukey's multiple comparisons post‐test. After one month of reduced renal mass, GFR was not altered in male and female controls (Table). However, GFR was significantly reduced in male and female UNI‐X IUGR relative to their same‐sex sham counterparts (Table). Although UNI‐X reduced GFR by 43% in male IUGR compared to a 28% reduction in female IUGR relative to same‐sex shams, this difference was not statistically significant. BP was not significantly altered post‐UNI‐X in any groups (data not shown). Renal cortical expression of TNF‐α and TGF‐β were significantly elevated by 9‐fold and 3‐fold (P&lt;0.05), respectively, in male UNI‐X IUGR compared to male sham IUGR and male control groups. This suggests that an exacerbated increase in renal inflammation in response to UNI‐X may be a contributory factor to enhanced susceptibility to chronic renal injury in male IUGR. Whether female UNI‐X IUGR exhibit a similar increase is not yet known. Therefore, these data suggest that both male and female IUGR demonstrate a greater susceptibility to chronic renal injury in later life than normal birthweight controls. Furthermore, this study indicates that optimization of treatments to improve the health of renal patients should involve a complete patient perinatal history and that investing in further efforts to understand the link between the in utero environment and adult health is invaluable.Support or Funding InformationNIH: F30DK112718‐01, T32HL105324‐07, HL074927, HL51971, P20GM104357; AHA: GRNT19900004. GFR (mL/min/100 g body weight) Male Female Control Sham 0.78 ± 0.06 0.89 ± 0.08 Control UNI‐X 0.54 ± 0.07 0.77 ± 0.03 IUGR Sham 0.71 ± 0.09 0.98 ± 0.07 IUGR UNI‐X 0.40 ± 0.10* 0.70 ± 0.02* P&lt;0.05 vs Same‐Sex Sham.