Sex-specific differences in the mechanisms for enhanced thromboxane A2-mediated vasoconstriction in adult offspring exposed to prenatal hypoxia.

BACKGROUND: Prenatal hypoxia, a common pregnancy complication, leads to impaired cardiovascular outcomes in the adult offspring. It results in impaired vasodilation in coronary and mesenteric arteries of the adult offspring, due to reduced nitric oxide (NO). Thromboxane A2 (TxA2) is a potent vasoconstrictor increased in cardiovascular diseases, but its role in the impact of prenatal hypoxia is unknown. To prevent the risk of cardiovascular disease by prenatal hypoxia, we have tested a maternal treatment using a nanoparticle-encapsulated mitochondrial antioxidant (nMitoQ). We hypothesized that prenatal hypoxia enhances vascular TxA2 responses in the adult offspring, due to decreased NO modulation, and that this might be prevented by maternal nMitoQ treatment. METHODS: Pregnant Sprague-Dawley rats received a single intravenous injection (100 µL) of vehicle (saline) or nMitoQ (125 µmol/L) on gestational day (GD)15 and were exposed to normoxia (21% O2) or hypoxia (11% O2) from GD15 to GD21 (term = 22 days). Coronary and mesenteric arteries were isolated from the 4-month-old female and male offspring, and vasoconstriction responses to U46619 (TxA2 analog) were evaluated using wire myography. In mesenteric arteries, L-NAME (pan-NO synthase (NOS) inhibitor) was used to assess NO modulation. Mesenteric artery endothelial (e)NOS, and TxA2 receptor expression, superoxide, and 3-nitrotyrosine levels were assessed by immunofluorescence. RESULTS: Prenatal hypoxia resulted in increased U46619 responsiveness in coronary and mesenteric arteries of the female offspring, and to a lesser extent in the male offspring, which was prevented by nMitoQ. In females, there was a reduced impact of L-NAME in mesenteric arteries of the prenatal hypoxia saline-treated females, and reduced 3-nitrotyrosine levels. In males, L-NAME increased U46619 responses in mesenteric artery to a similar extent, but TxA2 receptor expression was increased by prenatal hypoxia. There were no changes in eNOS or superoxide levels. CONCLUSIONS: Prenatal hypoxia increased TxA2 vasoconstrictor capacity in the adult offspring in a sex-specific manner, via reduced NO modulation in females and increased TP expression in males. Maternal placental antioxidant treatment prevented the impact of prenatal hypoxia. These findings increase our understanding of how complicated pregnancies can lead to a sex difference in the programming of cardiovascular disease in the adult offspring.

Prenatal hypoxia, when the fetus does not receive enough oxygen, is a common problem during pregnancy that impacts the developing fetus. It is associated with an increased risk of cardiovascular disease in the offspring in adulthood. While the mechanisms are not fully understood, the blood vessel function in the offspring may be impacted by prenatal hypoxia. We hypothesize that prenatal hypoxia increases the constriction of the blood vessels in the offspring. The placenta, an essential organ for fetal development, supplies oxygen and nutrients to the fetus. In prenatal hypoxia pregnancies, the placenta does not work properly. We have been studying a placental treatment (called nMitoQ) to improve placenta function and thereby the blood vessel function of the offspring. We used a rat model of prenatal hypoxia, where pregnant rats (dams) were placed in a low oxygen environment (hypoxia) during the last trimester of pregnancy. Control rats were kept in normal oxygen conditions. The dams were treated with nMitoQ, or with saline (control). Next, we studied the blood vessels of the offspring in adulthood. We found that prenatal hypoxia increases the constriction of the blood vessels, which was prevented by treating the dams with nMitoQ. Interestingly, this impact was more severe in females compared to males, and the mechanisms were different between the sexes. This study helps in the understanding of how complicated pregnancies can impair cardiovascular health in the offspring, and in a potential development of targeted and sex-specific therapies for those offspring at high risk for future cardiovascular disease.

Prenatal Hypoxia Triggers a Glucocorticoid-Associated Depressive-like Phenotype in Adult Rats, Accompanied by Reduced Anxiety in Response to Stress.

Fetal hypoxia and maternal stress frequently culminate in neuropsychiatric afflictions in life. To replicate this condition, we employed a model of prenatal severe hypoxia (PSH) during days 14-16 of rat gestation. Subsequently, both control and PSH rats at 3 months old were subjected to episodes of inescapable stress to induce learned helplessness (LH). The results of the open field test revealed an inclination towards depressive-like behavior in PSH rats. Following LH episodes, control (but not PSH) rats displayed significant anxiety. LH induced an increase in glucocorticoid receptor (GR) levels in extrahypothalamic brain structures, with enhanced nuclear translocation in the hippocampus (HPC) observed both in control and PSH rats. However, only control rats showed an increase in GR nuclear translocation in the amygdala (AMG). The decreased GR levels in the HPC of PSH rats correlated with elevated levels of hypothalamic corticotropin-releasing hormone (CRH) compared with the controls. However, LH resulted in a reduction of the CRH levels in PSH rats, aligning them with those of control rats, without affecting the latter. This study presents evidence that PSH leads to depressive-like behavior in rats, associated with alterations in the glucocorticoid system. Notably, these impairments also contribute to increased resistance to severe stressors.

The Prenatal Hypoxic Pathology Associated with Maternal Stress Predisposes to Dysregulated Expression of the chrna7 Gene and the Subsequent Development of Nicotine Addiction in Adult Offspring.

INTRODUCTION: Previous studies have shown that fetal hypoxia predisposes individuals to develop addictive disorders in adulthood. However, the specific impact of maternal stress, mediated through glucocorticoids and often coexisting with fetal hypoxia, is not yet fully comprehended. METHODS: To delineate the potential effects of these pathological factors, we designed models of prenatal severe hypoxia (PSH) in conjunction with maternal stress and prenatal intrauterine ischemia (PII). We assessed the suitability of these models for our research objectives by measuring HIF1α levels and evaluating the glucocorticoid neuroendocrine system. To ascertain nicotine dependence, we employed the conditioned place aversion test and the startle response test. To identify the key factor implicated in nicotine addiction associated with PSH, we employed techniques such as Western blot, immunohistochemistry, and correlational analysis between chrna7 and nr3c1 genes across different brain structures. RESULTS: In adult rats exposed to PSH and PII, we observed increased levels of HIF1α in the hippocampus (HPC). However, the PSH group alone exhibited reduced glucocorticoid receptor levels and disturbed circadian glucocorticoid rhythms. Additionally, they displayed signs of nicotine addiction in the conditioned place aversion and startle response tests. We also observed elevated levels of phosphorylated DARPP-32 protein in the nucleus accumbens (NAc) indicated compromised glutamatergic efferent signaling. Furthermore, there was reduced expression of α7 nAChR, which modulates glutamate release, in the medial prefrontal cortex (PFC) and HPC. Correlation analysis revealed strong associations between chrna7 and nr3c1 expression in both brain structures. CONCLUSION: Perturbations in the glucocorticoid neuroendocrine system and glucocorticoid-dependent gene expression of chrna7 associated with maternal stress response to hypoxia in prenatal period favor the development of nicotine addiction in adulthood.

Nicotinamide Riboside, an NAD + Precursor, Protects Against Cardiac Mitochondrial Dysfunction in Fetal Guinea Pigs Exposed to Gestational Hypoxia.

Gestational hypoxia inhibits mitochondrial function in the fetal heart and placenta contributing to fetal growth restriction and organ dysfunction. NAD + deficiency may contribute to a metabolic deficit by inhibiting oxidative phosphorylation and ATP synthesis. We tested the effects of nicotinamide riboside (NR), an NAD + precursor, as a treatment for reversing known mitochondrial dysfunction in hypoxic fetal hearts. Pregnant guinea pigs were housed in room air (normoxia) or placed in a hypoxic chamber (10.5%O2) for the last 14 days of gestation (term = 65 days) and administered either water or NR (1.6 mg/ml) in the drinking bottle. Fetuses were excised at term, and NAD + levels of maternal liver, placenta, and fetal heart ventricles were measured. Indices of mitochondrial function (complex IV activity, sirtuin 3 activity, protein acetylation) and ATP synthesis were measured in fetal heart ventricles of NR-treated/untreated normoxic and hypoxic animals. Hypoxia reduced fetal body weight in both sexes (p = 0.01), which was prevented by NR. Hypoxia had no effect on maternal liver NAD + levels but decreased (p = 0.04) placenta NAD + levels, the latter normalized with NR treatment. Hypoxia had no effect on fetal heart NAD + but decreased (p < 0.05) mitochondrial complex IV and sirtuin 3 activities, ATP content, and increased mitochondrial acetylation, which were all normalized with maternal NR. Hypoxia increased (p < 0.05) mitochondrial acetylation in female fetal hearts but had no effect on other mitochondrial indices. We conclude that maternal NR is an effective treatment for normalizing mitochondrial dysfunction and ATP synthesis in the hypoxic fetal heart.

Effects of Gestational Hypoxia on PGC1α and Mitochondrial Acetylation in Fetal Guinea Pig Hearts.

Chronic intrauterine hypoxia is a significant pregnancy complication impacting fetal heart growth, metabolism, and mitochondrial function, contributing to cardiovascular programming of the offspring. PGC1α (peroxisome proliferator-activated receptor γ co-activator 1α) is the master regulator of mitochondrial biogenesis. We investigated the effects of hypoxia on PGC1α expression following exposure at different gestational ages. Time-mated pregnant guinea pigs were exposed to normoxia (NMX, 21% O2) or hypoxia (HPX, 10.5% O2) at either 25-day (early-onset) or 50-day (late-onset) gestation, and all fetuses were extracted at term (term = ~65-day gestation). Expression of nuclear PGC1α, sirtuin 1 (SIRT1), AMP-activated protein kinase (AMPK), and mitochondrial sirtuin 3 (SIRT3) was measured, along with SIRT3 activity and mitochondrial acetylation of heart ventricles of male and female fetuses. Early-onset hypoxia increased (P<0.05) fetal cardiac nuclear PGC1α and had no effect on mitochondrial acetylation of either growth-restricted males or females. Late-onset hypoxia had either no effect or decreased (P<0.05) PCC1α expression in males and females, respectively, but increased (P<0.05) mitochondrial acetylation in both sexes. Hypoxia had variable effects on expression of SIRT1, AMPK, SIRT3, and SIRT3 activity depending on the sex. The capacity of the fetal heart to respond to hypoxia differs depending on the gestational age of exposure and sex of the fetus. Further, the effects of late-onset hypoxia on fetal heart function impose a greater risk to male than female fetuses, which has implications toward cardiovascular programming effects of the offspring.

Creatine supplementation reduces the cerebral oxidative and metabolic stress responses to acute in utero hypoxia in the late-gestation fetal sheep.

Prophylactic creatine treatment may reduce hypoxic brain injury due to its ability to sustain intracellular ATP levels thereby reducing oxidative and metabolic stress responses during oxygen deprivation. Using microdialysis, we investigated the real-time in vivo effects of fetal creatine supplementation on cerebral metabolism following acute in utero hypoxia caused by umbilical cord occlusion (UCO). Fetal sheep (118 days' gestational age (dGA)) were implanted with an inflatable Silastic cuff around the umbilical cord and a microdialysis probe inserted into the right cerebral hemisphere for interstitial fluid sampling. Creatine (6 mg kg-1  h-1 ) or saline was continuously infused intravenously from 122 dGA. At 131 dGA, a 10 min UCO was induced. Hourly microdialysis samples were obtained from -24 to 72 h post-UCO and analysed for percentage change of hydroxyl radicals (• OH) and interstitial metabolites (lactate, pyruvate, glutamate, glycerol, glycine). Histochemical markers of protein and lipid oxidation were assessed at post-mortem 72 h post-UCO. Prior to UCO, creatine treatment reduced pyruvate and glycerol concentrations in the microdialysate outflow. Creatine treatment reduced interstitial cerebral • OH outflow 0 to 24 h post-UCO. Fetuses with higher arterial creatine concentrations before UCO presented with reduced levels of hypoxaemia ( PO2${P_{{{\rm{O}}_{\rm{2}}}}}$ and SO2${S_{{{\rm{O}}_{\rm{2}}}}}$ ) during UCO which associated with reduced interstitial cerebral pyruvate, lactate and • OH accumulation. No effects of creatine treatment on immunohistochemical markers of oxidative stress were found. In conclusion, fetal creatine treatment decreased cerebral outflow of • OH and was associated with an improvement in cerebral bioenergetics following acute hypoxia. KEY POINTS: Fetal hypoxia can cause persistent metabolic and oxidative stress responses that disturb energy homeostasis in the brain. Creatine in its phosphorylated form is an endogenous phosphagen; therefore, supplementation is a proposed prophylactic treatment for fetal hypoxia. Fetal sheep instrumented with a cerebral microdialysis probe were continuously infused with or without creatine-monohydrate for 10 days before induction of 10 min umbilical cord occlusion (UCO; 131 days' gestation). Cerebral interstitial fluid was collected up to 72 h following UCO. Prior to UCO, fetal creatine supplementation reduced interstitial cerebral pyruvate and glycerol concentrations. Fetal creatine supplementation reduced cerebral hydroxyl radical efflux up to 24 h post-UCO. Fetuses with higher arterial creatine concentrations before UCO and reduced levels of systemic hypoxaemia during UCO were associated with reduced cerebral interstitial pyruvate, lactate and • OH following UCO. Creatine supplementation leads to some improvements in cerebral bioenergetics following in utero acute hypoxia.

Tissue-specific responses that constrain glucose oxidation and increase lactate production with the severity of hypoxemia in fetal sheep.

Fetal hypoxemia decreases insulin and increases cortisol and norepinephrine concentrations and may restrict growth by decreasing glucose utilization and altering substrate oxidation. Specifically, we hypothesized that hypoxemia would decrease fetal glucose oxidation and increase lactate and pyruvate production. We tested this by measuring whole body glucose oxidation and lactate production, and molecular pathways in liver, muscle, adipose, and pancreas tissues of fetuses exposed to maternal hypoxemia for 9 days (HOX) compared with control fetal sheep (CON) in late gestation. Fetuses with more severe hypoxemia had lower whole body glucose oxidation rates, and HOX fetuses had increased lactate production from glucose. In muscle and adipose tissue, expression of the glucose transporter GLUT4 was decreased. In muscle, pyruvate kinase (PKM) and lactate dehydrogenase B (LDHB) expression was decreased. In adipose tissue, LDHA and lactate transporter (MCT1) expression was increased. In liver, there was decreased gene expression of PKLR and MPC2 and phosphorylation of PDH, and increased LDHA gene and LDH protein abundance. LDH activity, however, was decreased only in HOX skeletal muscle. There were no differences in basal insulin signaling across tissues, nor differences in pancreatic tissue insulin content, ß-cell area, or genes regulating ß-cell function. Collectively, these results demonstrate coordinated metabolic responses across tissues in the hypoxemic fetus that limit glucose oxidation and increase lactate and pyruvate production. These responses may be mediated by hypoxemia-induced endocrine responses including increased norepinephrine and cortisol, which inhibit pancreatic insulin secretion resulting in lower insulin concentrations and decreased stimulation of glucose utilization.NEW & NOTEWORTHY Hypoxemia lowered fetal glucose oxidation rates, based on severity of hypoxemia, and increased lactate production. This was supported by tissue-specific metabolic responses that may result from increased norepinephrine and cortisol concentrations, which decrease pancreatic insulin secretion and insulin concentrations and decrease glucose utilization. This highlights the vulnerability of metabolic pathways in the fetus and demonstrates that constrained glucose oxidation may represent an early event in response to sustained hypoxemia and fetal growth restriction.

Placental pathology: Pathways leading to or associated with perinatal brain injury in experimental neurology, special issue: Placental mediated mechanisms of perinatal brain injury.

Perinatal brain injury is a multifactorial process. In utero placental physiology plays a major role in neuroprotection and the normal development of the fetal central nervous system. Advances in placental pathology have clarified several specific mechanisms of injury and the histologic lesions most strongly associated with them. This review provides an updated summary of the relevant placental anatomy and physiology, the specific placental pathways leading to brain injury, the revised Amsterdam classification system for placental pathology, and the known associations of specific placental lesions with subtypes of adverse neurologic outcomes.

Chronic Hypoxia Inhibits Respiratory Complex IV Activity and Disrupts Mitochondrial Dynamics in the Fetal Guinea Pig Forebrain.

Mitochondrial dysfunction is an underlying cause of childhood neurological disease secondary to the crucial role of mitochondria in proper neurodevelopment. We hypothesized that chronic intrauterine hypoxia (HPX) induces mitochondrial deficits by altering mitochondrial biogenesis and dynamics in the fetal brain. Pregnant guinea pigs were exposed to either normoxia (NMX, 21%O2) or HPX (10.5%O2) starting at 28-day (early onset, EO-HPX) or 50-day (late onset, LO-HPX) gestation until term (65 days). Near-term male and female fetuses were extracted from anesthetized sows, and mitochondria were isolated from excised fetal forebrains (n = 6/group). Expression of mitochondrial complex subunits I-V (CI-CV), fission (Drp-1), and fusion (Mfn-2) proteins was measured by Western blot. CI and CIV enzyme activities were measured by colorimetric assays. Chronic HPX reduced fetal body wts and increased (P < 0.05) brain/body wt ratios of both sexes. CV subunit levels were increased in EO-HPX males only and CII levels increased in LO-HPX females only compared to NMX. Both EO- and LO-HPX decreased CIV activity in both sexes but had no effect on CI activity. EO-HPX increased Drp1 and decreased Mfn2 levels in males, while LO-HPX had no effect on either protein levels. In females, both EO-HPX and LO-HPX increased Drp1 but had no effect on Mfn2 levels. Chronic HPX alters abundance and activity of select complex subunits and shifts mitochondrial dynamics toward fission in a sex-dependent manner in the fetal guinea pig brain. This may be an underlying mechanism of reduced respiratory efficiency leading to disrupted metabolism and increased vulnerability to a second neurological injury at the time of birth in HPX fetal brains.

Late-Pregnancy Fetal Hypoxia Is Associated With Altered Glucose Metabolism and Adiposity in Young Adult Offspring of Women With Type 1 Diabetes.

Objective: To investigate associations between exposure to fetal hypoxia and indicators of metabolic health in young adult offspring of women with type 1 diabetes (OT1D). Methods: 156 OT1D born between 7/1995 and 12/2000 at Helsinki University Hospital, Finland, were invited for follow-up between 3/2019 and 11/2019. A control group of 442 adults born from non-diabetic pregnancies, matched for date and place of birth, was obtained from the Finnish Medical Birth Register. In total, 58 OT1D and 86 controls agreed to participate. All OT1D had amniotic fluid (AF) sampled for erythropoietin (EPO) measurement within two days before delivery in order to diagnose fetal hypoxia. In total, 29 OTID had an AF EPO concentration <14.0 mU/l, defined as normal, and were categorized into the low EPO (L-EPO) group. The remaining 29 OT1D had AF EPO ≥14.0 mU/ml, defined as fetal hypoxia, and were categorized into the high EPO (H-EPO) group. At the age of 18-23 years, participants underwent a 2-h 75g oral glucose tolerance test (OGTT) in addition to height, weight, waist circumference, body composition, blood pressure, HbA1c, cholesterol, triglyceride, high-sensitivity CRP and leisure-time physical activity measurements. Results: Two OT1D were diagnosed with diabetes and excluded from further analyses. At young adult age, OT1D in the H-EPO group had a higher BMI than those in the L-EPO group. In addition, among female participants, waist circumference and body fat percentage were highest in the H-EPO group. In the OGTTs, the mean (SD) 2-h post-load plasma glucose (mmol/L) was higher in the H-EPO [6.50 (2.11)] than in the L-EPO [5.21 (1.10)] or control [5.67 (1.48)] offspring (p=0.009). AF EPO concentrations correlated positively with 2-h post-load plasma glucose [r=0.35 (95% CI: 0.07 to 0.62)] and serum insulin [r=0.44 (95% CI: 0.14 to 0.69)] concentrations, even after adjusting for maternal BMI, birth weight z-score, gestational age at birth and adult BMI. Control, L-EPO and H-EPO groups did not differ with regards to other assessed parameters. Conclusions: High AF EPO concentrations in late pregnancy, indicating fetal hypoxia, are associated with increased adiposity and elevated post-load glucose and insulin concentrations in young adult OT1D.

Chronic perinatal hypoxia delays cardiac maturation in a mouse model for cyanotic congenital heart disease.

Compared with acyanotic congenital heart disease (CHD), cyanotic CHD has an increased risk of lifelong mortality and morbidity. These adverse outcomes may be attributed to delayed cardiomyocyte maturation, since the transition from a hypoxic fetal milieu to oxygen-rich postnatal environment is disrupted. We established a rodent model to replicate hypoxic myocardial conditions spanning perinatal development, and tested the hypothesis that chronic hypoxia impairs cardiac development. Pregnant mice were housed in hypoxia beginning at embryonic day 16. Pups stayed in hypoxia until postnatal day (P)8 when cardiac development is nearly complete. Global gene expression was quantified at P8 and at P30, after recovering in normoxia. Phenotypic testing included electrocardiogram, echocardiogram, and ex vivo electrophysiology study. Hypoxic P8 animals were 47% smaller than controls with preserved heart size. Gene expression was grossly altered by hypoxia at P8 (1,427 genes affected), but normalized after recovery (P30). Electrocardiograms revealed bradycardia and slowed conduction velocity in hypoxic animals at P8, with noticeable resolution after recovery (P30). Notable differences that persisted after recovery (P30) included a 65% prolongation in ventricular effective refractory period, sinus node dysfunction, 23% reduction in ejection fraction, and 16% reduction in fractional shortening in animals exposed to hypoxia. We investigated the impact of chronic hypoxia on the developing heart. Perinatal hypoxia was associated with changes in gene expression and cardiac function. Persistent changes to the electrophysiological substrate and contractile function warrant further investigation and may contribute to adverse outcomes observed in the cyanotic CHD population.NEW & NOTEWORTHY We utilized a new mouse model of chronic perinatal hypoxia to simulate the hypoxic myocardial conditions present in cyanotic congenital heart disease. Hypoxia caused numerous abnormalities in cardiomyocyte gene expression, the electrophysiologic substrate of the heart, and contractile function. Taken together, alterations observed in the neonatal period suggest delayed cardiac development immediately following hypoxia.

CD15 immunostaining improves placental diagnosis of fetal hypoxia.

INTRODUCTION: Fetal hypoxic events with unclear predictive value are a common indication for placenta examination. We evaluated whether the use of CD15 immunostaining can improve the assessment of severity and duration of fetal hypoxia. METHODS: We compared placentas (37-42 gestational weeks) from stillborns/newborns with birth asphyxia (BA) and non-hypoxic newborns. Placental findings were studied in following groups: (1) acute BA (n = 11) due to placental abruption, (2) non-acute BA (n = 121) due to non-acute conditions, (3) non-BA (n = 46) in pregnancies with preeclampsia and gestational diabetes, and (4) controls (n = 30). RESULTS: A high expression of CD15 in feto-placental resistance vessels (FRVs) was present in non-acute BA (95.9%), but absent in acute BA, non-BA and controls (p < 0.0001). Furthermore, we found no causal relationship of high expression of CD15 in FRVs to coexisting placental conditions, including severity and mechanisms/patterns of placental injury, fetal erythroblastosis, and maternal conditions. According to a multivariate analysis, only a high expression of CD15 in FRVs was independently associated with severe non-acute fetal hypoxia ([OR] = 15.52; 95% [CI] = 5.92-40.67). DISCUSSION: We have defined a characteristic pattern of CD15 expression in FRVs that allows to interpret various clinical/placental conditions with respect to fetal hypoxia, with an improved predictability compared to conventional analyses. This approach represents a novel diagnostic strategy for placenta examination, which could indirectly assess severity and duration of intrauterine hypoxia in a heterogeneous population of newborns.

Cinnamaldehyde mitigates placental vascular dysfunction of gestational diabetes and protects from the associated fetal hypoxia by modulating placental angiogenesis, metabolic activity and oxidative stress.

Gestational diabetes mellitus (GDM) is a major pregnancy-related disorder with an increasing prevalence worldwide. GDM is associated with altered placental vascular functions and has severe consequences for fetal growth. There is no commonly accepted medication for GDM due to safety considerations. Actions of the currently limited therapeutic options focus exclusively on lowering the blood glucose level without paying attention to the altered placental vascular reactivity and remodelling. We used the fat-sucrose diet/streptozotocin (FSD/STZ) rat model of GDM to explore the efficacy of cinnamaldehyde (Ci; 20 mg/kg/day), a promising antidiabetic agent for GDM, and glyburide/metformin-HCl (Gly/Met; 0.6 + 100 mg/kg/day), as a reference drug for treatment of GDM, on the placenta structure and function at term pregnancy after their oral intake one week before mating onward. Through genome-wide transcriptome, biochemical, metabolome, metal analysis and histopathology we obtained an integrated understanding of their effects. GDM resulted in maternal and fetal hyperglycemia, fetal hyperinsulinemia and placental dysfunction with subsequent fetal anemia, hepatic iron deficiency and high serum erythropoietin level, reflecting fetal hypoxia. Differentially-regulated genes were overrepresented for pathways of angiogenesis, metabolic transporters and oxidative stress. Despite Ci and Gly/Met effectively alleviated the maternal and fetal glycemia, only Ci offered substantial protection from GDM-associated placental vasculopathy and prevented the fetal hypoxia. This was explained by Ci's impact on the molecular regulation of placental angiogenesis, metabolic activity and redox signaling. In conclusion, Ci provides a dual impact for the treatment of GDM at both maternal and fetal levels through its antidiabetic effect and the direct placental vasoprotective action. Lack of Gly/Met effectiveness to restore it's impaired functionality demonstrates the vital role of the placenta in developing efficient medications for GDM.

Hypoxia-induced inhibition of mTORC1 activity in the developing lung: a possible mechanism for the developmental programming of pulmonary hypertension.

Perinatal hypoxia induces permanent structural and functional changes in the lung and its pulmonary circulation that are associated with the development of pulmonary hypertension (PH) in later life. The mechanistic target of the rapamycin (mTOR) pathway is vital for fetal lung development and is implicated in hypoxia-associated PH, yet its involvement in the developmental programming of PH remains unclear. Pregnant C57/BL6 dams were placed in hyperbaric (760 mmHg) or hypobaric chambers during gestation (505 mmHg, day 15 through postnatal day 4) or from weaning through adulthood (420 mmHg, postnatal day 21 through 8 wk). Pulmonary hemodynamics and right ventricular systolic pressure (RVSP) were measured at 8 wk. mTOR pathway proteins were assessed in fetal (day 18.5) and adult lung (8 wk). Perinatal hypoxia induced PH during adulthood, even in the absence of a sustained secondary hypoxic exposure, as indicated by reduced pulmonary artery acceleration time (PAAT) and peak flow velocity through the pulmonary valve, as well as greater RVSP, right ventricular (RV) wall thickness, and RV/left ventricular (LV) weight. Such effects were independent of increased blood viscosity. In fetal lung homogenates, hypoxia reduced the expression of critical downstream mTOR targets, most prominently total and phosphorylated translation repressor protein (4EBP1), as well as vascular endothelial growth factor, a central regulator of angiogenesis in the fetal lung. In contrast, adult offspring of hypoxic dams tended to have elevated p4EBP1 compared with controls. Our data suggest that inhibition of mTORC1 activity in the fetal lung as a result of gestational hypoxia may interrupt pulmonary vascular development and thereby contribute to the developmental programming of PH.NEW & NOTEWORTHY We describe the first study to evaluate a role for the mTOR pathway in the developmental programming of pulmonary hypertension. Our findings suggest that gestational hypoxia impairs mTORC1 activation in the fetal lung and may impede pulmonary vascular development, setting the stage for pulmonary vascular disease in later life.

Hypoxia in utero increases the risk of pulmonary hypertension in rat offspring and is associated with vasopressin type­2 receptor upregulation.

Pulmonary hypertension (PH) in newborns and adults is a disease that can lead to right heart failure and result in a shorter lifespan. PH was induced by maintaining pregnant rats in a hypoxic chamber for 4 h twice a day, from days 7­21 of pregnancy. Hypoxia was confirmed by a decrease in the partial pressure of oxygen (PaO2) and the oxygen saturation (SaO2) of arterial blood in the aorta. The body weight of newborns from hypoxic rats was ~20% decreased compared with the control newborns of normoxic rats. The vascular wall thickness/vascular diameter values of hypoxia treated pubs were increased compared with that of control newborns 7 days after birth; however, it decreased to similar levels than in the control group after 3 months, and then further decreased to significantly lower levels than in the control group at 6 months after birth. At birth, the lung tissues of newborns from hypoxic rats exhibited an increase in the levels of mRNA and proteins associated with PH such as HIF­1α, HIF­2α, V2R, TGF­ß, TNF­α, Ang­2 and α­SMA. At 3 and 6 months after birth, the levels of both V2R mRNA and protein in offspring from hypoxic rats were at least 2­fold higher, whereas the expression of all other factors decreased compared with the control offspring. By contrast, HIF­2α and Ang­2 expression levels were significantly increased in the 6­month­old control offspring from normoxic rats. V2R overexpression in pups induced by hypoxia in maternal rats was sustained until their adulthood. V2R may be a marker for detecting PH.

Fetal Hypoxia Impacts on Proliferation and Differentiation of Sca-1+ Cardiac Progenitor Cells and Maturation of Cardiomyocytes: A Role of MicroRNA-210.

Hypoxia is one of the most frequent and severe stresses to an organism's homeostatic mechanisms, and hypoxia during gestation has profound adverse effects on the heart development increasing the occurrence of congenital heart defects (CHDs). Cardiac progenitor cells (CPCs) are responsible for early heart development and the later occurrence of heart disease. However, the mechanism of how hypoxic stress affects CPC fate decisions and contributes to CHDs remains a topic of debate. Here we examined the effect of hypoxic stress on the regulations of CPC fate decisions and the potential mechanism. We found that experimental induction of hypoxic responses compromised CPC function by regulating CPC proliferation and differentiation and restraining cardiomyocyte maturation. In addition, echocardiography indicated that fetal hypoxia reduced interventricular septum thickness at diastole and the ejection time, but increased the heart rate, in mouse young adult offspring with a gender-related difference. Further study revealed that hypoxia upregulated microRNA-210 expression in Sca-1+ CPCs and impeded the cell differentiation. Blockage of microRNA-210 with LNA-anti-microRNA-210 significantly promoted differentiation of Sca-1+ CPCs into cardiomyocytes. Thus, the present findings provide clear evidence that hypoxia alters CPC fate decisions and reveal a novel mechanism of microRNA-210 in the hypoxic effect, raising the possibility of microRNA-210 as a potential therapeutic target for heart disease.

Impact of Chronic Fetal Hypoxia and Inflammation on Cardiac Pacemaker Cell Development.

Chronic fetal hypoxia and infection are examples of adverse conditions during complicated pregnancy, which impact cardiac myogenesis and increase the lifetime risk of heart disease. However, the effects that chronic hypoxic or inflammatory environments exert on cardiac pacemaker cells are poorly understood. Here, we review the current evidence and novel avenues of bench-to-bed research in this field of perinatal cardiogenesis as well as its translational significance for early detection of future risk for cardiovascular disease.

Oscillations and concentration dynamics of brain tissue oxygen in neonates and adults.

The brain is a metabolically demanding organ and its health directly depends on brain oxygen dynamics to prevent hypoxia and ischemia. Localized brain tissue oxygen is characterized by a baseline level combined with spontaneous oscillations. These oscillations are attributed to spontaneous changes of vascular tone at the level of arterioles and their frequencies depend on age. Specifically, lower frequencies are more typical for neonates than for adults. We have built a mathematical model which analyses the diffusion abilities of oxygen based on the frequency of source brain oxygen oscillations and neuronal demand. We have found that a lower frequency of spontaneous oscillations of localized brain tissue oxygen can support higher amplitudes of oxygen concentration at areas distant from a source relative to oscillations at higher frequencies. Since hypoxia and ischemia are very common events during early development and the neurovascular unit is underdeveloped in neonates, our results indicate that lower frequency oxygen oscillations can represent an effective passive method of neonatal brain protection against hypoxia. These results can have a potential impact on future studies aiming to find new treatment strategies for brain ischemia.

Evidence of increased hypoxia signaling in fetal liver from maternal nutrient restriction in mice.

BACKGROUND: Intrauterine growth restriction (IUGR) is a pregnancy condition where fetal growth is reduced, and offspring from IUGR pregnancies are at increased risk for type II diabetes as adults. The liver is susceptible to fetal undernutrition experienced by IUGR infants and animal models of growth restriction. This study aimed to examine hepatic expression changes in a maternal nutrient restriction (MNR) mouse model of IUGR to understand fetal adaptations that influence adult metabolism. METHODS: Liver samples of male offspring from MNR (70% of ad libitum starting at E6.5) or control pregnancies were obtained at E18.5 and differential expression was assessed by RNAseq and western blots. RESULTS: Forty-nine differentially expressed (FDR < 0.1) transcripts were enriched in hypoxia-inducible pathways including Fkbp5 (1.6-fold change), Ccng2 (1.5-fold change), Pfkfb3 (1.5-fold change), Kdm3a (1.2-fold change), Btg2 (1.6-fold change), Vhl (1.3-fold change), and Hif-3a (1.3-fold change) (FDR < 0.1). Fkbp5, Pfkfb3, Kdm3a, and Hif-3a were confirmed by qPCR, but only HIF-2a (2.2-fold change, p = 0.002) and HIF-3a (1.3 p = 0.03) protein were significantly increased. CONCLUSION: Although a moderate impact, these data support evidence of fetal adaptation to reduced nutrients by increased hypoxia signaling in the liver.

Multi-Omics Integration Reveals Short and Long-Term Effects of Gestational Hypoxia on the Heart Development.

Antenatal hypoxia caused epigenetic reprogramming of methylome and transcriptome in the developing heart and increased the risk of heart disease later in life. Herein, we investigated the impact of gestational hypoxia in proteome and metabolome in the hearts of fetus and adult offspring. Pregnant rats were treated with normoxia or hypoxia (10.5% O2) from day 15 to 21 of gestation. Hearts were isolated from near-term fetuses and 5 month-old offspring, and proteomics and metabolomics profiling was determined. The data demonstrated that antenatal hypoxia altered proteomics and metabolomics profiling in the heart, impacting energy metabolism, lipid metabolism, oxidative stress, and inflammation-related pathways in a developmental and sex dependent manner. Of importance, integrating multi-omics data of transcriptomics, proteomics, and metabolomics profiling revealed reprogramming of the mitochondrion, especially in two clusters: (a) the cluster associated with "mitochondrial translation"/"aminoacyl t-RNA biosynthesis"/"one-carbon pool of folate"/"DNA methylation"; and (b) the cluster with "mitochondrion"/"TCA cycle and respiratory electron transfer"/"acyl-CoA dehydrogenase"/"oxidative phosphorylation"/"complex I"/"troponin myosin cardiac complex". Our study provides a powerful means of multi-omics data integration and reveals new insights into phenotypic reprogramming of the mitochondrion in the developing heart by fetal hypoxia, contributing to an increase in the heart vulnerability to disease later in life.