Normal Pregnant Rats Research Articles

Microbial reconstitution reverses prenatal stress-induced cognitive impairment and synaptic deficits in rat offspring

Stress during pregnancy is often linked with increased incidents of neurodevelopmental disorders, including cognitive impairment. Here, we report that stress during pregnancy leads to alterations in the intestinal flora, which negatively affects the cognitive function of offspring. Cognitive impairment in stressed offspring can be reproduced by transplantation of cecal contents of stressed pregnant rats (ST) to normal pregnant rats. In addition, gut microbial dysbiosis results in an increase of β-guanidinopropionic acid in the blood, which leads to an activation of the adenosine monophosphate-activated protein kinase (AMPK) and signal transducer and activator of transcription 3 (STAT3) in the fetal brain. Moreover, β-guanidinopropionic acid supplementation in pregnant rats can reproduce pregnancy stress-induced enhanced glial differentiation of the fetal brain, resulting in impaired neural development. Using probiotics to reconstruct maternal microbiota can correct the cognitive impairment in the offspring of pregnant stressed rats. These findings suggest that microbial reconstitution reverses gestational stress-induced cognitive impairment and synaptic deficits in male rat offspring.

Computational model captures cardiac growth in hypertensive pregnancies and in the postpartum period.

Heart growth in the pregnant patient helps maintain cardiovascular function while supporting the growing fetus. However, in some cases, the cardiovascular demand of pregnancy can trigger life-threatening conditions, including hypertensive disorders of pregnancy and peripartum cardiomyopathy. The mechanisms that control heart growth throughout pregnancy are unclear, and treating these diseases remains elusive. We previously developed a computational model that accounts for hormonal and hemodynamic interactions throughout pregnancy and demonstrated its ability to capture realistic cardiac growth in normal rat pregnancy. In this study, we evaluated whether this model could capture heart growth beyond normal pregnancy. After further validation of our normal pregnancy predictions, we tested our model predictions of three rat studies of hypertensive pregnancies. We next simulated the postpartum period and examined the impact of lactation on cardiac growth in rats. We demonstrate that our multiscale model can capture cardiac growth associated with new-onset hypertension during pregnancy and lactation status in the postpartum period. We conclude by elaborating on the potential clinical utility of our model in the future.NEW & NOTEWORTHY Our multiscale model predicts appropriate heart growth beyond normal pregnancy, including elevated heart weights in rats with induced hypertension during pregnancy and in lactating mice and decreased heart weight in nonlactating mice. Our model captures distinct mechanisms that result in similar organ-level growth, highlighting its potential to distinguish healthy from diseased pregnancy-induced growth.

Increased uterine arterial tone, stiffness and remodeling with augmented matrix metalloproteinase-1 and -7 in uteroplacental ischemia-induced hypertensive pregnancy

Preeclampsia is a pregnancy-related disorder manifested as hypertensive pregnancy (HTN-Preg) and often fetal growth restriction (FGR), but the mechanisms involved are unclear. We have reported enhanced reactivity of systemic vessels in HTN-Preg rats, but the critical changes in the uterine circulation are less clear. We tested whether HTN-Preg involves localized aberrations in uterine arterial tone, stiffness and remodeling by matrix metalloproteinases (MMPs). Blood pressure (BP) and litter size were recorded in normal pregnant (Preg) rats and Preg rats with reduced uteroplacental perfusion pressure (RUPP). Isolated uterine arteries were placed in a pressure myograph for measuring intrinsic and extrinsic tone and arterial stiffness. Arteries were bathed in normal Krebs solution (2.5 mM Ca2+), Ca2+-free (2 mM EGTA) Krebs, treated with sodium nitroprusside (SNP), or endothelium denuded, then pressurized at 10 mmHg steps from 10 to 110 mmHg, and the % change in diameter was analyzed to measure total (active + passive), active Ca2+-dependent myogenic, passive, and endothelium-dependent tone, respectively. BP was higher and the litter size and pup weight were reduced in RUPP vs Preg rats. In normal Krebs, increasing intraluminal pressure caused smaller increments in diameter in arteries of RUPP vs Preg rats, suggesting greater total vascular tone. Arterial incubation in Ca2+-free Krebs, treatment with SNP or endothelium-removal abolished the differences in vascular tone, and subtraction of each of these components from total vascular tone revealed significant active Ca2+-dependent myogenic, passive, and endothelium-dependent tone, respectively, in RUPP vs Preg rats. The total and passive strain–stress curves were shifted leftward in arteries of RUPP vs Preg rats, indicating increased uterine arterial stiffness. Arterial sections showed decreased lumen/total and increased wall/total area, and immunohistochemistry revealed greater MMP-1 and MMP-7 staining particularly in the media, suggesting uterine arterial remodeling by MMPs in RUPP vs Preg rats. The increased uterine arterial active myogenic, passive, and endothelium-dependent tone, arterial stiffness and remodeling by MMPs would further reduce uterine blood flow and exacerbate uteroplacental ischemia, FGR and HTN-Preg.

IL-33 supplementation improves uterine artery resistance and maternal hypertension in response to placental ischemia.

Preeclampsia (PE), a leading cause of maternal/fetal morbidity and mortality, is a hypertensive pregnancy disorder with end-organ damage that manifests after 20 wk of gestation. PE is characterized by chronic immune activation and endothelial dysfunction. Clinical studies report reduced IL-33 signaling in PE. We use the Reduced Uterine Perfusion Pressure (RUPP) rat model, which mimics many PE characteristics including reduced IL-33, to identify mechanisms mediating PE pathophysiology. We hypothesized that IL-33 supplementation would improve blood pressure (BP), inflammation, and oxidative stress (ROS) during placental ischemia. We implanted intraperitoneal mini-osmotic pumps infusing recombinant rat IL-33 (1 µg/kg/day) into normal pregnant (NP) and RUPP rats from gestation day 14 to 19. We found that IL-33 supplementation in RUPP rats reduces maternal blood pressure and improves the uterine artery resistance index (UARI). In addition to physiological improvements, we found decreased circulating and placental cytolytic Natural Killer cells (cNKs) and decreased circulating, placental, and renal TH17s in IL-33-treated RUPP rats. cNK cell cytotoxic activity also decreased in IL-33-supplemented RUPP rats. Furthermore, renal ROS and placental preproendothelin-1 (PPET-1) decreased in RUPP rats treated with IL-33. These findings demonstrate a role for IL-33 in controlling vascular function and maternal BP during pregnancy by decreasing inflammation, renal ROS, and PPET-1 expression. These data suggest that IL-33 may have therapeutic potential in managing PE.NEW & NOTEWORTHY Though decreased IL-33 signaling has been clinically associated with PE, the mechanisms linking this signaling pathway to overall disease pathophysiology are not well understood. This study provides compelling evidence that mechanistically links reduced IL-33 with the inflammatory response and vascular dysfunction observed in response to placental ischemia, such as in PE. Data presented in this study submit the IL-33 signaling pathway as a possible therapeutic target for the treatment of PE.

Cardiovascular effect of preeclampsia upon offspring development: Are (Pro) renin-renin receptor ((P)RR) and gender related?

Preeclampsia (PE) is a complication of pregnancy that might increase progeny risk of cardiovascular and metabolic problems, mainly in males. Renin angiotensin aldosterone system is known to be involved. (Pro) renin/renin receptor ((P)RR) has been shown to participate in cardiovascular pathology. The aim of this work was to evaluate (P)RR expression and function upon cardiovascular and renal tissues from PE dams' offspring. We used offspring from normal pregnant and preeclamptic rats, evaluating body, heart, aorta and kidney weight, length, and blood pressure along 3 months after birth. Subsets of animals received handle region peptide (HRP) (0.2 mg/Kg, sc). Another group received vehicle. Animals were sacrificed at first, second, and third months of age, tissues were extracted and processed for immunoblot to detect (P)RR, PLZF, β-catenin, DVL-1, and PKCα. (P)RR and PLZF were also measured by RT-PCR. We found that offspring developed hypertension. Male descendants remained hypertensive throughout the whole experiment. Female animals tended to recover at second month and returned to normal blood pressure at third month. HRP treatment diminished hypertension in both male and female animals. Morphological evaluations showed changes in heart, aorta, and kidney weight, and HRP reverted this effect. Finally, we found that (P)RR, PLZF, and canonical WNT transduction pathway molecules were stimulated by PE, and HRP treatment abolished this increase. These findings suggest that PE can induce hypertension in offspring, and (P)RR seems to play an important role through the canonical WNT pathway and that gender seems to influence this response.

Progesterone-induced blocking factor blockade causes hypertension in pregnant rats.

Preeclampsia (PE) is a multisystem disorder characterized by new onset hypertension in mid-late gestation and can include multi-organ dysfunction with or without proteinuria. It affects 5%-7% of all pregnancies in the U.S., making PE a major contributor to maternal and fetal morbidity and mortality. Currently, there is no cure for this pregnancy complication except for early delivery of the placenta and fetus. Moreover, the therapeutic options to treat PE are very limited. One potential trigger for the development of PE is progesterone deficiency-induced imbalance between T Helper 1(Th1)/Th2 cells, an increase in cytolytic natural killer (NK) cells and inflammatory cytokines that in turn leads to endothelial dysfunction, intrauterine growth restriction (IUGR) and hypertension. Importantly, progesterone signals the synthesis of progesterone-induced blocking factor (PIBF) which has anti-inflammatory effects and could promote the regulation of inflammation balance during pregnancy. However, the role of progesterone and PIBF in the pathophysiology of PE is still not fully understood. Thus, this current study was designed to test the hypothesis that inhibition of PIBF causes signs of PE in pregnant Sprague Dawley rats. In order to address our hypothesis, rabbit anti-PIBF IgG (0.25, low dose-LD or 0.50mg/mL, high dose-HD) was administered intraperitoneally on gestation day (GD) 15 to normal pregnant Sprague Dawley (NP) rats. On GD 18, carotid catheters were inserted and on GD 19 mean blood pressure (MAP) and samples were collected for further analysis. MAP in normal pregnant rats (NP) rats (n=7) was 99±3mmHg, which increased to 116±2 mmHg in NP+ anti-PIBF LD (n=10) and 113±4mmHg in NP+ anti-PIBF HD (n=4), p<0.05. Plasma TNF-alpha levels were 35±8 pg/mL in NP rats and increased to 84±21 pg/mL in NP+ Anti-PIBF HD (n=4), p<0.05. Plasma IL-4 and IL-10 levels were 22±5 and 25+6 pg/mL in NP (n=5), which decreased to 6±1 and 8±1 pg/mL in NP+ Anti-PIBF LD (n=6, p<0.05) and 16±4 and 15±5 pg/mL in NP+ Anti-PIBF HD (n=4). Circulating total NK cells were 67±11 % gatein NP rats (n=3), which decreased to 28±7% gate in NP+ Anti-PIBF LD and 45±6% gate in NP+ Anti-PIBF HD. Cytolytic NK cells were increased in NP+ Anti-PIBF HD, p<0.05. Moreover, circulating NO levels were significantly decreased while renal cortex PPET-1 levels increased NP+ Anti-PIBF HD. Our study demonstrates that PIBF blockade causes hypertension, inflammation and signs of endothelial dysfunction, all of which are associated with PE, thus indicating the importance of progesterone signalling pathways during a healthy pregnancy.

Microglia are necessary for probiotics supplementation to improve impaired fear extinction caused by pregnancy stress in adult offspring of rats

The prevention and treatment of fear-related disorders in offspring affected by pregnancy stress remains challenging at clinic. Here, we examined the effects of gut microbiota of stressed pregnant rats on the fear extinction of their offsprings, and the potential mechanisms. We found that gut microbiota transplantation from rats with pregnancy stress to normal pregnant rats impaired fear extinction, induced microglial activation and synaptic phagocytosis, increased synapse loss in offsprings. Probiotics supplement during pregnancy stress partly normalized pregnancy stress-induced gut microbiota dysbiosis of pregnant rats, and promoted fear memory extinction, inhibited fear memory reappearance, and limited microglial activation and synaptic phagocytosis in offsprings. These data revealed that gut microbiota of stressed pregnant mother improved the development of fear-related disorders of offspring, which may be associated with microglial synaptic pruning.

Abstract P426: Novel Peptide B7-33 Demonstrates Efficacy In Rat Models Of Preeclampsia

Background: Preeclampsia (PreE) is a hypertensive pregnancy disorder, which occurs in approximately 10% of all gestations. The literature suggests potential therapeutic role of H2 relaxin in PreE. Due to the complex insulin-like structure of relaxin (A- and B- chains, 53 amino acids, 3 disulfide bonds), a novel H2 relaxin B-chain-only peptide variant B7-33 (27 amino acids without any disulfide bonds) has recently been developed. Objective: This single-chain peptide displayed equivalent efficacy to the natural H2 relaxin in several functional assays both in vitro and in vivo . The Aim of this study is to evaluate whether B7-33 attenuates preE syndrome in rat models of PreE. Methods: Rat Model 1. The efficacy of B7-33 was evaluated in the Reduction of Uterine Perfusion Pressure (RUPP) model as described previously. RUPP rats are randomly assigned to 4 groups (N=8/group): 1) vehicle, 2) B733; 3) B733-Fc; and 4) B733-HSA. Rats are dosed twice weekly (i.v.) from GD10 to GD 20. Rat Model 2. Timed-pregnant rats are used, and the model is conducted as previously described. DOCA and saline administration begin at approximately GD2. Three groups of animals are studied (n=8 per group). 1) normal pregnant rats; 2) pregnant animals injected (i.p.) initially with 12.5 mg of DOCA, followed by a weekly injection of 6.5 mg, and whose drinking water was replaced with 0.9% saline; 3) rats administered DOCA and saline as for Group 2, and given B733 35 ug i.p. biweekly from GD10-20. BP, proteinuria and inflammatory markers were evaluated. Statistical comparisons were performed using analysis of variance with Duncan’s post hoc test. Results: RUPP rats have increased MAP, plasma TNF-α, and plasma sFlt-1 along with decreased NO index compared to normal pregnancy. Treatment with B733 and B7-33 fusion proteins lowers MAP, TNF-α, and sFlt-1 back to that of normal pregnancy. B7-33 data are consistent with earlier data with serelaxin in the RUPP model. All fusions tested ameliorate hypertension in the RUPP animals. B7-33 normalizes BP and proteinuria in the DOCA rat model of preE. Conclusion: Both B7-33 and B7-33 fusion proteins attenuate preE syndrome in RUPP and DOCA rat models of PreE. We conclude that B7-33 is an ideal candidate for development as a novel therapeutic in preE.

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 &lt;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 &lt;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 066: Decoy Peptide Attenuates The Syndrome Of preeclampsia In A Rodent Model

Background: Preeclampsia (preE) is a serious complication of pregnancy manifested by high blood pressure, proteinuria, and edema. In a previous study, we demonstrated that circulating levels of soluble (pro)renin receptor ((P)RR) were elevated at delivery in patients with preE, and that both plasma and placental (pro)renin were elevated in preE patients and in a rat model of preE. Decapeptide based on this handle-region sequence (handle-region peptides or HRP) can block binding of (pro)renin to (P)RR. Objective: The goal of this study is to develop and characterize HRP as an innovative treatment for preE. Methods and Materials: The human extravillous cytotrophoblast (CTB) cell line Sw-71 used in these studies was derived from first trimester chorionic villous tissue. (P)RR (2 nM) was incubated in 1.0 mL medium with the receptor-expressing CTB cells for 0, 6, 12, 18, or 24 hours, followed by renin activity assay to determine the percent binding and activation of (P)RR. Female Sprague-Dawley rats (200-250 g, first-time pregnant with confirmation by vaginal plug, Harlan) are randomly assigned to three groups (n=8 per group): 1) NP: normal pregnant rats; 2) PDS: pregnant animals injected initially with 12.5 mg of DOCA in a depot form intraperitoneally, followed by a weekly injection of 6.5 mg, and whose drinking water was replaced with 0.9% saline; 3) PDS-HRP: rats administered DOCA and saline as for Group 2, and also given HRP (10 mg/kg, i.p.) daily from day 10 through day 20 of pregnancy. Results: (Pro)renin was non-proteolytically activated by binding to (P)RR on the cell membrane of CTB cells, while HRP inhibited binding and attenuated activation of (P)RR. The HRP decoy peptide normalizes blood pressure, proteinuria, and birth numbers in DOCA model of preE. When HRP was administered following the onset of hypertension, the BP, proteinuria, and number of pups were normalized to control normal pregnancy. Conclusions: HRP inhibits the binding of (pro)renin to (P)RR. This study in animal model of preE suggests HRP blockade of (P)RR can attenuate the disease. (P)RR and (pro)renin levels are elevated in preE patients and in their placenta, as well as in a rat model of preE. This data suggests HRP as a potential therapeutic for intervention of RAS signaling in preE.

Abstract 022: Il-33 Supplementation Improves Vascular Function And Maternal Hypertension In Response To Placental Ischemia

Preeclampsia (PE), a leading cause of maternal/fetal morbidity and mortality, is a hypertensive pregnancy disorder with end-organ damage that manifests after 20 weeks of gestation. PE is characterized by chronic immune activation and endothelial dysfunction. Clinical studies report reduced IL-33 signaling in PE. We use the R educed U terine P erfusion P ressure (RUPP) rat model, which mimics many PE characteristics including reduced IL-33, to identify mechanisms mediating PE pathophysiology. We hypothesized IL-33 supplementation would improve blood pressure (BP), inflammation, and oxidative stress (ROS) during placental ischemia. We implanted i.p. mini-osmotic pumps infusing recombinant rat IL-33 (1 μg/kg/day) into normal pregnant (NP) and RUPP rats from gestation day 14-19. We assessed maternal BP, cytolytic NK (cNK) and T H 17 cells, cNK cell activity, ROS, preproendothelin-1 (PPET) expression, and uterine artery resistance index (UARI). MAP (mmHg) was 104±4 in NP vs 124±7 in RUPP (p&lt;0.05 vs NP) and lowered to 108±10 in RUPP+IL-33 (p&lt;0.05 vs RUPP, n=8/group). Circulating cNKs (% gated) were 0.9±0.2% in NP vs 3.9±1.1% in RUPP (p&lt;0.05 vs NP) and decreased to 1.1±0.6% in RUPP+IL-33 (p&lt;0.05 vs RUPP, n=8/group). Circulating T H 17s (%CD4 + T cells) were 1.0±0.6% in NP vs 7.2±0.5% in RUPP (p&lt;0.05 vs NP) and 0.8±0.4% in RUPP+IL-33 (p&lt;0.05 vs RUPP, n=8/group). Renal T H 17s, placenta cNKs and T H 17s followed similar trends. cNK cell activity (fold change) was 3.92±0.6 fold higher in RUPP vs NP (p&lt;0.05 vs NP) and decreased to -0.24±0.1 in RUPP+IL-33 (p&lt;0.05 vs RUPP, n=3/group). Renal ROS (relative light units/min/mg protein) was 452±33 in NP vs 676±44 in RUPP (p&lt;0.05) and decreased to 258±50 in RUPP+IL-33 (p&lt;0.05 vs RUPP, n=5/group). Placental PPET (fold change) was 2.36±0.4 fold higher in RUPP vs NP (p&lt;0.05) and decreased to 1.22±0.2 in RUPP+IL-33 (p=0.05 vs RUPP, n=6-7/group). UARI (arbitrary units) was 0.71±0.01 in RUPP vs 0.54±0.01 in NP (p&lt;0.05) and was lowered to 0.57±0.02 in RUPP+IL-33 (p&lt;0.05 vs RUPP, n=6/group). These findings demonstrate a role for IL-33 in controlling vascular function and maternal BP during pregnancy by decreasing inflammation, ROS, and PPET expression. These data suggest IL-33 may have therapeutic potential in managing PE.

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 &lt; 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 &lt; 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 &lt; 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 &lt; 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.

Abstract P422: CD4+ T Cells From Preeclamptic Patients During Pregnancy Causes Cognitive Dysfunction Postpartum In A Pregnant Rat Model

Preeclampsia (PE), new onset hypertension (HTN) during pregnancy, is associated with impaired cognitive function and increased cerebral blood flow (CBF). Although we have previously shown adoptive transfer of placental CD4+ T cells from PE women into athymic nude pregnant rats causes HTN and circulating factors associated with PE, we do not know the role of CD4+ T cells in contributing to impaired cerebral blood flow and cognitive dysfunction in the postpartum period. Therefore, we hypothesize that adoptive transfer of placental CD4+ T cells from PE patients into pregnant athymic nude recipient rats will cause maternal hypertension and cognitive dysfunction and impaired CBF postpartum. Placental CD4+ T cells were isolated from normotensive (NP) and preeclamptic (PE) pregnant women at delivery. One million CD4+ T cells were injected interperitoneally (i.p.) into normal pregnant nude athymic rats on gestational day (GD) 12. Dams were allowed to deliver and aged 12-weeks after delivery. Blood pressure (MAP) was measured by carotid catheter and at 12-weeks postpartum CBF autoregulation was measured by laser Doppler flowmetry. The Barnes maze and the novel object recognition behavioral assays were used to assess cognitive function on gestational day (GD) 15-19 and 12-weeks postpartum. A student’s t-test was used for statistical analysis. MAP increased during pregnancy in PE recipient rats (115±2 mmHg, n=5) compared to NP recipient rats (99±3 mmHg, n=4, p&lt; 0.05). MAP significantly increased in the postpartum period in PE recipient rats (114±2 mmHg, n=5) compared to NP controls (98±2 mmHg, n=7, p&lt;0.05). PE recipient rats exhibited latency with errors navigating in the Barnes maze compared to NP controls during pregnancy and postpartum. Locomotor activity was decreased in PE recipient rats compared to NP controls during pregnancy (p&lt;0.05) and postpartum. PE recipient rats had increased CBF autoregulation compared to NP rat recipients in the postpartum period. Our findings indicate that rat recipients of CD4+ T cells from PE patients demonstrate maternal HTN and increased CBF and cognitive dysfunction in the postpartum period compared to recipients of NP CD4 + T cells. These data indicate long-term cognitive and cerebral dysfunction as a result of preeclampsia.

RUPP Th17s cause hypertension and mitochondrial dysfunction in the kidney and placenta during pregnancy.

Preeclampsia (PE), new-onset hypertension (HTN), and organ dysfunction during the second half of pregnancy, is associated with an increase in inflammatory immune cells, including T helper 17 (Th17) cells. Studies have demonstrated that mitochondrial (mt) dysfunction is important in the pathogenesis of PE though causative factors have yet to be fully identified. Although Th17 cells, natural killer (NK) cells, and mt dysfunction contribute to HTN in the reduced uterine perfusion pressure (RUPP) rat model, the role of Th17 cells or IL-17 in mt dysfunction is unknown. Therefore, we hypothesize that RUPP stimulated Th17 cells cause HTN and mt dysfunction, which is alleviated with the blockade of IL-17. On gestational day 12 (GD12), RUPP Th17 cells were transferred into normal pregnant (NP) Sprague Dawley rats. A subset of NP+RUPPTh17 rats received IL-17RC (100pg/day) on GD14-19. Blood pressure (MAP), NK cells, and mt function were measured on GD19 in all groups. MAP increased in response to NP+RUPP Th17 compared to NP rats and was lowered with IL-17RC. Circulating and placental NK cells increased with NP+RUPP Th17 compared to NP and were lowered with IL-17RC. Renal mtROS increased in NP+RUPP Th17 compared to NP and was normalized with IL-17RC. Similar to PE women, placental mtROS decreased in NP+RUPP Th17 and was normalized with IL-17RC. Our results indicate that IL-17RC inhibition normalizes HTN, NK cell activation, and multi-organ mt dysfunction caused by Th17 cells stimulated in response to placental ischemia.

Adoptive transfer of CD4+ T cells from women with preeclampsia with or without a history of COVID during pregnancy contribute to hypertension, AT1-AA, and neurovascular dysfunction in pregnant rats

Objective: Preeclampsia (PE) is characterized by new onset hypertension during pregnancy and is associated with chronic inflammation and neurovascular dysfunction. The inflammation in PE includes activated CD4+ T cells and B cells producing agonistic antibodies against the angiotensin II type 1 receptor (AT1AA) which has also been shown to be produced in patients with COVID-19 (CV) infection. Interestingly, having had CV during pregnancy increases the incidence of a PE-like phenotype in that pregnancy. Both PE and CV have long lasting neurological implications and we have shown that AT1AA mediates neurovascular dysfunction in PE. Moreover, we have shown that CD4+ T cells from women with PE induce a PE syndrome in pregnant athymic nude recipient rats. Although CD4+T cells have been shown to contribute to neurovascular dysfunction, we do not know the role of CD4+ T cells in contributing to neurovascular dysfunction in PE or in patients with a history (Hx) CV during pregnancy. Therefore, we sought examine a role for CD4+ T cells from PE with or without a CV Hx to contribute to AT1AA in pregnancy as well as hypertension and cerebrovascular dysfunction postpartum. We utilized adoptive transfer of placental CD4+ T cells from PE and normotensive (NT) patients with or without a CV Hx during pregnancy into athymic nude recipient rats and determined the presence of a PE phenotype during pregnancy or postpartum. Methods: CD4+ T cells were isolated from placentas of normal pregnant (NP), PE, CV Hx normotensive (CV Hx NT), and Hx of CV with PE (CV Hx PE) at delivery. One million CD4+ T cells were injected i.p. into gestational day (GD) 12 nude athymic rats. At GD19 blood samples were collected to measure AT1AA by cardiomyocyte assay. At three months postpartum, MAP and CBF were measured by arterial catheter and laser Doppler flowmetry. A two-way ANOVA was used for statistical analysis. Results: In pregnant recipient rats, AT1AA was increased in PE (8±2 ΔBPM, n=4) and CV Hx PE (4±2 ΔBPM, n=4) compared to NP (-3±2 ΔBPM, n=4, p&lt;0.05) and CV Hx NT (2±2 ΔBPM, n=2) respectively. At 3 months postpartum, MAP was increased in PE (115±2 mmHg, n=5) and CV Hx PE (111±4, n=4) compared to NP (99±3 mmHg, n=4 p&lt;0.05) and CV Hx NT (100±4, n=5). PE and CV Hx PE had impaired autoregulation of CBF compared to CV Hx NT and NP rats. Conclusion: Our findings indicate that CD4+ T cells from PE or CV HX PE patients cause AT1-AA in pregnancy as well as hypertension and impaired CBF 3 months postpartum. Collectively, the data demonstrates that CD4+ T cells cause hypertension and inflammation in PE or CV Hx PE and may contribute to neurovascular dysfunction in patients with previous PE or CV Hx PE during pregnancy. RO1HD067541-06, P20GM121334, and AHA 19CDA34670055 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The protective effect of magnesium sulfate on placental inflammation via suppressing the NF-κB pathway in a preeclampsia-like rat model.

Abnormal placental inflammation has a role in the pathophysiology of preeclampsia. Magnesium sulfate (MgSO4) has anti-inflammatory properties and is a fetal neuroprotective agent. MgSO4 is often used to treat severe preeclampsia; however, the specificmechanisms of action underlyingthistherapeutic effect remain unclear. The objective of this study was to investigate the effects of MgSO4 (270mg/kg) on placental inflammation in a rat model of lipopolysaccharide (LPS; 1.0µg/kg)-induced preeclampsia. Compared to normal pregnant rats, LPS-treated pregnant rats had higher blood pressure, proteinuria, and expression of the anti-angiogenic factor sFlt-1 and the pro-inflammatory factors interleukin-1β (IL-1β) and IL-12 in placental tissue. LPS-treated pregnant rats had placental insufficiency, poor fetal outcomes, and significantly decreased expression of the anti-inflammatory factors apolipoprotein E (APOE) and IL-10 in placental tissue. MgSO4 treatment had favorable effects on maternal and fetal outcomes. MgSO4 treatment improved placental function by repressing an exaggerated inflammatory response in the placenta and promoting angiogenesis via the NF-κB pathway. These findings suggest MgSO4 has a potential role in the prevention of preeclampsia and in the treatment of mild and moderate preeclampsia.

NLRP3 inhibition improves maternal hypertension, inflammation, and vascular dysfunction in response to placental ischemia.

Preeclampsia (PE) is a pregnancy-specific hypertensive disorder with end-organ damage that presents after 20 wk of gestation. PE pathophysiology often includes vascular dysfunction and increased inflammation that continues to damage patient health even after PE resolves. Currently, there is no cure for PE beyond delivery of the fetal-placental unit. Previous clinical studies have identified elevated placental NLRP3 expression in patients with PE and suggest NLRP3 as a potential therapeutic target. In this study, we examined the effect of NLRP3 inhibition on PE pathophysiology in the reduced uterine perfusion pressure (RUPP) model rat using MCC950 (20 mg/kg/day) or esomeprazole (3.5 mg/kg/day). We hypothesized that increased NLRP3 in response to placental ischemia impairs anti-inflammatory IL-33 signaling to induce T-helper 17 cell (TH17) and cytolytic NK cell (cNK) activation, which is known to mediate oxidative stress and vascular dysfunction leading to maternal HTN and intrauterine growth restriction. RUPP rats had significantly higher placental NLRP3 expression, maternal blood pressure, fetal reabsorption rate, vascular resistance, oxidative stress, cNKs and TH17s, and decreased IL-33 compared with normal pregnant (NP) rats. NLRP3 inhibition, with either treatment, significantly reduced placental NLRP3 expression, maternal blood pressure, fetal reabsorption rates, vascular resistance, oxidative stress, cNK, and TH17 populations in RUPP rats. Based on our findings, NLRP3 inhibition reduces PE pathophysiology and esomeprazole may be a potential therapeutic for PE treatment.

Effect of Pravastatin on Levels of Malondealdehyde (MDA) And Endothelin-1 (ET-1) Preeclampsia Model Rats

Introduction: Pravastatin is known to have a number of pleiotropic effects including reducing endothelial dysfunction, anti-inflammatory, antioxidants, conangiogenic, and antitrombotic. Pravastatin through the pleitropic effect is expected to be one of the alternative therapies to prevent preeclampsia. The limited strategy for prevention and treatment of preeclampsia is due to the unknown etiology and pathogenesis. These two markers are thought to contribute to the occurrence of preeclampsia although they cause it in two different pathways. MDA is a marker of oxidative stress as an end product of lipid peroxidation. ET-1 is a vasoconstrictor that plays a role in the pathogenesis of preeclampsia through increasing anti-angiogenic properties. Aim: to determine the effect of pravastatin on serum levels of MDA and ET-1 in preeclampsia rat models. Methods: This study consisted of 5 groups; negative control/ K(-) consisted of normal pregnant rats, positive control/ K(+) consisted of rat model of preeclampsia (rat model of preeclampsia induced by administration of L-NAME at a dose of 125 mg/kg BW/day since gestational age 13-19 days), treatment groups 1, 2, and 3 (rat model of preeclampsia given pravastatin with 3 different doses; 2 mg/day (P1), 4 mg/day (P2) and 8 mg/day(P3)) at 13-19 days of gestation. The rat model of preeclampsia was determined based on blood pressure &gt; 140/90 with urine protein &gt; +1. After termination, blood was drawn to measure serum MDA and ET-1 levels. Results: Serum levels of MDA and ET-1 were decreased in groups P2 and P3 compared to groups K(+). Statistically, there was a significant difference in the mean levels of MDA (p=0.001) and ET-1 (p=0.000) between each group. Conclusion: Pravastatin can prevent preeclampsia by decreasing MDA and ET-1.

PS-B12-10: CLASSIC CARDIOVASCULAR PROTECTIVE SIGNALING IN ANGIOTENSIN II-INDUCED NOVEL PREECLAMPSIA RAT MODEL

Angiotensin II (Ang II)-induces vasoconstriction, in vivo perfusion has been shown to induce preeclampsia (PE) in pregnant rats. In this study, we aimed to examine the protein and mRNA expressions of endothelial and neuronal NOS (eNOS, nNOS) as well as angiotensin II type 2 receptor (AT2R) in normal pregnant and Ang II-induced PE rats. Pregnant (8 weeks) and non-pregnant SD rats (8 weeks, GD-8) were infused with Ang II via an osmotic minipump (1 ug/kg/min), and the rats (GD-21) were sacrificed 13 days later. Pregnant and non-pregnant rats without Ang II were taken as sham controls. The fetal weight, crown-rump length and placental weight of PE rats were significantly lower compared to those without Ang II infusion. Immunohistochemistry results showed an approximately 30% increase in glomerular volume, suggesting morphological changes in the kidney and Ang II infusion exhibits a PE phenotype. Ang II infusion did not affect AT2R, AT1R and nNOS protein or mRNA expression in the uterus of non-pregnant or pregnant rats. However, eNOS mRNA expression was reduced in PE rats (p &lt; 0.0001) although protein expression was not affected by Ang II infusion. However, in the placenta, AT1R and AT2R protein expression were not different in PE, but AT2R mRNA was decreased (p &lt; 0.0001). eNOS protein was only expressed in fetal vascular endothelial cells, and eNOS protein expression was higher in PE rats (P = 0.001) but eNOS mRNA was decreased (p = 0.0096). Notably, strong nNOS expression was observed in placental trophoblast cell groups of sham and PE rats, and both mRNA and protein expression of nNOS was decreased in placental lysates of Ang II rats (respectively p = 0.0287 and p &lt; 0.0001). These results demonstrate that we have recapitulated the PE model of Ang II infusion in rats. The expression of nNOS was reduced, and the localization of nNOS in the trophoblast suggested a new functional remodeling in the fetal vasculature. In the placenta, decreased expression of AT2R and nNOS may be a novel mechanism of impaired fetal development.

Adoptive transfer of diabetic CD4+ T-cells into normal pregnant rats induces characteristics of gestational diabetes

Gestational diabetes mellitus (GDM) is characterized by hyperglycemia, β-cell dysfunction, and insulin sensitivity during pregnancy, and is associated with increased T cell activation. We hypothesize, CD4+ T cells play a role in the pathophysiology of GDM by stimulating antibodies leading to β-cell destruction causing glucose intolerance, renal injury and hypertension during pregnancy.