Purpose of reviewEndothelial dysfunction is a major risk factor for many cardiovascular diseases, notably hypertension. Obesity increases the risk of endothelial dysfunction in association with increasing production of the adipokine leptin. Preclinical studies have begun to unravel the mechanisms whereby leptin leads to the development of endothelial dysfunction, which are sex-specific. This review will summarize recent findings of mechanisms of leptin-induced endothelial impairment in both male and females and in pregnancy.Recent findingsLeptin receptors are found in high concentrations in the central nervous system (CNS), via which leptin promotes appetite suppression and upregulates sympathetic nervous system activation. However, leptin receptors are expressed in many other tissues, including the vascular endothelial cells and smooth muscle cells. Recent studies in mice with vascular endothelial or smooth muscle-specific knockdown demonstrate that endothelial leptin receptor activation plays a protective role against endothelial dysfunction in male animals, but not necessarily in females. Clinical studies indicate that women may be more sensitive to obesity-associated vascular endothelial dysfunction. Emerging preclinical data indicates that leptin and progesterone increase aldosterone production and endothelial mineralocorticoid receptor activation, respectively. Furthermore, decades of clinical studies indicate that leptin levels increase in the hypertensive pregnancy disorder preeclampsia, which is characterized by systemic endothelial dysfunction. Leptin infusion in mice induces the clinical characteristics of preeclampsia, including endothelial dysfunction.SummaryNovel preclinical data indicate that the mechanisms whereby leptin promotes endothelial dysfunction are sex-specific. Leptin-induced endothelial dysfunction may also play a role in hypertensive pregnancy as well.

Preeclampsia (PE) is a hypertensive disorder of pregnancy that causes adverse pregnancy outcomes. Progesterone in pregnancy increases endothelial mineralocorticoid receptor (ECMR) expression, which we showed mediates endothelial function in hypertensive female mice, however, whether ablating endothelial dysfunction in PE improves pregnancy outcomes is unknown. We hypothesized that improving endothelial function via ECMR deletion increases fetal growth and ablates hypertension in the reduced uterine perfusion pressure (RUPP) PE mouse. Pregnant ECMR-intact (WT) and -deficient (KO) mice underwent RUPP or sham surgery on GD13 and sacrificed on GD18 (WT Sham=9, WT RUPP=5, KO Sham=6, KO RUPP=6). Vascular function was measured via wire myography on 2 nd order mesenteric arteries and thoracic aorta. Systolic blood pressure (BP) was measured by radiotelemetry in a subset of mice. RUPP did not decrease maternal weights in either WT or KO mice (P>0.05) but decreased pup weight (726±20 WT+Sham vs 660±11mg WT+RUPP, *P<0.05) and fetal demise (16±4 vs 28±4% resoprtions/litter, *P<0.05) in WT pregnant mice, however, not in KO mice (710±15 KO+Sham vs 693±19 mg KO+RUPP, P>0.05, and 13±5 vs 28±12% resoprtions/litter, P>0.05), indicating KO protected fetal growth. RUPP reduced endothelial-dependent, acetylcholine-mediated vascular relaxation (10 -9 -3x10 -5 M) in WT, but not KO pregnant mice (*P<0.05, 2-way ANOVA w/ RM), but not aorta ACh-mediated relaxation (P>0.05). Nitric oxide synthase (NOS) inhibitor LNAME ablated differences in ACh-mediated relaxation between WT Sham and WT RUPP (P>0.05), with no differences observed in KO Sham or KO RUPP, in both aorta and mesenteric arteries, indicating RUPP impaired endothelial function via decreasing NOS activity. Preliminary data demonstrates RUPP increased BP on GD15-17 in WT (127±21 mmHg, N=3), however, not in KO pregnant mice (111±4, N=2). RUPP also ablated circadian rhythm in WT, but not KO, pregnant mice from GD15-18. Collectively, these data indicate that ECMR activation induces reduced pup weight, endothelial dysfunction and hypertension in the RUPP model which may lead to clinical studies for consideration of specific MR antagonists as therapeutic options for endothelial dysfunction in PE pregnancies.

Clinical data report a 3-5-fold increase in adverse pregnancy outcomes in women with a history of acute kidney injury (AKI), despite clinical recovery of renal function (creatinine clearance) before pregnancy. We recently showed that female Sprague-Dawley (SD) rats recovered from AKI develop increased blood pressure, uterine artery resistance and decreased fetal growth in pregnancy. Healthy pregnancies are characterized by robust nitric oxide (NO) production and reduction in total peripheral vascular resistance, however, our lab has shown decreased renal nitric oxide synthase (NOS) expression in pregnant rats post-AKI. We, therefore, tested the hypothesis that AKI prior to pregnancy decreases NO which induces endothelial dysfunction and fetal growth restriction. Female SD rats received either 45-min bilateral renal ischemia-reperfusion (AKI) or sham surgery. We utilized sildenafil to increase NO bioavailability. Sham and AKI rats were given vehicle or sildenafil chow 15 days (n=8/grp) post-AKI or sham surgery and mated 15 days later. A separate group of rats (prolonged AKI/Sham) (n=3-6/grp) were mated for pregnancy 6 months post-AKI or sham surgery. Gestational day 1 (GD1) was identified through vaginal smear. On GD19, rats were placed in metabolic cages for urine collection and euthanized on GD20. Pups were weighed and vascular function was assessed via wire myography on 3rd order mesenteric arteries. AKI prior to pregnancy reduced urinary nitrate/nitrite levels assessed via ELISA (3.2±0.9 AKI vs 8.9±1.6 μM/day Sham, *P<0.05) and trended to decrease aortic protein expression of endothelial NOS (eNOS; 0.5±0.1 AKI phospho-/total-eNOS vs 0.8±0.1 Sham, n=3/group, P=0.1) compared to sham pregnant rats. Pup weight decreased in post-AKI pregnant rats (2.14±0.02 g Sham Vehicle vs 2.02±0.05 AKI Vehicle, P=0.06), however, sildenafil restored pup weights in post-AKI rats (2.17±0.04 Sham Sildenafil vs 2.08±0.06 AKI Sildenafil). Pre-pregnancy AKI reduced endothelial-dependent relaxation to acetylcholine (ACh) compared to sham (concentration-response, 1 nM-10 μM, 2-way ANOVA, RM, *P<0.05), indicating endothelial dysfunction. Sildenafil restored endothelial relaxation responses to ACh in post-AKI pregnant rats (P>0.05). None of our groups demonstrated differences in vascular responses to the constrictor phenylephrine or relaxation to sodium nitroprusside. Interestingly, the prolonged 6-month post AKI/Sham group showed increased constrictor responses to phenylephrine (*P<0.05) and KCl (*P<0.05) in post-AKI pregnant rats compared to sham, suggesting prolonged time removed from AKI increases vascular stiffness in pregnancy. Taken together, these data indicate AKI prior to pregnancy induces endothelial dysfunction and that sildenafil, potentially by increasing NO, rescues endothelial function and fetal growth in post-AKI pregnant rats. In addition, our data show that increased recovery time does not ablate vascular dysfunction in pregnancies with history of AKI. R00 HL146948-03 to DM; AHA CDA858380 to JLF; AHA DIVSUP0224732 to DM 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.

Acute kidney injury (AKI), a sudden loss of kidney function, is responsible for 1 in 5 US hospitalizations. Although women are considered to be protected from AKI, they make up 40% of AKI patients. Recent clinical data report a 3-5-fold increase in adverse pregnancy outcomes among women with a history of AKI, despite apparent clinical recovery of renal function before pregnancy. These women have increased odds for developing preeclampsia and fetal growth restriction, via mechanisms unknown. To address this clinical issue, we developed a rat model of pregnancy post-AKI. We show that female Sprague-Dawley (SD) rats recover renal function as measured by creatinine clearance 30 days post-AKI. These rats exhibit decreased creatinine clearance, increased blood pressure, reduced plasma volume expansion and fetal growth restriction during pregnancy compared to sham rats. This study tested the hypothesis that AKI prior to pregnancy induces renal inflammation, fibrosis and dysregulates the renin-angiotensin aldosterone system (RAAS). Female SD rats were randomized to 45-minute warm sham or bilateral renal ischemia reperfusion (N=6/grp) and mated 30 days post sham/AKI surgery. Gestational day 1 (GD1) was identified through vaginal smear. Rats were placed in metabolic cages on GD19 for urine collection and euthanized on GD20. Urinary volume/day (30±5 vs 19±2 ml/day, P=0.08) and water intake (55±5 ml vs 41±1 ml, *P<0.05) increased in post-AKI pregnant rats compared to sham. In addition, urinary aldosterone levels measured by ELISA decreased in post-AKI pregnant rats compared to sham (188±34 pg/ml/day vs 312±35, *P<0.05). These data suggest disruption of fluid homeostasis and RAAS by pre-pregnancy AKI. Kidney injury marker-1 (KIM-1) measured via ELISA trended to increase in pre-pregnant AKI rats compared to sham at 30 days post AKI (580±169 pg/ml vs 257±17, P=0.09) and in pregnancy (283±28 vs 216±12 pg/ml, P=0.06), suggesting a lingering kidney injury despite creatinine clearance returning to sham levels. Mason Trichrome renal staining at GD20 revealed increases in both glomerular fibrosis (0.93±0.08 vs 0.04±0.04 score, *P<0.05) and tubular fibrosis (1.7±0.19 vs 0.0±0.0 score, *P<0.05) in post-AKI pregnant rats compared to sham. T regulatory immune (Tregs) cell populations in blood and kidney were measured via flow cytometry on GD20. Blood Tregs (16195±2935 cells/ml vs 26740±2508, *P<0.05) as well as renal Tregs (1811±364 cells/gram kidney vs 3241±405.6, *P<0.05) decreased in post-AKI pregnant rats compared to sham, suggesting AKI prior to pregnancy increases renal inflammation. Taken together, these data suggest that history of AKI predisposes to pregnancy induced dysregulation of fluid hemodynamics, renal inflammation, renal fibrosis and sustained renal injury mediating adverse fetal and maternal outcomes. R00 HL146948-03 to DM; AHA CDA858380 to JLF; AHA DIVSUP0224732 to DM 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.

Elevated leptin (lep) in pregnant women strongly associates with preeclampsia (PE), a hypertensive pregnancy disorder. Our lab established a mid-late gestation lep-infusion model of PE, which induces endothelial dysfunction, hypertension and fetal growth restriction (FGR); deletion of endothelial mineralocorticoid receptors (MR) ablated these PE characteristics in pregnant mice. Whether pharmacological MR antagonism ablates lep-induced PE, however, is unknown. We hypothesized that MR antagonism (eplerenone) ablates lep-induced PE characteristics in pregnant mice. We infused pregnant BALB/c mice with leptin (lep, 0.9mg/kg/day) or saline (sham) via osmotic minipump alongside eplerenone (Eplr, 200mg/kg/day) or vehicle in drinking water from gestation day (GD)11-18. We measured vascular function via wire myography of 2 nd order mesenteric vessels and pup/placental weights on GD18 (n=5-7). We implanted radiotelemeters in separate mice (n=1-3) prior to mating and measured mean arterial pressure (BP) from GD11-18. Pre-pregnancy BP did not differ between treatment groups. Preliminary data indicates lep increased BP compared to sham from GD11-18 (112.7±4.5 lep vs 89.4 sham mmHg,) and Eplr decreased BP in lep pregnant mice (97.2±7.2 lep+Eplr mmHg, P=0.09 vs lep student’s t-test). Vascular relaxation to endothelial-dependent acetylcholine (ACh) decreased in lep+vehicle compared to sham+vehicle (*P<0.05, 2-Way ANOVA w/ RM). Eplr+lep restored ACh-mediated vascular relaxation (*P<0.05 vs lep+vehicle). LNAME pre-incubation did not ablate differences in ACh-mediated relaxation between lep+vehicle and lep+eplr (P>0.05) indicating that Eplr restored a non-NO mechanism to improve endothelial function. Lep as a variable decreased pup weight in both vehicle and Eplr pregnant mice, however, Eplr did not increase pup weight in either vehicle or lep mice (0.74±0.01sham+veh, 0.71±0.01 lep+veh, 0.70±0.01 sham+eplr, 0.67±0.01 lep+eplr (g), *P<0.05, 2-Way ANOVA). Collectively these data indicate that Eplr improves lep-induced endothelial dysfunction and hypertension, but may not prevent FGR when given from GD11-18. Therefore, further studies will determine whether MR antagonism promises therapeutic potential in lep-induced PE.