Placental Ischemic Rats Research Articles

Abstract 281: HO-1 Induction Increases ßENaC in Ischemic Placentas and Cultured Cytotrophoblasts

Preeclampsia is characterized by abnormal placentation that is thought to arise from inadequate cytotrophoblast migration and invasion of the spiral arteries. The resulting reduced perfusion of the utero-placental unit promotes the development of hypertension in preeclampsia. Beta Epithelial Na+ Channel (βENaC) protein, a mediator of cytotrophoblast migration in vitro, is decreased in placentas of preeclamptic patients. We have recently reported that hemeoxygenase-1 (HO-1) and its by-products reduce blood pressure in placental ischemic rats. Since HO and its byproducts can regulate ENaC expression, including βENaC, are important mediators of migration, the goal of this study was to test the hypothesis that HO-1 induction 1) stimulates βENaC expression and migration in cultured cytotrophoblasts and 2) restores βENaC expression in ischemic placentas. Cobalt protoporhyrin (CoPP, 1 - 10 μM, 48 hrs) stimulated HO-1 induction in cultured cytotrophoblasts (BEWO cells), upregulated βENaC expression, and increased spontaneous migration by 39.2 ± 6.0% (n=7, p<0.001). To determine if HO-1 induction increased placental βENaC expression and prevented hypertension, CoPP was injected subcutaneously (5mg/kg) on gestational day 14 in timed pregnant Sprague-Dawley rats with placental ischemia. MAP was increased on gestational day 19 in placental ischemic rats (141 ± 4 vs.101 ± 1 mmHg, p<0.001; n=4 per group). While HO-1 induction did not affect MAP in normal pregnant rats (101 ± 5 mmHg), MAP was significantly attenuated (p<0.01) in placental ischemic rats treated with CoPP (117 ± 1 mmHg). Western blot analysis revealed a 62 ± 22% decrease in βENaC expression in ischemic placentas, which was restored by HO-1 induction (n= 4, p<0.05). These findings suggest that HO-1 induction stimulates βENaC expression in cultured cytotrophoblasts and ischemic placentas. Future studies will determine if the beneficial effects of HO-1 occur through βENaC-mediated cytotrophoblast migration and spiral artery remodeling in ischemic placentas. Targeting the HO-1 pathway presents a potential therapeutic approach for improving placentation in preeclamptic women.

Abstract 209: Hemeoxygenase-1 Induction Restores Beta ENaC Expression in Ischemic Placentas

Preeclampsia is a pregnancy-specific disease characterized by hypertension, proteinuria, and vascular abnormalities; the etiology of which is under active investigation. Utilizing the rat model of reduced uterine perfusion pressure (RUPP), our lab previously demonstrated that induction of hemeoxygenase-1 (HO-1) reduces blood pressure and oxidative stress in placental ischemic rats. While epithelial sodium channel proteins (specifically β-ENaC) are important in maintaining vascular myogenic tone and have been implicated in cytotropholast invasion, the effect of placental ischemia and induction of HO-1 on β-ENaC expression in placentas have not been established. Therefore, the purpose of this study is to test the hypothesis that placental β-ENaC expression is reduced in ischemic placentas and restored by HO-1 induction. Normal pregnant and RUPP rats were treated with the HO-1 inducer, cobalt protoporphyrin (CoPP) (n=4 per group/treatment). Blood pressure was measured and placentas were harvested on day 19 of gestation. Blood pressure increased from 101±1 to 141±4 mmHg (p<0.0001) in response to the RUPP procedure but was reduced by HO-1 induction (117±2 mmHg). RUPP rats also had fewer pups compared to normal pregnant rats (14±1 vs. 4±2 pups). HO-1 induction did not prevent the reduction in pup number. Placental β-ENaC protein expression was reduced by 62% in the RUPPs compared to the normal pregnant rats (p=0.036). HO-1 induction restored placental expression of β-ENaC to the levels of the normal pregnant rats. These findings suggest that HO-1 may exert its protective effects by altering the placental environment through changes in β-ENaC expression.

Activating autoantibodies to the angiotensin II type I receptor play an important role in mediating hypertension in response to adoptive transfer of CD4+ T lymphocytes from placental ischemic rats

Hypertension in rats with chronic placental ischemia (reduced uterine perfusion pressure, RUPP) is associated with elevated inflammatory cytokines, agonistic autoantibodies to the angiotensin II type I receptor (AT1-AA) and CD4(+) T cells; all of which are elevated in preclamptic women. Additionally, we have shown that adoptive transfer of RUPP CD4(+) T cells increases blood pressure, inflammatory cytokines, and sFlt-1. The objective of this study was to determine the long-term effects of RUPP CD4(+) T cells on AT1-AA, renal and systemic hemodynamics in pregnant rats. To answer this question CD4(+) T splenocytes were magnetically isolated on day 19 of gestation from control RUPP and normal pregnant (NP) rats and injected into a new group of NP rats at day 13 of gestation. On day 19 of gestation mean arterial pressure (MAP) and renal function (glomerular filtration rates, GFR) were analyzed and serum collected for AT1-AA analysis. To determine a role for AT1-AA to mediate RUPP CD4(+) T cell-induced blood pressure increases, MAP was analyzed in a second group of rats treated with AT1 receptor blockade losartan (10 mg·kg(-1)·day(-1)) and in a third group of rats treated with rituximab, a B cell-depleting agent (250 mg/kg) we have shown previously to decrease AT1-AA production in RUPP rats. MAP increased from 101 ± 2 mmHg NP to 126 ± 2 mmHg in RUPP rats (P < 0.001) and to 123 ± 1 mmHg in NP rats injected with RUPP CD4(+) T cells (NP+RUPP CD4(+)T cells) (P < 0.001). Furthermore, GFR decreased from 2.2 ml/min (n = 7) in NP rats to 1.0 ml/min (n = 5) NP+RUPP CD4(+)T cell. Circulating AT1-AA increased from 0.22 ± 0.1 units in NP rats to 13 ± 0.7 (P < 0.001) units in NP+RUPP CD4(+)T cell-treated rats but decreased to 8.34 ± 1 beats/min in NP+RUPP CD4(+) T cells chronically treated with rituximab. Hypertension in NP+RUPP CD4(+)T cell group was attenuated by losartan (102 ± 4 mmHg) and with B cell depletion (101 ± 5 mmHg). Therefore, we conclude that one mechanism of hypertension in response to CD4(+) T lymphocytes activated during placental ischemia is via AT1 receptor activation, potentially via AT1-AA during pregnancy.

Role of angiotensin II type I receptor agonistic autoantibodies (AT1-AA) in preeclampsia

Despite being one of the leading causes of maternal death and a major contributor of maternal and perinatal morbidity, the mechanisms responsible for the pathogenesis of preeclampsia are unclear. One important initiating event in preeclampsia is thought to be reduced placental perfusion leading to the production of a variety of factors that cause widespread dysfunction of the maternal vasculature. The major objective of this review is to discuss the potential role of a novel agonistic autoantibody to the angiotensin II type I receptor (AT1-AA) in mediating hypertension during pregnancy. Although animal studies suggest that increasing plasma AT1-AA concentration in pregnant rats to levels observed in preeclamptic women or placental ischemic rats result in significant increases in arterial pressure, the quantitative importance of AT1-AA in the pathophysiology of preeclampsia in humans has yet to be fully elucidated.

Hypertension in response to autoantibodies to the angiotensin II type I receptor (AT1-AA) in pregnant rats: role of endothelin-1.

Agonistic autoantibodies to the angiotensin II type I receptor (AT1-AA) and endothelin -1 (ET-1) are suggested to be important links between placental ischemia and hypertension during preeclampsia. Activation of the angiotensin II type 1 receptor (AT1R) increases endothelial cell production of ET-1; however, the importance of ET-1 in response to AT1-AA-mediated AT1 R activation during preeclampsia is unknown. Furthermore, the role of AT1-AA-mediated increases in blood pressure during pregnancy remains unclear. The objective of this study was to test the hypothesis that AT1-AA, increased to levels observed in preeclamptic women and placental ischemic rats, increases mean arterial pressure (MAP) by activation of the ET-1 system. Chronic infusion of purified rat AT1-AA into normal pregnant (NP) rats for 7 days increased AT1-AA from 0.68+/-0.5 to 10.88+/-1.1 chronotropic units (P<0.001). The increased AT1-AA increased MAP from 99+/-1 to 119+/-2 mm Hg (P<0.001). The hypertension was associated with significant increases in renal cortices (11-fold) and placental (4-fold) ET-1. To determine whether ET-1 mediates AT1-AA-induced hypertension, pregnant rats infused with AT1-AA and NP rats were treated with an ET(A) receptor antagonist. MAP was 100+/-1 mm Hg in AT1-AA+ET(A) antagonist-treated rats versus 98+/-2 mm Hg in ET(A) antagonist-treated rats. Collectively, these data support the hypothesis that one potential pathway whereby AT1-AAs increase blood pressure during pregnancy is by an ET-1-dependent mechanism.