Abstract 548: Reduced Urinary Angiotensinogen Excretion in Preeclampsia

Objective: Preeclampsia (preE) is a hypertensive disorder of pregnancy. We reported the suppression of circulatory renin-angiotensin system (RAS) in a rat model of preE. Urinary angiotensinogen has been considered as an indicator of intrarenal angiotensin status in hypertension. Little is known about the urinary angiotensinogen in preE. This study evaluates the level of urinary excretion of angiotensinogen in preE to assess the RAS status. Methods: Normal pregnant (n=57) and preE (n=32) patients were recruited from Scott & White Hospital and had their blood drawn between 21 to 40 weeks of pregnancy. Criteria for diagnosis of preE included blood pressure >140/90 mm Hg and proteinuria >300 mg of protein/24h. Two groups of rats were used: normal pregnant (n=10) and preE rats (n=10) which were given weekly injections of desoxycorticosterone acetate and 0.9% saline to drink. Urinary angiotensinogen levels and the plasma AngII levels were assayed by ELISA. The kidney expression of (pro)renin receptor, AT 1 receptor, AT 2 receptor, and renin for the two groups of rats was measured by western blot. Results are expressed as means with SD and comparisons made using Student’s t test with p<0.05 considered significant. Results: In preE patients, the mean urinary excretion of angiotensinogen (2.0 ± 1.1 ng/mg creatinine) differed (p<0.05) from that in patients with normal pregnancy (2.7 ± 1.5 ng/mg creatinine). The urinary excretion of angiotensinogen (PreE: 1.5 ± 0.3, NP: 2. 3 ± 0.4 nmol/day) also differed (p<0.05) in preE rats compared to NP. The plasma concentration of Ang II for preE rats (25 ± 3 fmol/ml) differed (p<0.05) compared to NP rats (39 ± 5 fmol/ml). The kidney expression of (pro)renin receptor was downregulated (p=0.008), while the AT 1 receptor was upregulated (p=0.01) in preE rats compared to NP. However, the kidney expression of AT 2 receptor (1.0 vs 1.1) and renin (1.0 vs 1.4) were similar in NP and PDS rats (p=0.63 and p=0.12, respectively). Conclusions: We demonstrated that urinary excretion of angiotensinogen is reduced in both patients with preE and in a rat model of preE. The kidney expression of RAS components was either downregulated or remained unchanged in preE rats. This finding indicates that preE is an Ang-II independent form of systemic hypertension.

Abstract 314: Elevation of (pro)renin and Its Receptor in Preeclampsia: A Rat Model and Human Patients

Preeclampsia (PreE), a syndrome manifesting with hypertension, proteinuria, and edema, is a leading cause of maternal and fetal morbidity and mortality. While preE triggers are likely many and elusive, the renin-angiotensin system (RAS) has been implicated in preE pathogenesis. However, there is no data showing involvement of (pro)renin and its receptor. We recruited 32 preE and 57 normal pregnant consenting patients. (Pro)renin levels were assayed in plasma samples using an ELISA kit. An established rat model of preE was used to evaluate the role of (pro)renin and its receptor in pathogenesis. We used normal pregnant rats (NP, n=10) and pregnant rats receiving weekly injections of desoxycorticosterone acetate and whose drinking water was replaced with 0.9% saline (PreE, n=10). The plasma and placental levels of (pro)renin were assayed by ELISA. The placental levels of (pro)renin receptor was measured by gel electrophoresis of the placental homogenate followed by detection with immunoblotting using anti-ATP6IP2 antibody. The ERK1/2 phosphorylation was analyzed by immunoblotting using antibodies to total and active ERK1/ERK2 in the placenta. The mean plasma (pro)renin of 0.27 ± 0.04 μg/mL in preE patients differ (p < 0.001 using Student’s t test) from 0.15 ± 0.05 μg/mL in those without preE. Both plasma and placental levels of (pro)renin were higher (p < 0.001 using Kolmogorov-Smirnov test) in PreE rats compared to NP (Plasma (pro)renin for NP:0.21 ± 0.04 and PreE:0.49 ± 0.09 pg/mL; placental (pro)renin for NP:152 ± 79 and PreE:302 ± 42 ng/g tissue). In addition to serving as a source of (pro)renin, the placenta is also a site for signaling as ERK1/2 phosphorylation is greater (p<0.05) in placental tissue of preE rats. These data show that circulatory and uteroplacental (pro)renin and its receptor are upregulated. Together with the upregulation of ERK1/2 phosphorylation in placenta of the rat model, there is now evidence of (pro)renin and its receptor associated novel RAS activation to play a role in preE pathogenesis through (pro)renin receptor-mediated detrimental cellular signaling at the placental boundary. This offers an opportunity for interventional treatments with signal inhibitors and interference with nonclassical (pro)renin activation of RAS.

Abstract 315: Supression of Aldosterone and Progesterone in Preeclampsia: Role of Marinobufagenin: A Translational Approach

Preeclampsia (preE) is a hypertensive disorder of pregnancy. In an animal model of preE, we have shown that the urinary excretion of marinobufagenin (MBG) is elevated prior to the development of hypertension and proteinuria. Several research groups provided evidence for reduced aldosterone (Aldo) and progesterone (Prog) availability in preE. This study focuses on the alterations in Aldo & Prog in a rat model of preE, in vitro study and in preE patients. Three groups of animals were studied: normal pregnant (NP, n = 9); pregnant rats that received weekly injections of desoxycorticosterone acetate and saline drinking water (PDS; preE rat model, n = 9); and NP rats injected with MBG (7.65 μg/kg/d, NPM, n = 8). The plasma and urinary levels of Aldo were assayed in all groups. To see the effect of MBG in the secretion of Aldo, adrenal cells were treated with DMSO (vehicle), 0.1, 1, 10 or 100 nM MBG. The levels of Aldo & Prog in the culture media of MBG-treated adrenal cells and in cell lysates were measured by ELISA. We recruited 17 PreE and 23 normally pregnant patients at Scott & White hospital. Aldo, Prog and MBG levels were assayed in plasma samples by ELISA. Both plasma and urinary levels of Aldo (NP: 578.5 ± 152.4, PDS: 379.5 ± 89.6, NPM: 348.7 ± 91.3 pg/mL for plasma; NP: 86.7± 10.7, PDS: 52.0 ± 5.2, NPM: 58.9 ± 6 for urine, p<0.05 for each case) & Prog (NP: 31.5 ± 2.9, PDS: 18.6 ± 2.1, NPM: 16.5 ± 91.3 ng/mL for plasma; NP: 6.5 ± 0.5, PDS: 3.5 ± 0.4, NPM: 3.2 ± 0.5 for urine, p<0.05 for each case) were significantly lower in PDS and NPM rats compared to NP. MBG ≥ 1 nM significantly decreased the secretion of Aldo in MBG- treated adrenal cells when compared to the vehicle-treated cells. The mean plasma Aldo of 398.6 ± 115.8 pg/mL and Prog of 46.2 ± 9.5 ng/ml in PreE patients were significantly suppressed (p<0.05) from 548.4 ± 137.5 pg/mL and 82.5 ± 13.6 ng/l, respectively, in NP women. On the contrary, plasma MBG was significantly higher (p<0.05) in preE compared to NP (preE: 59 ± 17 & NP: 12 ± 2 pg/mL). These data suggest that Aldo & Prog are paradoxically concealed in PreE. Moreover, it has been suggested that MBG is involved in the downregulation of Aldo & Prog expression in preE rat model and in in vitro adrenal cells. This study provides new evidence that MBG plays a key role in the suppression of Aldo & Prog in PreE.