NCT01956110.

Background: Previous studies implicated cardiotonic steroids, including Na/K-ATPase inhibitor marinobufagenin (MBG), in the pathogenesis of preeclampsia (PE). Immunoneutralization of heightened MBG by Digibind, a digoxin antibody, reduces blood pressure (BP) in patients with PE, and anti-MBG monoclonal antibody lessens BP in a rat model of PE. Recently, we demonstrated that MBG induces fibrosis in cardiovascular tissues via a mechanism involving inhibition of Fli-1, a nuclear transcription factor and a negative regulator of collagen-1 synthesis. Objectives and Methods: We hypothesized that in PE, elevated placental MBG levels are associated with development of fibrosis in umbilical arteries. Eleven patients with PE (mean BP 124 ± 4 mmHg; age 29 ± 2 years; 39 weeks gest. age) and 10 gestational age-matched normal pregnant subjects (mean BP 92 ± 2 mmHg; controls) were enrolled in the clinical study. Results: PE was associated with a higher placental (0.04 ± 0.01 vs. 0.49 ± 0.11 pmol/g; p < 0.01) and plasma MBG (0.5 ± 0.1 vs. 1.6 ± 0.5 nmol/L; p < 0.01), lower Na/K-ATPase activity in erythrocytes (2.7 ± 0.2 vs. 1.5 ± 0.2 µmol Pi/mL/hr; p < 0.01), 9-fold decrease of Fli-1 level and 2.5-fold increase of collagen-1 in placentae (p < 0.01) vs. control. Incubation of umbilical arteries from control patients with 1 nmol/L MBG was associated with four-fold decrease in Fli-1 level and two-fold increase in collagen-1 level vs. those incubated with placebo (p < 0.01), i.e., physiological concentration of MBG mimicked effect of PE in vitro. Collagen-1 abundance in umbilical arteries from PE patients was 4-fold higher than in control arteries, and this PE-associated fibrosis was reversed by monoclonal anti-MBG antibody ex vivo. Conclusion: These results demonstrate that elevated placental MBG level is implicated in the development of fibrosis of the placenta and umbilical arteries in PE.

Elevated levels of endogenous Na/K-ATPase (NKA) inhibitors, cardiotonic steroids (CTS) including marinobufagenin (MBG), contribute to pathogenesis of preeclampsia (PE) and represent a target for immunoneutralization by Digibind (Ovine Digoxin Immune Antibody, Glaxo-Smith Kline). Because Digibind is no longer commercially available we studied whether DigiFab (BTG International Ltd, UK) can substitute Digibind for immunoneutralization of CTS in patients with PE. We compared DigiFab, Digibind and anti-MBG monoclonal antibody (mAb) with respect to their ability to interact with CTS in PE plasma and to restore NKA activity in erythrocytes from patients with PE. Using immunoassays based on DigiFab, Digibind, and anti-MBG mAb we studied the elution profile of CTS following HPLC-fractionation of PE plasma. Seven patients with mild PE (28±2 years; gestational age, 39±0.5 weeks; blood pressure 156±5/94±2 mmHg) and six normotensive pregnant subjects (28±1 years; gestational age, 39±0.4 weeks; blood pressure 111±2/73±2 mmHg) were enrolled. PE was associated with a substantial inhibition of erythrocyte NKA (1.47±0.17 vs. 2.65±0.16 μmol Pi/mL/hr in control group, P<0.001). Ex vivo, at concentration 10 μg/mL, which is consistent with the clinical dosing of Digibind administered previously in PE, DigiFab and Digibind as well as anti-MBG mAb (0.5 μg/mL) restored erythrocyte NKA activity. Following HPLC fractionation of pooled PE and control plasma, PE-associated increase in CTS material was detected by Digibind (176 vs. 75 pmoles), DigiFab (221 vs. 70 pmoles) and anti-MBG mAb (1056 vs. 421 pmoles). Therefore, because DigiFab interacts with CTS from PE plasma and reverses PE-induced NKA inhibition, it can substitute Digibind for immunoneutralization of CTS in patients with PE.

Our aim was to study the expression of adipokine-encoding genes (leptin, adiponectin, and angiopoietin-like protein 4 (ANGPTL4)) in human umbilical vein endothelial cells (HUVECs) and adipokine concentration in cord blood from women with gestational diabetes mellitus (GDM) depending on glycaemic targets. GDM patients were randomised to 2 groups per target glycaemic levels: GDM1 (tight glycaemic targets, fasting blood glucose < 5.1 mmol/L and <7.0 mmol/L postprandial, N = 20) and GDM2 (less tight glycaemic targets, <5.3 mmol/L and < 7.8 mmol/L, respectively, N = 21). The control group included 25 women with normal glucose tolerance. ANGPTL4 expression was decreased in the HUVECs from GDM patients versus the control group (23.11 ± 5.71, 21.47 ± 5.64, and 98.33 ± 20.92, for GDM1, GDM2, and controls; p < 0.001) with no difference between GDM1 and GDM2. The level of adiponectin gene expression was low and did not differ among the groups. Leptin gene expression was undetectable in HUVECs. In cord blood, leptin/adiponectin ratio (LAR) was increased in GDM2 compared to controls and GDM1 (p = 0.038) and did not differ between GDM1 and controls. Tight glycaemic targets were associated with normalisation of increased LAR in the cord blood. ANGPTL4 expression was downregulated in HUVECs of newborns from GDM mothers and was not affected by the intensity of glycaemic control.

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