Activin and NADPH-oxidase in preeclampsia: insights from in vitro and murine studies

doi:10.1016/j.ajog.2014.07.021

American Journal of Obstetrics and Gynecology

Volume 212, Issue 1, January 2015, Pages 86.e1-86.e12

https://doi.org/10.1016/j.ajog.2014.07.021 Get rights and content

Objective

Clinical management of preeclampsia has remained unchanged for almost 5 decades. We now understand that maternal endothelial dysfunction likely arises because of placenta-derived vasoactive factors. Activin A is one such antiangiogenic factor that is released by the placenta and that is elevated in maternal serum in women with preeclampsia. Whether activin has a role in the pathogenesis of preeclampsia is not known.

Study Design

To assess the effects of activin on endothelial cell function, we cultured human umbilical vein endothelial cells in the presence of activin or serum from normal pregnant women or pregnant women with preeclampsia, with or without follistatin, a functional activin antagonist or apocynin, a NADPH oxidase (Nox2) inhibitor. We also administered activin to pregnant C57Bl6 mice, with or without apocynin, and studied maternal and fetal outcomes. Last, we assessed endothelial cell Nox2 and nitric oxide synthase expression in normal pregnant women and pregnant women with preeclampsia.

Results

Activin and preeclamptic serum induced endothelial cell oxidative stress by Nox2 up-regulation and endothelial cell dysfunction, which are effects that are mitigated by either follistatin or apocynin. The administration of activin to pregnant mice induced endothelial oxidative stress, hypertension, proteinuria, fetal growth restriction, and preterm littering. Apocynin prevented all of these effects. Compared with normal pregnant women, women with preeclampsia had increased endothelial Nox2 expression.

Conclusion

An activin-Nox2 pathway is a likely link between an injured placenta, endothelial dysfunction, and preeclampsia. This offers opportunities that are not novel therapeutic approaches to preeclampsia.

Section snippets

Blood and tissue collection

Blood and tissues were collected from women after written, informed consent was obtained and with the approval of the Southern Health Human Research Ethics Committee. Maternal blood was collected from the antecubital vein of women with a singleton pregnancy in absence of labor. Samples were obtained from women with established preeclampsia, as defined by the Society of Obstetric Medicine of Australia and New Zealand,¹⁶ who were receiving labetalol and/or nifedipine and from women with a healthy

Effects of activin on HUVECs in vitro

Activin A (50 ng/mL) increased the production of ROS (Figure 1, A) and the lipid peroxidation product 8-isoprostane (Figure 1, B) from HUVECs, increased transendothelial permeability, and reduced transendothelial resistance in a HUVEC monolayer (Figure 1, C and D). All of these effects were prevented by the addition of follistatin (FS288), which is an activin-binding protein that blocks activin activity (Figure 1). Exploring the mechanism of impaired endothelial integrity after activin

Comment

In this study, we have shown that activin can induce endothelial dysfunction by NOX-mediated oxidative stress and that, in vitro, antagonizing either activin or NOX activity mitigates the disruptive endothelial effects of preeclamptic serum. We have also shown that the administration of activin to pregnant mice induces a preeclampsia-like condition that is prevented by the coadministration of the Nox-inhibitor apocynin. Taken together with the clinical observation that maternal circulating

References (45)

J.M. Roberts et al.
Pre-eclampsia: more than pregnancy-induced hypertension
Lancet
(1993)
B. Sibai et al.
Pre-eclampsia
Lancet
(2005)
S. Muttukrishna et al.
Activin A and inhibin A as possible endocrine markers for pre-eclampsia
Lancet
(1997)
P. Lagrange et al.
Transendothelial permeability changes induced by free radicals in an in vitro model of the blood-brain barrier
Free Rad Biol Med
(1999)
P. Ancuta et al.
Transendothelial migration of CD16+ monocytes in response to fractalkine under constitutive and inflammatory conditions
Immunobiology
(2004)
J.D. Morrow et al.
The isoprostanes: current knowledge and directions for future research
Biochem Pharmacol
(1996)
L.J. Roberts et al.
Measurement of F(2)-isoprostanes as an index of oxidative stress in vivo
Free Radic Biol Med
(2000)
E.T. McKinney et al.
Plasma, urinary, and salivary 8-epi-prostaglandin f2alpha levels in normotensive and preeclamptic pregnancies
Am J Obstet Gynecol
(2000)
J.M. Roberts et al.
Is oxidative stress the link in the two-stage model of pre-eclampsia?
Lancet
(1999)
X.L. Cui et al.
Expression of NADPH oxidase isoform 1 (Nox1) in human placenta: involvement in preeclampsia
Placenta
(2006)

L. Poston et al.

2006 Vitamin C and vitamin E in pregnant women at risk for pre-eclampsia (VIP trial): randomised placebo-controlled trial

Lancet

(2006)

C.W. Redman et al.

Latest advances in understanding preeclampsia

Science

(2005)

J.S. Gilbert et al.

Pathophysiology of hypertension during preeclampsia: linking placental ischemia with endothelial dysfunction

Am J Physiol Heart Circ Physiol

(2007)

C.W.G. Redman et al.

Placental stress and pre-eclampsia: a revised view

Placenta

(2009)

S.E. Maynard et al.

Excess placental soluble fms-like tyrosine kinase 1 (sFlt1) may contribute to endothelial dysfunction, hypertension, and proteinuria in preeclampsia

J Clin Invest

(2003)

R.J. Levine et al.

Circulating angiogenic factors and the risk of preeclampsia

N Engl J Med

(2004)

R.J. Levine et al.

Soluble endoglin and other soluble antiangiogenic factors in preeclampsia

N Engl J Med

(2006)

S. Venkatesha et al.

Soluble endoglin contributes to the pathogenesis of preeclampsia

Nat Med

(2006)

R. Thadhani et al.

Pilot study of extracorporeal removal of soluble Fms-like tyrosine kinase 1 in preeclampsia

Circulation

(2011)

W. Vale et al.

Activins and inhibins and their signalling

Ann N Y Acad Sci

(2004)

U. Manuelpillai et al.

Activin A and activin receptors in gestational tissue from preeclamptic pregnancies

J Endocrinol

(2001)

D. D’Antona et al.

Increased maternal serum activin A but not follistatin levels in pregnant women with hypertensive disorders

J Endocrinol

(2000)

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Ferroptosis, a novel mode of cell death characterized by lipid peroxidation and oxidative stress, plays an important role in the pathogenesis of preeclampsia (PE). The aim of this study is to determine the role of Nox2 in the ferroptosis of trophoblast cells, along with the underlying mechanisms.
The mRNA and protein levels of Nox2, STAT3, and GPX4 in placental tissues and trophoblast cells were respectively detected by qRT-PCR and western blot analysis. CCK8, transwell invasion and tube formation assays were used to evaluate the function of trophoblast cells. Ferroptosis was evaluated using flow cytometry and the lipid peroxidation assay. Glycolysis and mitochondrial respiration were investigated by detecting the extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) using Seahorse extracellular flux technology. The t-test or one-way ANOVA was used for statistical analysis.
Nox2 was up-regulated while STAT3 and GPX4 were down-regulated in PE placental tissues. Nox2 knockdown inhibited ferroptosis in trophoblast cells, which was shown by enhanced proliferation and invasion, decreased ROS and lipid peroxide levels, and reduced glycolysis and mitochondrial dysfunction. Nox2 negatively correlated with MVD in PE placentas, and Nox2 knockdown restored ferroptosis-inhibited tube formation. Nox2 could interact with STAT3. Inhibiting Nox2 restored ferroptosis-induced alterations in the mRNA and protein levels of STAT3 and GPX4.
Nox2 may trigger ferroptosis through the STAT3/GPX4 pathway, subsequently leading to regulation of mitochondrial respiration, transition of glycolysis, and inhibition of placental angiogenesis. Therefore, targeted inhibition of Nox2 is expected to become a new therapeutic target for PE.
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Early diagnosis and efficient treatment of preeclampsia remains a medical challenge and etiological factors converge in a deficient placentation that triggers oxidative stress. There is evidence that statins show antioxidant effects that can improve endothelial function without adverse perinatal effects. We aimed to compare early vs. late pravastatin treatment on the oxidative stress and cardiovascular features of an experimental model of preeclampsia. Female Wistar rats were randomly divided into preeclampsia phenotype rats (PEP) developed by sub renal aortic coarctation (SRAC) and healthy pregnant rats (C). Each group received pravastatin (5 mg/Kg) p.o. either for one week before and during the first week or during the last two weeks of gestation. Blood pressure was determined using the plethysmographic method. Phenylephrine (Phe)-induced contractility was evaluated in isolated thoracic and abdominal aortic rings with or without endothelium. Blood samples were obtained to determine anion superoxide concentration as indicator of NADPH activity. Two-way ANOVA and Bonferroni post hoc tests were used to define statistical significance. Early or late pravastatin treatment decreased hypertension of PEP animals but did not change BP of the healthy pregnant group. Thoracic and abdominal aorta from PEP rats showed increased contractility that was reverted by pravastatin early treatment in endothelium intact rings. Pravastatin did not significantly change contractility neither in the thoracic nor in the abdominal aorta segments from healthy pregnant control rats (C), and decrease anion superoxide concentration by NADPH activity. We conclude pravastatin can improve both blood pressure and endothelium-dependent Phe-induced contractility in an experimental model of preeclampsia by reducing oxidative stress.
Melatonin-mediated actions and circadian functions that improve implantation, fetal health and pregnancy outcome
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This review summarizes data related to the potential importance of the ubiquitously functioning antioxidant, melatonin, in resisting oxidative stress and protecting against common pathophysiological disorders that accompany implantation, gestation and fetal development. Melatonin from the maternal pineal gland, but also trophoblasts in the placenta, perhaps in the mitochondria, produce this molecule as a hedge against impairment of the uteroplacental unit. We also discuss the role of circadian disruption on reproductive disorders of pregnancy. The common disorders of pregnancy, i.e., stillborn fetus, recurrent fetal loss, preeclampsia, fetal growth retardation, premature delivery, and fetal teratology are all conditions in which elevated oxidative stress plays a role and experimental supplementation with melatonin has been shown to reduce the frequency or severity of these conditions. Moreover, circadian disruption often occurs during pregnancy and has a negative impact on fetal health; conversely, melatonin has circadian rhythm synchronizing actions to overcome the consequences of chronodisruption which often appear postnatally. In view of the extensive findings supporting the ability of melatonin, an endogenously-produced and non-toxic molecule, to protect against experimental placental, fetal, and maternal pathologies, it should be given serious consideration as a supplement to forestall the disorders of pregnancy. Until recently, the collective idea was that melatonin supplements should be avoided during pregnancy. The data summarized herein suggests otherwise. The current findings coupled with the evidence, published elsewhere, showing that melatonin is highly protective of the fertilized oocyte from oxidative damage argues in favor of its use for improving pregnancy outcome generally.
Association of activin A and postpartum blood pressure in peripartum cardiomyopathy
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Activin A has been implicated in the pathogenesis of patients with chronic hypertension and heart failure as well as patients with hypertensive disorders of pregnancy (HDP). Whether activin A correlates with blood pressure in patients with peripartum cardiomyopathy (PPCM) and HDP history has not previously been explored.
82 women with PPCM w/ and w/out HDP or hypertension history were selected for analysis from the Investigations in Pregnancy Associated Cardiomyopathy (IPAC) study. Serum biomarkers and blood pressure were assessed at the time of enrollment (median postpartum day 24). Levels of both sFlt-1 (SBP: r 0.47, p = 0.008; DBP: r 0.57, p < 0.001) and activin A (SBP: r 0.59, p < 0.001; DBP: r 0.68, p < 0.001) were noted to significantly correlate with blood pressure in patients with a history of HDP who went on to develop PPCM, but not in patients with chronic hypertension or no hypertensive history. The strongest correlation was between activin A levels and postpartum diastolic blood pressure for the subset with preeclampsia (DBP: r 0.82, p < 0.001). This remained significant in multivariable linear regression analysis (DBP: β = 0.011, p = 0.015).
In patients with PPCM, activin A and sFlt-1 levels had direct correlations with both systolic (SBP) and diastolic blood pressures (DBP), but only in participants with history of HDP. This correlation was more evident for activin A and strongest with a history of preeclampsia. Our findings suggest that activin A may play an important role in blood pressure modulation in women with HDP who subsequently develop PPCM.
Activin A and pathologies of pregnancy: a review
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Activin A is a two-subunit protein belonging to the transforming growth factor β superfamily. First discovered almost three decades ago, it has since been implicated in diverse physiological roles, ranging from wound repair to reproduction. After 30 years of research, altered activin A levels are now understood to be associated with the development of various diseases, making activin A a potential therapeutic target. In pregnancy, the placenta and fetal membranes are major producers of activin A, with significantly enhanced serum concentrations now recognised as a contributor to numerous gestational disorders. Evidence now suggests that circulating levels of activin A may be clinically relevant in the early detection of pregnancy complications, including miscarriage and preeclampsia. This review aims to summarise our current understanding of activin A as a potential diagnostic marker in common pregnancy pathologies.
Circulating Activin A is elevated at 36 weeks’ gestation preceding a diagnosis of preeclampsia
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Citation Excerpt :
Activin A is an established anti-angiogenic factor, that functions independently to the endothelial dysfunction mechanisms induced by the two best-performing preeclampsia biomarkers, soluble fms-like tyrosine kinase 1 (sFlt1) and soluble endoglin (sENG) [19]. This is predominantly through the induction of oxidative stress pathways in endothelial cells, and endothelial dysfunction through the reactive oxygen species (ROS)-generating enzyme, Nox2 [20]. It is now widely hypothesised that term preeclampsia may be more associated with maternal endothelial dysfunction than poor placental invasion [21,22], which is more likely to present with preterm disease [23–25].
Activin A is aberrantly expressed by the preeclamptic placenta and circulating levels have been investigated as a potential biomarker for the disease. In a nested case-control study we measured Activin A levels in maternal plasma at 28- and 36-weeks’ gestation preceding term preeclampsia diagnosis. At 28 weeks Activin A was not significantly altered (n = 73 destined to develop preeclampsia vs n = 191 controls). At 36 weeks’ gestation Activin A was significantly increased in 40 women destined to develop preeclampsia relative to 201 controls (p < 0.0001). These findings provide further validation of Activin A as a potential biomarker for subsequent term preeclampsia.

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: Supported by funding from the Patricia Penrose bequest, Monash University Faculty of Medicine, Nursing, and Health Sciences Strategic Grant Scheme; the Royal Australian and New Zealand College of Obstetricians and Gynaecologists Research Foundation; National Health and Medical Research Council (NHMRC) Project Grant number 1029148; and the Victorian Government’s Operational Infrastructure Support Program.

: The authors report no conflict of interest.

: Cite this article as: Lim R, Acharya R, Delpachitra P, et al. Activin and NADPH-oxidase in preeclampsia: insights from in vitro and murine studies. Am J Obstet Gynecol 2015;212:86.e1-12.

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ResearchObstetricsActivin and NADPH-oxidase in preeclampsia: insights from in vitro and murine studies

Objective

Study Design

Results

Conclusion

Section snippets

Blood and tissue collection

Effects of activin on HUVECs in vitro

Comment

Lancet

Lancet

Lancet

Free Rad Biol Med

Immunobiology

Biochem Pharmacol

Free Radic Biol Med

Am J Obstet Gynecol

Lancet

Placenta

Lancet

Latest advances in understanding preeclampsia

Science

Pathophysiology of hypertension during preeclampsia: linking placental ischemia with endothelial dysfunction

Am J Physiol Heart Circ Physiol

Placental stress and pre-eclampsia: a revised view

Placenta

Excess placental soluble fms-like tyrosine kinase 1 (sFlt1) may contribute to endothelial dysfunction, hypertension, and proteinuria in preeclampsia

J Clin Invest

Circulating angiogenic factors and the risk of preeclampsia

N Engl J Med

Soluble endoglin and other soluble antiangiogenic factors in preeclampsia

N Engl J Med

Soluble endoglin contributes to the pathogenesis of preeclampsia

Nat Med

Pilot study of extracorporeal removal of soluble Fms-like tyrosine kinase 1 in preeclampsia

Circulation

Activins and inhibins and their signalling

Ann N Y Acad Sci

Activin A and activin receptors in gestational tissue from preeclamptic pregnancies

J Endocrinol

Increased maternal serum activin A but not follistatin levels in pregnant women with hypertensive disorders

J Endocrinol

Research
Obstetrics
Activin and NADPH-oxidase in preeclampsia: insights from in vitro and murine studies