American Journal of Obstetrics and Gynecology (2006) 194, 1228–33

www.ajog.org

Is vitamin E a safe prophylaxis for preeclampsia?

Subhasis Banerjee, DVM, PhD,* Anne E. Chambers, PhD, Stuart Campbell, MD, DSc

Harris Birthright Research Centre for Fetal Medicine, King’s College Hospital Medical School, London, UK

Received for publication July 25, 2005; revised October 25, 2005; accepted November 21, 2005

KEY WORDSVitamin E

PreeclampsiaTh1/Th2 switchAtopic diseases

The prophylactic use of vitamins E and C for the prevention of preeclampsia is currently beingevaluated in multiple clinical trials in Canada, Mexico, the United Kingdom, the United States,and other developing countries. In addition to its antioxidant capacity, exogenous vitamin E may

prevent an immunologic switch (Th1 to Th2) that is vital for early-to late transition in normalpregnancies. Moreover, vitamin E could be a potential interferon-gamma (IFN-g) mimic facilitat-ing persistent proinflammatory reactions at the fetal-maternal interface. These untoward effectsof dietary intake of vitamin E may be more pronounced in those treated cases that fail to develop

preeclampsia. A critical test of this hypothesis would be to establish whether, under variable O2

tension, vitamin E is capable of affecting cytokine signaling in placental trophoblasts and mater-nal immune effector cells, both in early and late human pregnancies.

� 2006 Mosby, Inc. All rights reserved.

Preeclampsia (PE) is a hypertensive pregnancy disor-der marked by superficial implantation and inadequateplacental perfusion that has been linked to increasedoxidative stress. Oxidative stress is a condition where anatural antioxidant system fails to eliminate highlyreactive oxygen species (ROS), including superoxide(O2c

�) and free radical intermediates (hydroxyl, cOH;peroxynitrite, ONOOc�) produced during reduction ofparamagnetic O2. ROS production in biologic systemscan occur by a variety of mechanisms: normal aerobicmetabolism during oxidative phosphorylation in mito-chondria, activation of NADP(H) oxidases, xanthineoxidase (XO), cytochrome P450, and uncoupling ofnitric oxide synthase (NOS). ROS production could beinduced by growth factors (angiotensin II, platelet-derived

* Reprint requests: Subhasis Banerjee, DVM, PhD, Harris Birth-

right Research Centre for Fetal Medicine, King’s College Hospital

Medical School, Denmark Hill, London SE5 9RS, UK.

E-mail: dr_sbanerjee@hotmail.com

0002-9378/$ - see front matter � 2006 Mosby, Inc. All rights reserved.

doi:10.1016/j.ajog.2005.11.034

growth factor [PDGF]) and cytokines (tumor necrosisfactor-a [TNF-a]). The best documented natural antioxi-dant systems to counter excess cellular ROS accumula-tion are superoxide dismutases, catalases, glutathioneperoxidase, thioredoxin-thioredoxin reductase, and otherredox regulatory systems.1,2

Traditionally, ROS have been considered to be acci-dental by products of mitochondrial oxidative phospho-rylation (electron leakage) or as inducible, manifested bythe robust induction of NADP(H) oxidases in phago-cytes to destroy invading micro-organisms and nonselfantigens.3 Emerging evidence in recent years suggeststhat ROS-generating nonphagocytic oxidase homo-logues of the catalytic subunit gp91phox are also presentin a variety of fetal, adult, and placental tissues and areexpressed in various cell types, including epithelium,endothelium, and smooth muscle cells.4-6 Most impor-tantly, these nonphagocytic NADP(H) oxidase systems(Nox/Duox,NADPHoxidase/dual oxidase) have distinc-tive roles in ROS-mediated cellular signal transduction

Banerjee, Chambers, and Campbell 1229

(growth and angiogenesis),7 innate and adaptive immu-nity,8,9 and in oxidative remodeling of the extracellularmatrix.5,10

Given that in hypoxia, ROS could be generated byinduction of nonphagocytic NADP(H) oxidases and arecritical mitogenic signals for neoplastic growth,11 vascu-lar smooth muscle cell proliferation,12 angiogenic switch(vascular endothelial growth factor [VEGF] receptorsand matrix metalloproteinase),7 and that tumor-like in-vasive placental trophoblasts13 abundantly express Nox/Duox homologues,6,10 it is highly likely that a part ofROS produced during early placental development(8-10 weeks of gestation) are integral to trophoblast pro-liferation and vascular remodeling. This paradigm shiftraised the possibility that early hypoxic developmentof the human embryo and the placenta could be a phys-iologic necessity. This scenario, nevertheless, does notcontradict the idea that the pathogenesis of PE couldbe a consequence of ROS-induced toxicity caused bychronic development under low oxygen tension. In PE,the continued development of the fetus and the placentaat relatively reduced O2-tension engenders persistentROS production, and this, together with the failure ofthe enzymatic regulatory network,14-17 leads to ROS-mediated toxicity.

Considering these factors, the most logical prophy-lactic measure against PE might be to administer anti-oxidants in an attempt to ameliorate the oxidative stresswithout affecting the physiologic progression of thepregnancy. It has been proposed that vitamin E, anatural cell membrane-associated lipophilic antioxidant,could be an effective prophylactic measure against PE.18

This idea has recently been tested in several well-controlled clinical trials that are currently underway inCanada, Mexico, the United Kingdom, the UnitedStates, and other countries.18,19 However, of late it hasbecome apparent that in addition to the free radicalscavenging activity of vitamin E, the principal antioxi-dant component of this vitamin (a-tocopherol) alsohas nonantioxidant pleiotropic effects. These includeeffects on redox-regulated transcription,20 cell cycle,21

cytokine signaling,22 and perinatal immunoglobulin E(IgE) production in atopic diseases.23 The complexityof the effects of vitamin E therefore raises the intriguingpossibility that, in addition to relieving oxidative stressin PE, vitamin E therapy might enhance Th1 induc-tion24-28 and also prevent the essential Th1 to Th2switch24,26,28 that is necessary for early-to-late transitionin normal human pregnancy.29 In early pregnancy be-tween 8 and 12 weeks of gestation, human chorionicgonadotropin (hCG), proinflammatory cytokine (Th1)production, trophoblast differentiation, and vascular re-modeling are at their peaks and this development occursat relatively low O2 tension. A gradually improved feto-placental circulation with a concomitant increase in O2

and nutrient supply to the fetus develops in the mid to

late phase of pregnancy marked by a decline in hCGlevels and the prevalence of Th2 cytokine profiles atthe fetal-maternal interface. The immunomodulatoryfunctions of vitamin E therefore may have importantconsequences in embryonic and postnatal development.

Human pregnancy has 2 aspects of development, thegrowth and differentiation of trophoblasts for optimumuterine implantation and the ability of the placenta toprevent activation of maternal leukocytes and macro-phages after recognition of placental/fetal antigen. Onthe basis of their reactivity to alloantigens, T-helperprecursors (Thp) differentiate into either Th1 or Th2phenotypes.30 Th1 responses are usually cell-mediatedimmunity (delayed-type hypersensitivity and macro-phage activation), whereas Th2 responses are antibody-mediated humoral responses that include activation ofB cells, mast cells, and eosinophils. In human pregnancy,the Th1 (IFN-g, TNF-a and interleukin-12 [IL-12]) andTh2 (IL-4 and IL-10) cytokines have opposite effects,and the inability of the mother to switch from Th1 toTh2 cytokine profiles at the fetal-maternal interface hasbeen proposed as one of the primary causes of miscar-riage, intrauterine growth restriction, and PE.31-35 Itshould be noted that Th1/Th2 classification is based ona dichotomous feedback regulatory model of the chronicimmune responses, whereby failure to elicit 1 Th subsetfacilitates the cytokine production of the other Th re-sponse.30 However, a number of recent studies suggestthat distinct subsets of regulatory T cells (Tr and Th3) ex-ist that have neither Th1 nor Th2 cytokine phenotype.These regulatory T cells secrete transforming growth fac-tor-b (TGF-b) which inhibits both Th1 and Th2 develop-ment and therefore suppresses inflammatory disease andinduces immune tolerance.27,36,37

Here, we propose a model through which the non-antioxidant effects of vitamin E are detrimental tohuman pregnancy (Figure). IFN-g and IL-4 productionare considered to be key parameters of Th1 and Th2 im-mune responses, respectively.29,30 In this model, the pre-dominance of Th1 or Th2 cytokine profile at a givenstage of pregnancy can be controlled by positive andnegative feedback loops acting upon the Thp cells(Figure). For example, IFN-g activates Th1 pathwaydirectly by inducing IL-12/IL-12 receptor expressionin macrophages and indirectly, by negatively regulatingIL-4 production (Th2 activation) via interferon regula-tory factor 1 and 2 (IRF-1 and IRF-2).38 On the otherhand, in addition to inducing Th2 differentiation ofnaı̈ve Thp, IL-4 negatively regulates Th1 differentiationby blocking IL-12 receptor expression.39 As well asrelieving oxidative stress by scavenging free radicals,vitamin E inhibits the production of active proteinkinase C theta (PKC-q). PKC-q belongs to a family ofserine/threonine kinases and transduces extracellularsignals in response to growth factors, hormones, andneurotransmitters.40 PKC-q is essential for activation

1230 Banerjee, Chambers, and Campbell

Figure Vitamin E may prevent the Th1 to Th2 transition: a proposed schema. A, IFN-g activates IL-12 in macrophages (Mf), IL-12 receptor (IL-12R) expression in Mf, Th1 cells and stimulates the differentiation of Thp to Th1 cells, and inhibits IL-4/IL-10

production in Th2 cells. IL-4 stimulates and inhibits differentiation of Th2 and Th1 by interacting with IL-4R, respectively. VitaminE mimics the action of IFN-g inhibiting the differentiation of Thp into Th2. In normal early pregnancies, low O2 tension predom-inates and the Th1 phenotype is expressed. In normal late pregnancies, O2 tension increases and the Th1 phenotype switches to the

Th2 phenotype (represented by a horizontal black arrow). In PE, this Th1 to Th2 switch is inhibited (horizontal red arrow); B, showsthe negative regulatory effect of vitamin E (inhibition of AP-1, NF-kB) and IFN-g (activation of IRF-1 and IRF-2) on IL-4 genetranscription. Black, Activation; Red, inhibition; Thp, T-helper cell precursor; Th1, T-helper cell 1; Th2, T-helper cell 2. IL-4R andIL-12R are IL-4 and IL-12 receptors, respectively.

of transcription factors AP-1 and nuclear factor-kB(NF-kB).41,42 However, PKC-q activation in phorbol es-ter-stimulated cells is dependent on its phosphorylationand membrane translocation. Vitamin E inhibits PKC-qactivation by activating protein phosphatase 2B.43 Be-cause transcription from the IL-4 gene requires NFkBin conjunction with AP-1,28 exogenous vitamin E wouldresult in reduced synthesis of IL-4 (Figure, B), which inturn would lead to a block in the Th1 to Th2 switch30,39

that is essential for the early-to-late transition in normalpregnancy.29,31-35

IFN-g inhibits IL-4 expression via transcriptionalrepressors IRF-1 and IRF-2 (Figure, B).38 Vitamin Eblocks binding of the transcription factors NF-kB andAP-1 to the IL-4 promoter, down-regulating the expres-sion of IL-4 in peripheral blood T cells (Figure, B).28

This would lead to the inhibition of Th2 differentiationand development as well as IL-10 production. Both IL-4and IL-10 negatively regulate Th1 development by in-hibiting IL-12 receptor (IL-12Rb2) in Th1 cells andIL-12 expression in macrophages, respectively.39 As aconsequence, vitamin E-induced inhibition of the nega-tive feedback loop would facilitate Th1 dominance.29,30

Thus, despite the mechanistic differences, both vitaminE and IFN-g inhibit IL-4 production, the former indi-rectly, the latter directly. Therefore, with respect toThp recruitment and differentiation, vitamin E mostlikely mimics the effect of IFN-g, which is known tobe detrimental in the later stages of pregnancy.44 Thesystemic effects of short- or long-term dietary intake ofvitamin E on Th1 induction in experimental animals25-28

and human patients23,24 support this hypothesis. Vitamin

Banerjee, Chambers, and Campbell 1231

E-mediated inhibition of IL-4 production is central topreventing the IgE switch in B cells and the developmentof atopic disorders such as rhinoconjunctivitis andasthma.45 By paradox, however, intake of combined vita-mins E and C during pregnancy may modulate fetal andneonatal Th-cell responses to allergen, increasing thesusceptibility to postnatal atopic diseases.23

These apparently disparate systemic and localizedeffects of dietary vitamin E on human pregnancy couldbe linked to its nonantioxidant regulatory role in pla-cental morphogenesis and fetal development. This hasto be reconciled with the current knowledge of innateand adaptive immunity in human pregnancy. Much ofthe past work centers around the reactivity of theimmune effector cells at the fetal-maternal interface inresponse to placental alloantigen. However, in preg-nancy, the cytokine profile of placental trophoblasts isintegral to the development of innate immunity.33 Fur-thermore, human placenta produces significant amountsof IFN-g and IL-444,46 at early and late pregnancies,respectively. These cytokines, as part of a complex net-work, could potentially affect the innate and adaptiveimmune response at the fetal-maternal interface. Giventhat vitamin E inhibits PKC-q in nonimmune cells43

and IL-4 expression in T cells,22 one would expect a sim-ilar effect on placental trophoblasts. Administration ofvitamin E to high-risk pregnancy groups between thefirst and second trimesters could have specific adverseeffects on those pregnancies that did not develop PE.The criteria used to select high-risk pregnancy groups(14-22 weeks gestation) for randomized vitamin E con-trol trials include the measurement of mean uterineartery pulsatility index by Doppler, history of PE,diabetes, and renal failure. Although the uterine arteryscreening has been shown to have a high sensitivity forthe severe forms of the disease,47 a large proportion (ap-proximately 40%-50%) of patients from these high-riskpregnancy groups might not develop PE, ie, the placentadevelops in the absence of persistent oxidative stress. Inthese pregnancies, vitamin E could potentially modulatethe physiologic cytokine profile at the fetal-placentalunit that appears to be linked to increased susceptibilityof children to atopy and asthma.48,49 Furthermore, vita-min E may enhance persistent IFN-g production bytrophoblasts at late pregnancy in patient groups withPE.44

A common theme that vitamin E promotes Th1cytokine production has emerged in both animal modelsand human clinical trials. For example, a short-term (2weeks) dietary supplementation of high doses of vitaminE (750 mg/d) in patients with advanced colorectal cancerselectively increased the recruitment of naı̈ve T cells(CD45RAhigh) and the production of Th1 cytokines IL-2and IFN-g, whereas IL-10 (Th2) was unaffected. Al-though vitamin E increased CD4:CD8 ratios, indicatinggeneralized recruitment of Thp, it had no effect on

memory or regulatory T cells that suppress T-cell func-tion.24 An association of maternal vitamin E intake dur-ing pregnancy and Th cell responses to allergen(CD45RAhigh, which predominates in fetal and neonatalblood) suggests that maternal antioxidants influence thedevelopment of the fetal immune system.50 A recentlypublished progress report of an ongoing trial suggeststhat dietary intake of antioxidants during pregnancymay increase the risk of wheeze and eczema in early child-hood.23 The vitamin E-induced Th1 cytokine responsesare also evident in animal models. Adolfsson et al51 sug-gested that vitamin E directly affects T cells by inducingIL-2 production in naı̈ve Th cells and subsequent clonalexpansion. Vitamin E supplementation in murineAIDS25,26 and nasal allergy27 models restored IL-2/IFN-g production and reduced IgE production, respec-tively. The general conclusions from these publisheddata are consistent with the hypothesis proposed herethat vitamin E treatment could have undesirable sideeffects on fetal immune maturation as well as fetal-mater-nal interactions. In addition, our model might explain theconflicting outcomes of reported clinical trials of vitaminE on hypertensive pregnancy disorders.18,52,53

These studies, together with the hypothesis proposedhere, raise the necessity for a collective investigation bythe various groups involved in vitamin E clinical trials toexamine the possibility that adverse immunologic effectsinduced by vitamin E may harm the offspring of thesepregnancies.

Finally, in the light of the diverse effects of vitamin E,it might be wise to further investigate the nonantiox-idant regulatory roles of vitamin E on PKC-q activationand cytokine signalling (IL-4, IFN-g), under variable O2

tension, in placental trophoblasts and maternal immuneeffector cells in early and late human pregnancies, beforeevaluating the therapeutic efficacy of this vitamin in PE.

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