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Epidemiología Molecular de la PE 2003
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Transcript of Epidemiología Molecular de la PE 2003
CHIEF EDITOR’S NOTE: This article is part of a series of continuing education activities in this Journal through which a totalof 36 AMA/PRA category 1 credit hours can be earned in 2003. Instructions for how CME credits can be earned appear onthe last page of the Table of Contents.
Molecular Epidemiology of PreeclampsiaMelissa L. Wilson, MPH,* Thomas Murphy Goodwin, MD,† Vivien L. Pan, MD‡
and Sue Ann Ingles, Dr PH¶*Research Assistant and PhD Candidate, ¶Assistant Professor of Preventive Medicine and Co-Director,
Program in Molecular Epidemiology, Department of Preventive Medicine, and ‡Fellow and ClinicalInstructor, Division of Maternal-Fetal Medicine, Department of Obstetrics and Gynecology, University of
Southern California Keck School of Medicine; and †Chief, Division of Maternal-Fetal Medicine, University ofSouthern California Women’s and Children’s Hospital, Los Angeles, California
Numerous articles have been published that address the possible genetic influences on thedevelopment of preeclampsia (PE). However, to our knowledge, a complete review of the resultshas not yet been completed. We undertook a MEDLINE search to identify English-language articlespublished after January 1, 1990 that examined the possible role of specific genes in the etiology ofPE. After a brief introduction and a concise review of the prevailing etiologic hypotheses, we havecategorized the candidate genes into six categories, based on their hypothesized role in PEetiology. The purpose of this paper is to review the literature, comment on its quality, and providea reference for researchers interested in the molecular epidemiology of preeclampsia.
Target Audience: Obstetricians & Gynecologists, Family PhysiciansLearning Objectives: After completion of this article, the reader will be able to list the prevailing
etiologic hypotheses of preeclampsia, to outline the published data on possible genetic influence on thedevelopment of preeclampsia, and to clearly state the definition of preeclampsia.
INTRODUCTION
Preeclampsia (PE) is a hypertensive disorder spe-cific to pregnancy and is a major cause of maternaland neonatal death and morbidity worldwide, affect-ing nearly 6% of all pregnancies (1, 2). As much as15% to 20% of maternal mortality in developedcountries can be attributed to PE (3). Severity rangesfrom a mild disorder associated with transient hyper-tension in the later part of pregnancy to a life-threat-ening disorder with seizures, HELLP (hemolysis,elevated liver enzymes, and low platelets) syndrome,fetal hypoxia, and growth retardation (4). These moresevere sequelae are much less common than milddisease, affecting approximately 0.56 pregnancies
per 1000 deliveries (5). PE also predisposes womento other serious pregnancy complications such asabruptio placentae, acute renal failure, cerebral hem-orrhage, disseminated intravascular coagulation, andcirculatory collapse (6). Despite the large number ofstudies focusing on PE, the etiology of this diseaseremains to be elucidated. Identification of geneticrisk factors, or of a population with an exceptionallyhigh risk of disease, could aid substantially in theunderstanding of this important public health prob-lem and provide clues for the prevention or treatmentof PE.
We undertook a literature review of preeclampsiaand genetics, focusing on studies that used epidemi-ologic methods to conduct their research. Specifi-cally, our aim is to provide a review of the candidategenes examined thus far, discuss how they fit into theprevailing etiologic hypotheses, and to evaluate thequality of the evidence that these genes affect the risk
Reprint requests to: Sue Ann Ingles, Dr PH, USC/Norris CancerCenter, Room 6419, 1441 Eastlake Avenue, MS 44, Los Angeles,CA 90089. Email: [email protected]
The authors have disclosed no significant financial or otherrelationship with any commercial entity.
CME REVIEWARTICLEVolume 58, Number 1OBSTETRICAL AND GYNECOLOGICAL SURVEY
Copyright © 2002by Lippincott Williams & Wilkins, Inc. 1
39
of PE. By doing so, we hope to provide a clearerdirection for future research.
METHODS
Three methods were used to select papers for thisreview. First, we undertook a MEDLINE search us-ing the keywords preeclampsia, genetics, and epide-miology. We focused on peer-reviewed papers pub-lished between January 1, 1990 and December 31,2001, written in the English language and limited tohumans. Next, we performed a search using Web ofScience (v4.3.1) using the keywords preeclampsiaand genetics and focusing the search on papers pub-lished after 1990 in the English language. From thelist generated, we selected papers that conductedtheir research in humans and performed their studiesusing epidemiological methods. Finally, we selectedpapers from the bibliography sections of the papersidentified above that dealt directly with the topics ofinterest.
In evaluating the quality of the evidence, we fo-cused primarily on issues of sample size, case defi-nitions, and possible confounding and bias. We havecategorized the candidate genes into six categories,based on their hypothesized mechanism of action.Results for each candidate gene will be summarizedand, where applicable, assessments of study qualitywill be included. We have only attempted to assessthe quality of the epidemiological evidence, althoughother evidence may be cited.
BACKGROUND
The epidemiology of PE has been complicated bythe varying definitions and criteria used to diagnoseit. To allow for comparisons across studies, a strict,consistent definition of the disease requiring bothhypertension and proteinuria is essential. To that end,PE is best defined as an increase in blood pressure to�140/90 mm Hg or a 30/15-mm Hg increase overbaseline, on at least two occasions at least 6 hoursapart after 20 weeks of gestation and resolvingshortly after delivery, accompanied by significantproteinuria (at least 30 mg/dl on a random urinesample or 300 mg in 24 hours) (7–9). Althoughhaving a history of severe or early PE increases one’srisk for developing PE in a subsequent pregnancy(10), PE is considered to be primarily a disease offirst pregnancies (11).
Further complicating matters, evidence suggeststhat PE is likely to be a two-stage process (12, 13),requiring input from both the fetus (via the placenta)
and the mother (via underlying susceptibility). Ac-cording to Ness and Roberts (13), PE can result frompoor placental perfusion (a component that is likelyto be hereditary) combined with an underlying ma-ternal condition that may or may not be diagnosedbefore pregnancy and may or may not have a hered-ity component. Ideally, a study of genetic suscepti-bility to PE would focus on hypertension and pro-teinuria that is primarily of placental origin since it isthis form that holds the key to the unique contribu-tion of pregnancy to this disease.
ETIOLOGY OF PREECLAMPSIA
The etiology of PE is unknown. However, PE isknown to be the result of pathological changes inplacental development with subsequent endothelialcell dysfunction, which accounts for its clinical signs.To understand what is abnormal about preeclampticplacentation, a brief review of normal placentation isnecessary.
The placenta develops primarily from fetally de-rived cells known as trophoblasts. The trophoblastsinitially differentiate into two types, the cytotropho-blasts, which are the precursors to all subsequenttrophoblast cells and the syncytiotrophoblasts, whichare responsible for the invasion into the decidua, andin particular, into the maternal spiral arteries. Thereare two waves of trophoblastic invasion, one at thebeginning of pregnancy and another later in preg-nancy, around 14 to 16 weeks of gestation (14). Theinvasion of the syncytiotrophoblasts into the spiralarteries results in a widening of these arteries toapproximately 4- to 6-fold their width in nonpregnantwomen, thereby increasing the blood flow availableto the developing fetus and placenta. In PE, tropho-blastic invasion and subsequent remodeling of thespiral arteries, especially during the second wave ofinvasion, is deficient, resulting in spiral artery diam-eters that are only about 40% as wide as those in anormal pregnancy. The result is placental ischemiaand poor placental perfusion in women who willeventually develop the clinical signs of PE.
There are four main etiologic factors believed to beinvolved in the development of PE (12, 15): 1) immunemaladaptation, 2) placental ischemia, 3) oxidativestress, and 4) genetic susceptibility (12, 15, 16). Thesecategories are not mutually exclusive, and in reality,the etiology is likely to be a combination of the four(Fig. 1).
40 Obstetrical and Gynecological Survey
Immune Maladaptation
PE may result from an abnormal maternal immuneresponse to paternally derived antigens on the tro-phoblast. The evidence for the involvement of anabnormal immune response in PE developmentcomes from epidemiologic research, including thefollowing observations: 1) the risk of preeclampsia isdecreased after the first pregnancy, 2) the protectiveeffect of multiparity is largely lost with a change inpartner, 3) prior abortion or blood transfusion pro-tects against preeclampsia, 4) artificial donor insem-ination and oocyte donation lead to an increase inrisk of PE, and 5) increased exposure to semen (e.g.,length of cohabitation, use of oral contraceptives)may be protective.
Evidence for the role of immune maladaptation inthe etiology of PE also comes from reports of immu-nologic phenomena occurring in women with PE.These include antibodies against endothelial cells;increased circulating immune complexes; comple-ment activation; complement and immune complexdeposition in spiral arteries, placenta, liver, kidney,and skin; altered TH1:TH2 profile; decreased sup-pression of T cell receptor chain CD3�; and elevatedconcentrations of proinflammatory cytokines (10).Thus, it is certain that women with PE manifestimmunopathology; however, it is not certain whetherthis immunopathology is the cause or the result ofPE.
The mechanism by which immune maladaptationis related to the endothelial cell dysfunction seen inPE is uncertain, but it is postulated that activatedimmune cells from the decidua may release media-tors that act on the endothelial cells. Several possiblemediators have been suggested. Specifically, plasmaelastase levels have been found to be elevated in PE(15). Elastase and other toxic proteases are releasedby activated neutrophils. Alternatively, cytokines(particularly tumor necrosis factor alpha [TNF-�]
and interleukin-1 [IL-1]) have been suggested as thecause of the endothelial dysfunction seen in PE be-cause both may be elevated in plasma and/or amni-otic fluid of preeclamptics and can have endothelialeffects that resemble the changes observed in PE(15).
Placental Ischemia
The placental ischemia hypothesis suggests that thedisease process begins with the failure of the spiralarteries to widen in response to the increased vascu-lar demands of pregnancy, leading to a deficientblood supply to the placenta [reviewed in 10, 15, 17].
At least two theories have been put forth to linkplacental ischemia to endothelial cell dysfunction.First, increased deportation of syncytiotrophoblastmicrovillous membrane (STBM) particles from pre-eclamptic compared with normal placentas has beendemonstrated to disrupt endothelial cells and inhibittheir proliferation (18, 19). Alternatively, oxidativestress secondary to placental ischemia might lead toendothelial cell dysfunction (12). Reduced organ per-fusion followed by the return of normal oxygenationis known to cause the formation of reactive oxygenspecies such as superoxide radicals (12). In concertwith maternal changes in lipid metabolism and/or amaternal predisposition such as hyperhomocysteine-mia or antioxidant deficiency, reduced placental per-fusion might result in oxidative stress that the motheris not capable of counteracting, thereby leading toendothelial cell dysfunction and PE.
Oxidative Stress
Pregnancy increases energy demands, a fact that isreflected by the accumulation of very low-densitylipoproteins (VLDL) throughout pregnancy (20).Free fatty acids (FFA) are increased in women with
Fig. 1. Postulated interplay between the various etiologic factors.
Molecular Epidemiology of Preeclampsia Y CME Review Article 41
PE as much as 15 to 20 weeks before the onset ofclinical disease (21). Plasma albumin, which nor-mally exerts a toxicity-preventing activity, is lesseffective in this respect when there are increasedamounts of circulating FFA. Thus, triglycerides tendto accumulate in the endothelial cells of preeclampticwomen.
According to the oxidative stress hypothesis, oxi-dative stress generated in the hypoxic placenta istransferred to the systemic circulation, resulting inoxidative damage to the vascular endothelial cellsthroughout the body (12, 22). Short-lived reactiveoxygen species may interact with lipids to form sta-ble lipid peroxidation products that are potentiallyvery damaging to cell structures. Dyslipidemia earlyin preeclamptic pregnancies can lead to the accumu-lation of small, dense LDL in the subendothelialspace, which are easily oxidized to form highly re-active oxidized LDL (22). Although lipid peroxidesare known to increase in normal pregnancy, thisincrease is usually offset by an increase in antioxi-dant activity. In preeclamptic pregnancies, there is adecrease in net antioxidant activity and, thus, thepotential for damage by oxygen free radicals (15).
Preeclampsia as a Genetic Disease
Evidence for a genetic component comes from theobservation that there is a marked increase in pre-eclampsia among mothers, daughters, sisters, andgranddaughters of women who have had preeclamp-sia, but not in women related through marriage (i.e.,in-laws) (23–25). Some studies have found that theincreased risk is greatest for the daughters of a pre-eclamptic pregnancy, even greater than the risk for asister born of a normotensive pregnancy (26), al-though others have found a similar increase in riskfor all daughters born to a mother with a history ofPE (24, 27).
Additionally, higher concordance rates amongmonozygotic twins compared with dizygotic twinssuggest a role for genetics in the development ofdisease (28). However, the observation that a highnumber of monozygotic twin sets are discordant forthe development of PE during their own pregnanciessuggests that the fetal genotype, as well as environ-mental factors, may also be important in determiningsusceptibility.
There are several other lines of evidence that sug-gest a fetal (paternal) component to PE susceptibility.For instance, the association between PE and fetalchromosomal abnormalities supports a fetal contri-bution to etiology (29) as does the observation that
the risk of developing PE is increased in women withcomplete hydatidiform moles, which are entirely ofpaternal origin (30). In addition, the small but statis-tically significant increase in incidence amongdaughters-in-law of index cases (24) and the obser-vation that men born of preeclamptic pregnancieswere more likely to father a preeclamptic pregnancythan their matched controls (31) also support the ideaof a fetal/paternal contribution to risk. Lastly, mul-tiparous women who change partners are at increasedrisk of PE, especially if their new partner is known tohave fathered a preeclamptic pregnancy with anotherwoman (32).
Although the role of genetic factors in the etiologyof PE is widely accepted, the mode of inheritance isstill the subject of vigorous debate. Some researchershave suggested that susceptibility to PE could beinherited via a single (usually maternal) autosomalrecessive gene (23, 24, 33–35) or a dominant genewith incomplete penetrance (10, 24, 35, 36). A rolefor the fetal genotype has also been hypothesized(26, 24). It has been suggested that PE susceptibilityis due to complex interactions between two or morematernal genes, environmental factors and fetal ge-notypes (4, 27, 37, 38), a combination of maternal,fetal, and paternal (via fetus) genetic contributions(39, 40) or maternal-fetal interactions (8, 40).
Intuitively, the genetic model for a condition that isrestricted to pregnancy would include componentsfrom both the mother and the fetus. Therefore, it hasbeen suggested that the PE phenotype is due to amaternal-fetal genotype-by-genotype interaction ei-ther at the same locus or at separate ones. It isprobable that susceptibility to PE is due to one ormore genes, acting in both the mother and her fetus,modified by various environmental factors (41). Fivegenome-wide scans have provided indirect supportfor the hypothesis that PE is genetically heteroge-neous and is not inherited in a simple Mendelianmanner (42–46). Only one scan has identified a locus(on chromosome 2p) meeting the criteria for ge-nome-wide statistical significance; however, this re-sult was generated largely by two Icelandic families(24) and has not been confirmed in other populations.Although several other loci have been identified thatare suggestive of linkage, little overlap has beenobserved from study to study. Such inconsistency isnot surprising and is even to be expected when con-ducting genome-wide scans for complex traits.
Because it is likely that no one major gene deter-mines PE risk and because most genetic studies ofpreeclampsia to date have concentrated on the ma-ternal genotype alone (31, 38), more research needs
42 Obstetrical and Gynecological Survey
to be conducted examining both the maternal andfetal genotypes and possible maternal-fetal genotypicinteractions at multiple loci.
CANDIDATE GENES
It seems likely that no one gene can account for allof the genetic risk in all women. More likely, poly-morphisms in a number of genes can affect PE risk.The specific gene(s) involved may depend, at least inpart, on the characteristics of the population beingstudied (e.g., ethnicity, severity of the disease, ma-ternal age, or gestational age at onset). As is illus-trated in Figure 1, genetic predisposition could beinvolved in any aspect of PE etiology: immune mal-adaptation, placental ischemia, or oxidative stress.Genes involved in blood pressure regulation, placen-tation, and vascular remodeling/injury may be in-volved in causing placental ischemia, whereas genesin the endothelial cell health category may lead tooxidative stress. A summary of candidate genes, theirproposed mechanism of action, and previously pub-lished allele frequencies appears in Table 1.
Blood Pressure Regulation
Genes that encode aspects of the renin-angiotensinsystem (RAS) (Fig. 2) seem to be good candidatesfor involvement in PE etiology due to their role inregulating blood pressure, body-fluid volume, andvascular remodeling during pregnancy. Because theplacenta has no autonomic nerve supply, it dependson humoral factors, such as angiotensin II to regulatevascular resistance. Briefly, renin, an enzyme presentin the uteroplacental unit, converts angiotensinogen(produced in the liver) into angiotensin I. Next, an-giotensin-converting enzyme (ACE) converts angio-tensin I to angiotensin II, a powerful vasoconstrictor.Increased sensitivity to angiotensin II as well asincreased angiotensin II type I receptor (AT1) ex-pression in placental tissue could lead to placentalischemia and play a part in the development of PE. Itis also possible, however, that the effects of the RAScould be a secondary response to some primary ma-ternal or fetal stimulus.
Angiotensinogen. One coding region polymor-phism, resulting in a substitution of threonine formethionine at amino acid 235, has been extensivelystudied (Table 2). The frequency of the variant T235allele varies greatly by ethnicity, occurring in ap-proximately 40% of whites, 71% of Hispanics, 75%of Asians, 75% of African Americans, and �90% ofAfricans (47). In addition, polymorphisms have been
identified in the 5' and 3' flanking regions. A com-mon variant, G(-6)A, in the proximal promoter re-gion has been reported to affect the basal transcrip-tion rate and the A(-6) allele is in complete linkagedisequilibrium with the T235 variant (48). Two otherpolymorphisms at 20 base pairs (bp) and 18 bpupstream of the initiation site are also in linkagedisequilibrium with M235T. All of these polymor-phisms have been associated with essential hyperten-sion but because they are linked, it is difficult todetermine which might be functionally important.Although a coding region variant such as the T235variant would not be expected to increase AGT levelson its own, linkage with a promoter mutation such asG(-6)A could explain observed associations betweenincreased AGT levels and the T235 polymorphism.Finally, there is also a highly polymorphic CA-repeatsite in the 3' flanking region of the AGT gene. Thusfar, 11 alleles have been identified and are designatedA1 to A11, with the most common allele being A7(41).
Evidence for a role for AGT variants in the devel-opment of PE comes from several sources. In a studyof decidual spiral artery cross-sections from normalpregnancies terminated at 8 weeks’ gestation, womenhomozygous for the T235 allele had a statisticallysmaller external diameter and greater area-to-diame-ter ratio than woman homozygous for the M235allele (49). This strongly suggests that T235 homozy-gotes have abnormal spiral artery remodeling, a hall-mark sign of PE. Additionally, T235 expression inheterozygous women was highly statistically signif-icantly elevated in decidual spiral arteries relative tothe M235 allele. This observation lends support tothe hypothesis that a promoter polymorphism in link-age disequilibrium with the T235 variant is in factresponsible for the observed elevation of AGT levelsseen in women with the T235 variant.
Linkage analyses using the CA-repeat in the 3'flanking region have shown mixed results. One studyreported no evidence of co-segregation between PEand the repeat (50), although another found evidenceof linkage (51).
Epidemiological evidence is also mixed, varyingsubstantially by population studied. Several studiesin white (52) and Japanese women (52–55) havefound a statistically significant increase in the fre-quency of the T235 variant among women with PEcompared with normotensive controls. However, thetwo studies conducted by Kobashi and colleagues(54, 55) seem to include some of the same subjects.In particular, the more recent study (55) included amore restrictive study population and as such, is the
Molecular Epidemiology of Preeclampsia Y CME Review Article 43
TABLE 1 Genes hypothesized to play a role in development of PE
GeneHigh-Risk
AlleleAllele Frequency*
(%) Proposed Mechanism for Increased Risk of PESelected
References
AGT T235 35–73% Elevated AGT levels lead to atherotic changes and abnormal spiralartery remodeling
(52)(56)(48)
Renin Unknown NA Increased renin activity leads to increased angiotensin II andplacental vascular contraction
(65)(63)
ACE Deletion 19–29% Increased ACE activity leads to increased angiotensin II andplacental vascular contraction
(68)(69)
(181)(66)
AT1 T573 47% Reduced placental expression may lead to impaired prostaglandinsecretion, inadequate dilation and ischemia or, low receptorexpression may lead to low trophoblast responsiveness to an-giotensin II and impaired placentation
(71)G1062 11%C1166 27%
A4 Repeat 46%ET-1 Asn198 22.8% Increased ET-1 could lead to increased vasoconstriction (76)
(75)ER� C (codon 10) 45% Increased ER� could upregulate gene expression for vasoactive
substances, thereby increasing vasocontraction(77)
C (codon 87) Very rareENOS Asp298 9–13% May be linked to a polymorphism which causes reduced eNOS
activity and a lack of vasodilation(82)(83)
Prothrombin A20210 2.5–4.1% Increased concentrations of prothrombin leads to increased risk ofthrombosis
(94)(93)
FVL FVL� 0.6–7% Predisposes to thrombophilic events and could lead to placentalinfarctions
(103)(101)
MTHFR T677 10–30% Reduced MTHFR activity increases levels of homocysteine, whichthen causes vascular injury
(9)(112)
CBS 844ins68 0–37.7% CBS deficiency may increase plasma homocysteine and lead tovascular injury
(123)(122)
GPIIIa T98 (33Pro) 13.9–19.1% Loss of functional GPIIIa could increase the risk of thrombosis andpossibly PE
(97)
MMP1 2G 30–43% Reduced MMP1 could lead to inadequate early interstitium remod-eling leading to inadequate trophoblast invasion
(128)
EPHX Tyr113 36% Fast hydrolysis of oxides may produce toxic intermediates whichcause endothelial cell dysfunction
(129)Arg139 19%
LPL S291 1.5% Reductions in LPL activity lead to dyslipidemia and endothelial celldysfunction
(130)N9/�93G 0.7% (131)
S447 9.4%SOD1 Not known NA Decreased activity leads to increased superoxides and oxidative stress (138)(CuZn-SOD) (137)LCHAD C1528 0.15% Impaired oxidation of long-chain fatty acids leads to triglyceride ac-
cumulation and endothelial cell injury(142)(141)
ApoE �4 8.4–18.7% Increased dyslipidemia leads to increased oxidative stress (146)(143)(145)
HLA-G T107 8.8% Impaired immune tolerance leads to rejection of baby by mother orvice versa. May also be important in adequate trophoblast in-vasion
(148)A110 8.1% (40)
Deletion 62% (150)HLA-DR� NA NA Impaired immune tolerance leads to rejection of baby by mother or
vice versa(40)
HLA-DR4 DR4 15–28% Impaired immune tolerance leads to rejection of baby by mother orvice versa
(155)Present (154)
(40)TNF-� TNFA-2 11–23% Higher TNF-� secretion may lead to higher levels of thromboxane,
causing vasoconstriction. TNF-� is important in immune func-tion and may also alter endothelial cell function via its ability togenerate reactive oxygen species.
(167)(165)(160)
IL-1� �511T 58.9% Increased inflammatory response may allow for rejection of fetus bymother or vice versa
(169)(promoter) (170)E2 (exon 5) 18.2%
IL-1RA IL-1RA*2 5.1–29% Increased inflammatory response may allow for rejection of fetus bymother or vice versa
(169)(170)
IGF-II B allele 39–42% Overexpression may have negative effects on intrauterine fetal growth (175)(Apa I) (in PE mothers,
fathers & children)(173)(151)
Mitochon-drial DNA
NA NA Defective mitochondria result in decreased energy supply and inad-equate trophoblast invasion
(176)(177)(180)
* Allele frequency in the normal population.
44 Obstetrical and Gynecological Survey
more rigorous of the two. Additionally, Morgan andcolleagues (41) found that the A9 allele of the CArepeat is transmitted from the mother to the fetussignificantly more frequently than would be expectedby chance, suggesting that maternal-fetal allele shar-ing might be involved in PE. Of the studies reportinga positive association between PE and AGT geno-type, two have either failed to provide details of thePE definition used or they included women withnonproteinuric hypertension, suggesting that the ob-served association could be due to the known effectof AGT genotype on chronic hypertension. The as-sociation between AGT genotype and PE was notsupported by several other studies in white(41, 56, 58), Japanese (57), Chinese (58, 59), andHispanic (60) populations.
The lack of conclusive findings can be attributed toa number of common problems encountered in thestudy of PE. For example, various PE definitions areused, making it difficult to compare results acrossstudies. Lack of adjustment for parity is anotherpossible explanation for inconsistent results. Lastly,differences in exclusion criteria can result in studiesthat are not comparable because the study popula-tions can be quite different. Although some research-ers do not seem to have excluded any subjects apriori (41, 56, 58), others have quite an extensive listof exclusion criteria (52) (Table 2).
Renin. Renin is expressed in the first trimesterdecidua (61) as well as in the chorionic villi (62),suggesting that it may be important in pregnancydevelopment. Renin gene expression in the deciduavera (DV) but not in the decidua basalis (DB) orchorionic villi (CV) portions of the placenta has beenobserved to be higher among those with preeclamp-sia than the controls (63). Additionally, the presenceof renin mRNA in the fetal side of the placentaindicates that the fetal genotype may be important indetermining risk of PE.
Although several restriction fragment length poly-morphisms are now known (BglI, TaqI, and HinfI)(64), no one has yet examined them in an associationstudy. An earlier linkage study found no evidence oflinkage or distorted allele distribution using a dial-lelic genomic probe containing exon 1 (65). How-ever, several limitations of this study may have con-tributed to the lack of positive findings. First, thestudy did not require proteinuria as part of the PEdefinition and may have included some women withconditions other than PE (e.g., essential or gesta-tional hypertension), creating a heterogeneous popu-lation. Additionally, this study was likely to have lowpower because of the limited sample size and the useof a diallelic probe.
Angiotensin-converting enzyme. An insertion-dele-tion polymorphism in intron 16 is associated with
Fig. 2. The renin-angiotensin system.
Molecular Epidemiology of Preeclampsia Y CME Review Article 45
changes in angiotensin-converting enzyme (ACE)activity. Subjects homozygous for the deletion (DD)have the highest ACE levels and subjects homozy-gous for the insertion (II) have the lowest levels (66).The frequency of this polymorphism varies signifi-cantly by ethnic group with 29% of African Ameri-cans, 19% of Indians (Indian or Pakistani), and 29%of whites being homozygous for the deletion (67).
There is some evidence that serum ACE activitymay be higher in women with pregnancy-inducedhypertension (PIH), with and without proteinuria,compared with normotensive pregnancies (68, 69).However, there was no evidence for an associationbetween the insertion-deletion polymorphism and PEin either maternal or fetal samples from a Britishpopulation (70) or in maternal samples from a Finn-ish population (71). Both studies included nullipa-rous and multiparous women. Additionally, bothstudies lacked the statistical power to detect the rel-atively small difference between PE cases and con-trols that would be expected given our understandingof genetic risk factors.
Angiotensin II type 1 receptors. Angiotensin II type1 (AT1) receptors, which are found in the resistancevessels of full-term placentas and are most probablyencoded by the fetal genome (72), may contribute tofetoplacental blood flow regulation and may impactplacental perfusion (72–74). Upregulation of placen-tal AT1 receptor activity has been found amongwomen with PE compared with their normotensivecounterparts (75). Because AT1 receptors coupledwith angiotensin II are strong vasoconstrictors, itseems reasonable that upregulation of these receptorscould be involved in the pathophysiology of PE. Incontrast, Morgan and colleagues (72) have suggestedthat a reduction in AT1 receptors or receptor activitycould lead to decreased trophoblast responsiveness toangiotensin II as well as reduced prostaglandin se-cretion, inadequate vasodilation, placental ischemia,and PE.
Morgan et al. (72) examined two diallelic nonfunc-tional polymorphisms in the coding region (C573Tand A1062G), one in the 3' untranslated region(A1166C) and one dinucleotide repeat in the 3' flank-ing region. Allele frequencies for the “at-risk” alleleswere 47% for T573, 11% for 1062G, 27% for 1166C,and 27% for the A4 repeat polymorphism in a sampleof healthy British women (72). Although Morganand colleagues (72) failed to find any differences infetal or maternal AT1 receptor allele or genotypefrequencies between PE cases and controls, they didfind statistically significant distortions in maternal-fetal transmission for both the dinucleotide repeat
(A4 allele) and the C573T (T573) polymorphisms,suggesting that these nonfunctional polymorphismsmay be in linkage disequilibrium with a functionalsite. However, 11% of the controls and nearly 20% ofthe cases were multigravid. The inclusion of womenwith previous pregnancies is significant becausewomen who are multigravida with previous PE mayhave a different mechanism of disease than womenwho are primipara. Thus, any association found be-tween the AT1 receptor polymorphism and PE, de-fined to include multigravid women, could in fact bean association with an underlying maternal conditionrather than PE itself.
Endothelin-1. Endothelin-1 gene (ET-1) is a vaso-constrictor expressed in the endothelium and smoothmuscle. ET-1 levels have been found to be increasedamong women with PE (76), but it is uncertainwhether the increase occurs before or after develop-ment of disease.
A substitution of asparagine for lysine at codon198 (Lys198Asn) in the ET-1 gene has been found tobe associated with higher resting blood pressuresamong overweight people (77). Barden et al. (76)found no association between the polymorphism andPE. However, in both the normotensive and pre-eclamptic groups, women with at least one variant(Asn198) allele had increased systolic blood pressurecompared to the wild type homozygotes (Lys198/Lys198). Additionally, pregnant women homozy-gous for the variant allele (Asn198/Asn198) had in-creased ET-1 levels. Thus, although the Lys198Asnpolymorphism seems to be associated with both ET-1levels and systolic blood pressure, it does not appearto directly affect PE risk. However, more research isneeded to clarify if fetal genotype, in combinationwith maternal genotype, can impact the risk of PE.
Estrogen Receptor �. Estrogen receptor � (ER�)can regulate vascular tone and structure through itsability to induce gene expression of vasoactive sub-stances in vascular tissues (78). Polymorphisms inexons 1 and 2 encoding the NH2-terminal portion ofER� have been associated with various pathologies,including essential hypertension (79). Thus, it hasbeen hypothesized that variation in the NH2-terminalportion could be related to PE risk.
Malamitsi-Puchner and colleagues (78) are theonly group thus far to address the possible relation-ship between PE and the ER� gene. In a very smallsample of preeclamptic women (n � 16) and preg-nant controls (n � 20), they found two polymor-phisms, a T3C substitution in codon 10 and a G3Csubstitution in codon 87. Neither of these polymor-phisms alters the amino acid sequence nor was asso-
46 Obstetrical and Gynecological Survey
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ryof
Stu
dy
Typ
eof
Cas
es
Typ
eof
Con
trol
sP
arity
PE
Def
initi
on
Cov
aria
tes
(Ad
just
men
ts)
Exc
lusi
ons
Cas
es/C
ontr
ols
T235
Alle
le
Freq
uenc
y%
(Cas
es/C
ontr
ols)
P
(52)
Uta
h&
Jap
anIn
cid
ent
case
s
ofan
yty
pe
PIH
*
Nor
mot
ensi
ve
pre
gnan
t
cont
rols
Mul
tipar
ous
case
s&
con-
trol
s
Not
spec
ified
Non
e,b
utst
rati-
fied
onp
arity
Cad
aver
icre
nalt
rans
pla
ntre
-
cip
ient
,m
ultip
lep
regn
anci
es,
del
iver
yat
�20
wee
ksge
sta-
tion
(Uta
hca
ses)
;p
rem
atur
e
del
iver
y,te
rmin
fant
�25
00
gm,
dia
stol
icB
P�
85,
sys-
tolic
BP
�14
0,ch
roni
cH
T,
activ
ere
nald
isea
se(U
tah
cont
rols
)
149/
571
(Uta
h)50
.0/4
1.0
(Uta
h)�
.05
Mul
tiple
pre
gnan
cies
,hi
stor
yof
HT,
dia
bet
icne
phr
opat
hy,
rena
ldis
ease
(Jap
anes
e
case
s&
cont
rols
)
41/8
0(J
apan
)90
.0/7
1.0
(Jap
an)
.000
6
NO
TE:
Ana
lysi
sre
stric
ted
toU
tah
Whi
tes
&Ja
pa-
nese
case
sw
ithsi
mila
r
PE
def
initi
on
(56)
UK
Inci
den
tP
EN
orm
oten
sive
pre
gnan
t
cont
rols
Not
spec
ified
“Str
ict
crite
ria”
not
spec
ified
,
but
mus
tha
veb
een
norm
o-
tens
ive
bef
ore
wee
k20
and
by
6w
eeks
pos
tpar
tum
Non
eN
one
spec
ified
15/1
543
.0/4
7.0
�.0
5
(58)
Aus
tral
ia&
Chi
naS
ever
eP
Eor
E
case
s
“Nor
mal
mat
er-
nalc
ontr
ols”
Not
spec
ified
BP
�14
0/90
mm
Hg,
pro
tein
-
uria
(�0.
3g/
L/24
h)&
gen-
eral
ized
edem
aaf
ter
20
wee
ks’
gest
atio
n
Non
eN
one
spec
ified
106/
81(A
ustr
alia
nW
hite
s)47
.0/3
8.0
(Aus
tra-
lia)
.08
72/4
8(C
hine
se)
78.0
/75.
0(C
hi-
nese
)
.53
(57)
Jap
anIn
cid
ent
case
s
ofP
IH
Nor
mot
ensi
ve
rem
aind
erof
coho
rt
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
elev
ated
over
two
visi
ts&
pro
tein
uria
(�0.
3g/
L/24
h)af
ter
wee
k
20,
&no
rmot
ensi
vew
ithno
pro
tein
uria
at6
wee
ksp
ost-
par
tum
Non
eH
isto
ryof
HT,
HT
bef
ore
wee
k
20,
dia
bet
icne
phr
opat
hy,
re-
nald
isea
se&
pro
tein
uria
in
early
pre
gnan
cy,
mul
tiple
pre
gnan
cies
,D
NA
sam
ple
unav
aila
ble
,su
bje
ctd
eliv
ered
atot
her
hosp
ital,
inco
mp
lete
med
ical
reco
rds,
hyp
erte
nsiv
e
6w
eeks
pos
tpar
tum
313
sub
ject
s;33
dev
el-
oped
PIH
72.2
(PE
only
)/
80.4
�.0
5
Not
e:If
nosi
gnifi
cant
pro
tein
-
uria
,su
bje
cts
wer
ecl
assi
fied
asG
H
(41)
UK
PE
case
s
(whi
te)
Nor
mot
ensi
ve
pre
gnan
t
cont
rols
(Whi
te)
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
inp
revi
-
ousl
yno
rmot
ensi
vew
oman
,
pro
tein
uria
(�0.
3g/
L/24
h
or�
�d
ipst
ick)
&no
rmot
en-
sive
3m
onth
sp
ostp
artu
m
Non
eN
one
spec
ified
43/8
4(M
ater
nal)
41/8
1(F
etal
)
Mat
erna
l:
48.0
/48.
0
Feta
l:48
.0/4
0.0
�.0
5
�.0
5
(54)
Jap
anC
ases
ofP
E,
ecla
mp
sia
or
tran
sien
tH
T
Nor
mot
ensi
ve
pre
gnan
t
cont
rols
Mul
tipar
ous
case
s&
con-
trol
s
�30
/15
mm
Hg
incr
ease
in
BP
over
aver
age
bef
ore
20
wee
ksor
BP
�14
0/90
afte
r
wee
k20
(&p
rote
inur
ia30
mg/
dL
or�
dip
stic
kfo
r
PE
/E)
Non
e,b
utst
rati-
fied
by
age,
grav
idity
&d
i-
agno
sis
(PE
/E
vs.
tran
sien
t
HT)
Mul
tiple
pre
gnan
cies
,ch
roni
c
HT,
HE
LLP
synd
rom
e,re
nal
dis
ease
s,d
iab
etes
,am
niot
ic
volu
me
abno
rmal
ities
,fe
tal
abno
rmal
ities
,H
Tor
pro
tein
-
uria
bef
ore
wee
k20
or4
wee
ksp
ostp
artu
m
115/
381
Prim
igra
vid
PE
/E
vs.
prim
igra
vid
cont
rols
:93
.0/
77.0
�.0
01
Molecular Epidemiology of Preeclampsia Y CME Review Article 47
ciated with PE. However, the extremely small samplesize may have precluded a statistically significantfinding.
Vascular Remodeling/Vascular Injury
Genes in this category affect aspects of normalvascular remodeling during pregnancy or cause vas-cular injury by increasing levels of homocysteine orby increasing the risk of a thrombotic event. Inade-quate vascular remodeling or vascular injury wouldlikely lead to placental ischemia, and possibly to anincrease in PE risk. Both fetal and maternal throm-bosis may predispose to placental thrombosis, andthus, the putative genes in this category may be ofeither fetal or maternal origin.
Endothelial NO Synthase. Nitric oxide (NO) in-creases in the placental tissue during normal preg-nancy, contributing to vasodilation, vascular remod-eling, and inhibition of platelet aggregation (80, 81).However, NO synthase (NOS) activity has beenshown to be decreased in the placentas of womenwith PE (82). Because the syncytiotrophoblasts (fe-tal) and the endothelium of blood vessels within themyometrium (maternal) both express endothelialNOS (eNOS), fetal and maternal eNOS genotypesmight both be important in determining nitric oxidesynthase activity (83).
A G3T transversion in exon 7 results in the re-placement of glutamic acid with aspartic acid atcodon 298; however, no functional change in eNOSactivity is yet known to result from this polymor-phism (84). The variant allele (T) has been found in9% to 13% of the general population and has beenshown to be associated with spastic angina (85),myocardial infarction (85), and essential hyperten-sion (86). In a sample of normal pregnant women,Savvidou et al. (87) found that maternal flow-medi-ated vasodilation (FMD) was 21% lower among ho-mozygotes for the variant allele (TT) than in womenhomozygous for the wild-type (GG) allele. Heterozy-gotes showed intermediate FMD. These results sug-gest that carriers of the exon 7 variant have dimin-ished NO bioactivity, possibly resulting in a loweredthreshold to PE development.
The results of linkage analyses have been contra-dictory and inconclusive. Whereas linkage analysesin Scotland, Iceland, and Australia have suggestedthat the region of chromosome 7q36 (that containsthe eNOS gene) may contain a PE susceptibilitylocus (4, 88). However, strong evidence against link-age to markers in and near the eNOS gene itself wasfound in this same Australian population (89). Nev-TA
BLE
2C
ont
inue
d
Ref
eren
ceC
ount
ryof
Stu
dy
Typ
eof
Cas
es
Typ
eof
Con
trol
sP
arity
PE
Def
initi
on
Cov
aria
tes
(Ad
just
men
ts)
Exc
lusi
ons
Cas
es/C
ontr
ols
T235
Alle
le
Freq
uenc
y%
(Cas
es/C
ontr
ols)
P
(55)
Jap
anP
rimip
arou
s
PE
case
s
Nor
mot
ensi
ve
pre
gnan
t
cont
rols
Prim
ipar
ous
case
s&
con-
trol
s
�30
/15
mm
Hg
incr
ease
inB
P
over
aver
age
bef
ore
20w
eeks
orB
P�
140/
90af
ter
wee
k20
&p
rote
inur
ia(�
30m
g/d
Lor
�d
ipst
ick)
Pre
pre
gnan
cyB
MI
�
24,
men
tally
stre
ss-
fulc
ond
ition
dur
ing
pre
gnan
cy,
salty
dis
hes
pre
ferr
ed
dur
ing
pre
gnan
cy
Mul
tiple
pre
gnan
cies
,re
nal
dis
ease
,d
iab
etes
,am
niot
ic
volu
me
abno
rmal
ities
,p
re-
exis
ting
HT,
feta
lab
norm
ali-
ties,
HT
orp
rote
inur
iab
e-
fore
wee
k20
or4
wee
ks
pos
tpar
tum
58/1
64U
nab
leto
calc
u-
late
from
dat
a
give
n
Not
e:M
ayin
clud
eso
me
of
the
sam
esu
bje
cts
as(5
4)
OR
�2.
5fo
rho
moz
y-
gous
T235
/T23
5ge
-
noty
pe
�.0
5
(60)
U.S
.M
exic
an/
Cen
tral
Am
eri-
can
His
pan
ic
PE
case
sW
omen
with
at
leas
t2
prio
r
norm
oten
sive
term
pre
g-
nanc
ies
Mul
tipar
ous
case
s&
con-
trol
s
�30
/15
mm
Hg
incr
ease
inB
P
over
aver
age
bef
ore
20w
eeks
orB
P�
140/
90&
pro
tein
uria
(�30
0m
g)
Non
esp
ecifi
ed(fo
r
asso
ciat
ion
bet
wee
n
dis
ease
stat
us&
geno
typ
e)
For
case
s:H
T&
pro
tein
uria
bef
ore
20w
eeks
gest
atio
n
orot
her
sign
ifica
ntm
edic
al
cond
ition
sth
atca
nca
use
hyp
erte
nsio
n&
pro
tein
uria
.
For
cont
rols
:es
sent
ialH
T,
chro
nic
rena
ldis
ease
,d
ia-
bet
es,
pla
tele
td
isor
der
sor
auto
imm
une
cond
ition
s
68/5
072
.0/7
0.0
.84
HT
�hy
per
tens
ion;
GH
�ge
stat
iona
lhyp
erte
nsio
n;B
P�
blo
odp
ress
ure;
BM
I�
bod
ym
ass
ind
ex.
*In
clud
esP
E(m
ildor
seve
re),
tran
sien
tH
T,ch
roni
cH
Tw
ithou
tsu
per
imp
osed
PE
,ch
roni
cH
Tw
ithsu
per
imp
osed
PE
,H
ELL
P,
und
erly
ing
auto
imm
une,
dia
bet
icor
othe
rne
phr
opat
hyan
ted
atin
gp
regn
ancy
,an
dso
me
whe
recl
inic
ald
ata
did
not
pro
vid
ead
equa
ted
iagn
ostic
crite
riato
mak
ea
det
erm
inat
ion.
48 Obstetrical and Gynecological Survey
ertheless, it should be noted that negative linkageresults do not necessarily exclude the possibility thateNOS is a susceptibility gene for PE. First, the mi-crosatellite marker (CA-repeat), for which strong ev-idence against linkage was found, is intronic and maynot be in linkage disequilibrium with potentiallycausal variants. Second, linkage analyses may not beable to identify the involvement of the eNOS genedue to the complex (multigenic/multifactorial) etiol-ogy of PE.
In association studies, the exon 7 variant has beenfound to be associated with PE. In a Japanese pop-ulation, the variant allele (genotypes TG � TT) wasfound more frequently among cases than controls(84). A second study of PIH reported similar results(90); however, this finding failed to reach statisticalsignificance when restricted to primiparous womenmeeting the more rigorous definition of PE, possiblydue to the reduction in sample size. Results usingintronic markers are more mixed. An eNOS intron 13CA-repeat was not associated with PE in either Chi-nese or Australian subjects (88). A 27 base pairrepeat polymorphism in intron 4, previously associ-ated serum NO levels and blood pressure amongnonpregnant women, was found to be associated withPE among Hispanic women (60). Additional researchis needed to clarify the role, if any, of the eNOS genein the development of PE.
Prothrombin. Placental infarctions and intervillousthrombosis are common events among women withPE, suggesting that genes predisposing to thrombo-philias are also candidates for susceptibility to PE.Specifically, in the prothrombin gene, a G3A mu-tation at nucleotide 20210 in the 3' untranslated re-gion is associated with higher plasma concentrationsof prothrombin and leads to increased risk of venousthromboembolism, myocardial infarction and cere-bral-vein thrombosis (91). Heterozygous carrier ratesamong controls who were white range from 2.5% to6%.
The epidemiologic data on whether this mutation isassociated with PE are mixed (Table 3). One study inItaly found a statistically significant association be-tween PE and the prothrombin gene mutation (92):11.4% of PE cases but only 4.1% of controls carriedthe mutation. This difference remained statisticallysignificant even after adjustment for maternal age,parity, factor V Leiden (FVL), and 5,10-methyl-enetetrahydrofolate reductase (MTHFR) genotypes.Similarly, investigators in Israel found that the pro-thrombin mutation was statistically significantly as-sociated with obstetric complications, including se-vere PE, but they did not have the power to detect an
association with PE alone (91, 93). In a later study,these same investigators reported a statistically sig-nificant association with severe PE, but only afteradjusting for other covariates (94).
In contrast, PE has not been found to be associatedwith the prothrombin mutation among Australian andNew Zealand women (95), women in the Nether-lands (96), white and African American women andtheir fetuses (97), or among a separate group of whitenorthern European women (98). However, the studyby Higgins et al. (95) was powered to find a three-fold increase in risk and any effect less than thatmight easily go undetected. Additionally, more thanhalf of the participants in the study by Livingston etal. (97) were of African American descent. Inasmuchas the prothrombin mutation is virtually nonexistentin nonwhite populations (98), any effect of this mu-tation would necessarily be restricted to whites. Al-though the authors might have simply stratified byethnicity, this would result in very small numbers ofsubjects in each ethnic group, thereby limiting thepower to detect an association.
FVL Mutation. The Leiden mutation is a G3Asubstitution at nucleotide position 1691 in exon 10 ofthe factor V gene. It results in the replacement ofglutamine for arginine at position 506 at the cleavagesite for activated protein C (APC) (99–101), causingactivated protein C resistance (APCR), retention ofprocoagulant activity and thus, increased susceptibil-ity to thrombophilic events (102). Pregnant womenwith APCR are at increased risk of PE (9). Addition-ally, placentas with greater than 10% placental in-farction have been associated with a more than 10-fold increase in fetal FVL carrier rate, suggesting thatfetal, as well as maternal, FVL genotype may be ofimportance (103). The frequency of the Leiden mu-tation ranges from 0.6% in Asia Minor to 7% amongGreeks, with a frequency among those in the UnitedKingdom of 4.4% (104).
The majority of epidemiological research also sug-gests an association between FVL and PE (Table 4).Studies of women in Hungary (105), Israel (91, 94),Italy (92, 102, 106), and Utah (107) have all found astatistically significant striking increase in risk of PEassociated with either being a homozygote or a het-erozygote carrier of FVL. In contrast, several studieshave failed to find an association (96, 97, 108–110).However, in a Japanese population, none of the casesor controls carried the FVL mutation (108). OneEuropean study that failed to find an associationfound a high FVL carrier rate among both cases(9.8%) and control women (9.2%) (96), suggestingthat selection bias among controls may be responsi-
Molecular Epidemiology of Preeclampsia Y CME Review Article 49
ble for the negative results. One U.S. study found astatistically nonsignificant elevation in carrier rateamong cases (110), although one European (96) andone American (97) study found no association at all.Additionally, Livingston and colleagues (96) exam-ined the possible role of fetal FVL mutation andfound no association. Although it seems clear thatFVL is associated with increased risk of thrombo-philia, the strength of its association with PE remainsto be clarified.
5,10-Methylenetetrahydrofolate Reductase. Increasedhomocysteine levels are known to cause vascular injury(111) and hyperhomocysteinemia has been reported inpreeclamptic patients (9). The mutation most com-monly reported to be associated with PE is a cytosine-to-thymine substitution at nucleotide 677, which resultsin an amino acid substitution (alanine3valine). Thetwo alleles are commonly referred to as C (wild type)and T (variant). The variant causes reduced 5,10-meth-ylenetetrahydrofolate reductase (MTHFR) activity andmodestly increases circulating homocysteine levels(109, 112). In a Los Angeles population, the frequencyof the TT genotype was highest among Hispanics with15.2% having the high-risk genotype, compared with10.2% among whites, 8.8% among Asians, and 2.4%among African Americans (113).
The epidemiologic data are mixed (Table 5). Stud-ies in Israel (91, 94), Japan (114), and Italy (106)have all found that the TT genotype is associatedwith a statistically significant increase in risk of PE.Interestingly, the PE risk conferred by the TT geno-type was highest among proteinuric nulliparae cases(92). In contrast, studies in the United States(97, 110, 115), Australia (116), the United Kingdom(109), Finland (109, 117), and the Netherlands (118)have all failed to find a direct association of theC677T mutation with PE in either maternal(110, 115, 119) or fetal (97) samples. It would beuseful to know what percentages of each of the casegroups used in this study were primiparous and tohave information on possible underlying maternalconditions.
A second variant, an adenine-to-cytosine substitu-tion at base 1298 (A1298C), has recently been de-scribed (120). The variant (C allele) also results indecreased MTHFR activity but is not associated withincreased plasma homocysteine or decreased plasmafolate (118). The frequency of the C allele has beenreported to be 33% in a European population (120).To date, only one study has examined the effect ofthe A1298C mutation on PE risk, finding no differ-ence in mutation frequency between PE cases andcontrols (118). However, the controls used in this
study were not screened for PE history, and thus,could have included some women with a history ofPE. The inclusion of such women in the controlgroup might have obliterated a true difference inmutation rate between women with normotensivepregnancy histories and those with a history of PE.
Because folate can decrease homocysteine levelsirrespective of MTHFR genotype, it is important toaddress the possibility of confounding due to folatestatus. Folate supplementation is more common inthe U.S. than in many other countries and is activelyencouraged in women of childbearing age. There-fore, differences in the folate status of the studypopulation could at least partially explain the differ-ing results obtained.
Cystathionine �-Synthase. Cystathionine �-syn-thase (CBS) is an enzyme that catalyzes the conden-sation of serine with homocysteine to produce cys-tathionine (121). It has been suggested that CBSdeficiency leads to high plasma levels of homocys-teine and might predispose to thrombophilic events(110). A T3C substitution at nucleotide 833 resultsin the substitution of threonine for isoleucine and hasbeen shown to be associated with mild hyperhomo-cysteinemia (110), but is relatively rare in the generalpopulation (�1%) (122). This mutation is known toco-segregate in cis with a common 68-bp insertion inthe coding region of exon 8 (844ins68) (123). Allelefrequencies for the insertion were 6.7% amongwhites, 0.45% among Amer-Indians, and 11%among blacks (123).
The increased incidence of PE among women withhyperhomocysteinemia has led to speculation about apotential role for CBS in PE development. Thus far,only one study has examined the role of the CBSinsertion mutation in relation to PE and has found noassociation with disease among nulliparous whitecarriers in the United States (110). Tsai and col-leagues (124) have provided a possible explanationfor the lack of association between the insertionpolymorphism and PE. They suggest that the muta-tion is not related to homocysteine levels because itcreates an alternate splicing site that eliminates theT833C polymorphic site (which is associated withhomocysteine levels) and produces a normal mRNAand CBS enzyme. This assertion is supported by thefinding that both heterozygous carriers of the inser-tion as well as homozygous subjects do not havelowered CBS enzyme activity (124) nor were plasmatotal homocysteine levels different between het-erozygous carriers and noncarriers (122).
�3 Integrin Glycoprotein IIIa. The �3 integringlycoprotein IIIa (GPIIIa) gene encodes the beta
50 Obstetrical and Gynecological Survey
TAB
LE3
Mo
lecu
lar
epid
emio
log
yst
udie
so
fth
eP
roth
rom
bin
G20
210A
po
lym
orp
hism
and
risk
of
PE
Ref
eren
ce
Cou
ntry
of
Stu
dy
Typ
eof
Cas
esTy
pe
ofC
ontr
ols
Par
ityP
ED
efin
ition
Cov
aria
tes
(Ad
just
men
ts)
Exc
lusi
ons
Cas
es/C
ontr
ols
P20
210A
Alle
leFr
eque
ncy
%
(96)
Net
her-
land
s
PE
hist
ory
Age
&d
eliv
ery
dat
e
mat
ched
;no
rmo-
tens
ive
pre
gnan
cy
hist
ory
Prim
ipar
ous
case
s&
cont
rols
�30
/15
mm
Hg
rise
in
BP
or�
BP
140/
90
mm
Hg
&p
rote
inur
ia
(�30
mg/
dL)
Fam
ilyhi
stor
yof
thro
mb
osis
orH
T,
bod
ym
ass,
smok
-
ing,
coag
ulat
ion
mut
atio
ns(F
VL�
)
Mul
tiple
pre
gnan
cies
,ch
roni
cH
T,
rena
ldis
ease
,d
iab
etes
,co
llage
n
vasc
ular
dis
ease
s,ca
ncer
,th
rom
-
bos
ishi
stor
y
163/
163
3.1/
3.7
�.0
5
(92)
Not
e.M
ayin
-
clud
eso
me
of
the
sam
esu
b-
ject
sas
(106
)
Italy
Inci
den
tG
H
(with
&w
ith-
out
pro
tein
-
uria
)
Nor
mot
ensi
vegr
avid
wom
enm
atch
ed
onet
hnic
ity
Mul
tipar
ous
case
s&
cont
rols
BP
�14
0/90
mm
Hg
on
2oc
casi
ons
�4
h
apar
t&
with
(n�
70)
orw
ithou
t(n
�70
)
pro
tein
uria
(�30
0
mg/
24h)
Mat
erna
lage
,p
arity
,
FVL
carr
ier
stat
us,
677T
MTH
FRho
-
moz
ygos
ity
Tran
sien
tH
T,d
iab
etes
,au
toim
mun
e
dis
ease
,ch
roni
cre
nalo
rp
ulm
o-
nary
dis
ease
&ch
roni
ces
sent
ial
HT
prio
rto
pre
gnan
cy
140/
216
11.4
/4.2
�.0
5
(91)
Isra
elIn
cid
ent
seve
re
PE
,A
P,
FGR
orst
illb
irth
Age
&ge
ogra
phi
cally
mat
ched
norm
o-
tens
ive
pre
gnan
cy
Mul
tipar
ous
case
s&
cont
rols
Sev
ere
PE
:B
P�
160/
110
mm
Hg,
pro
tein
-
uria
(�50
0m
g/24
h),
HE
LLP
orec
lam
psi
a
Non
e,b
utst
ratif
ied
by
dia
gnos
is
Var
ied
by
dia
gnos
is-n
one
spec
ified
for
seve
reP
E
110/
110
For
seve
reP
E
(n�
34):
5.9/
3.0
�.0
5
(95)
Aus
tral
ia&
New
Zea
land
Cas
esof
ecla
mp
sia
or
PE
(84/
189
of
case
sst
udie
d
wer
efr
om
mul
ticas
e
fam
ilies
)
Par
ous
wom
enw
ith
norm
oten
sive
pre
g-
nanc
yhi
stor
y
Not
spec
ified
�25
/15
mm
Hg
rise
in
BP
from
bas
elin
eor
BP
�14
0/90
mm
Hg
onat
leas
t2
occa
-
sion
s6
hap
art
&p
ro-
tein
uria
(�0.
3g/
l/24
h)
Non
eH
isto
ryof
chro
nic
HT,
dia
bet
esor
rena
ldis
ease
189/
119
3.6
(com
bin
ed
case
s)/2
.5
.73
(94)
Isra
elIn
cid
ent
seve
re
PE
Age
&et
hnic
ity
mat
ched
wom
en
with
�1
norm
al
pre
gnan
cy
Mul
tipar
ous
case
s&
cont
rols
BP
�16
0/11
0m
mH
g&
pro
tein
uria
(�50
0
mg/
24h)
,H
ELL
Por
ecla
mp
sia
Not
spec
ified
His
tory
ofth
rom
bop
hilic
even
t,H
T
inea
rlyp
regn
ancy
,m
ultip
lep
reg-
nanc
ies
63/1
268.
0/3.
0.1
4(u
n-
adj)
.03
(ad
j)
(93)
Not
e:In
clud
ed
90su
bje
cts
(con
-
trol
s)al
sod
e-
scrib
edin
(91)
Isra
elW
omen
with
inci
den
tob
-
stet
ricco
m-
plic
atio
ns*
Hea
lthy
wom
enw
ith
�1
norm
alp
reg-
nanc
y
Mul
tipar
ous
case
s&
cont
rols
Per
AC
OG
Non
eN
ohi
stor
yof
ath
rom
bop
hilic
even
t22
2/15
6Fo
rP
Eon
ly
(n�
80):
8.8/
3.2
�.0
5
(97)
U.S
.In
cid
ent
seve
re
PE
Nor
mot
ensi
ve
thro
ugho
utp
reg-
nanc
yw
ithno
his-
tory
ofP
E
Mul
tipar
ous
case
s&
cont
rols
(im-
plie
d)
BP
�16
0/11
0m
mH
g&
pro
tein
uria
(�30
0
mg/
24h)
,ec
lam
psi
aor
HE
LLP
Non
eC
hron
icH
T,d
iab
etes
,p
reex
istin
g
rena
ldis
ease
,hi
stor
yof
thro
mb
o-
emb
olis
m,
mul
tifet
alge
stat
ion,
maj
orfe
talc
onge
nita
lano
mal
y
Mat
erna
lsam
-
ple
s:
Mat
erna
lsam
-
ple
s:
110/
970/
1.0
�.0
5
Feta
lsam
ple
s:Fe
tals
amp
les:
75/9
21.
3/2.
2�
.05
(98)
Not
e:In
clud
es
200
sub
ject
s
(con
trol
s)&
may
also
incl
ude
som
eof
the
sam
eca
ses
as
des
crib
edin
(109
)
UK
Inci
den
tP
EA
ge-m
atch
edno
rmo-
tens
ive
pre
gnan
t(n
�10
0)&
norm
o-
tens
ive
nonp
reg-
nant
with
noP
E/
PIH
hist
ory
&ag
e
�50
atsc
reen
(n
�10
0)
Mul
tipar
ous
case
s&
cont
rols
BP
�14
0/90
mm
Hg
afte
rw
eek
20w
ithat
leas
t25
mm
Hg
in-
crea
sein
dia
stol
icB
P
&p
rote
inur
ia(�
300
mg/
24h
or�
�d
ip-
stic
k);
reso
lved
by
3
mon
ths
pos
tpar
tum
Non
eM
ultip
leb
irths
,d
iab
etes
,re
nald
is-
ease
,es
sent
ialH
T
356/
200
2.0/
1.1
�.0
5
HT
�hy
per
tens
ion;
GH
�ge
stat
iona
lHT;
FVL
�fa
ctor
VLe
iden
mut
atio
n;A
P�
abru
ptio
pla
cent
ae;
FGR
�fe
talg
row
thre
tard
atio
n.*
Ob
stet
ricco
mp
licat
ions
incl
ude:
seve
reP
E,
intr
aute
rine
grow
thre
tard
atio
n,se
vere
abru
ptio
pla
cent
ae,
unex
pla
ined
still
birt
h,se
cond
trim
este
rlo
ss,
and
thre
eor
mor
eco
nsec
utiv
esp
onta
neou
sab
ortio
ns.
Molecular Epidemiology of Preeclampsia Y CME Review Article 51
TAB
LE4
Mo
lecu
lar
epid
emio
log
yst
udie
so
fth
efa
cto
rV
Leid
enm
utat
ion
and
risk
of
PE
Ref
eren
ce
Cou
ntry
of
Stu
dy
Typ
eof
Cas
esTy
pe
ofC
ontr
ols
Par
ityP
ED
efin
ition
Cov
aria
tes
(Ad
just
men
ts)
Exc
lusi
ons
Cas
es/C
ontr
ols
FVL
Car
rier
Rat
e%
P
(107
)U
tah
Inci
den
tse
vere
PE
Nor
mot
ensi
vete
rmgr
avid
wom
en
Not
spec
ified
Sus
tain
edH
T(B
P�
160/
110
mm
Hg)
&p
rote
inur
ia(�
5
ggm
/24
hor
3�or
4�)
Non
eP
reex
istin
gH
T,re
nald
isea
se,
HE
LLP
synd
rom
e
158/
403
8.9/
4.2
.03
(106
)Ita
lyIn
cid
ent
PIH
Hea
lthy
par
ous
pre
gnan
t
cont
rols
;m
atch
edon
ethn
icity
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
onat
leas
t2
occa
sion
s�
4h
apar
t(n
�51
)
Non
eTr
ansi
ent
HT,
dia
bet
es,
chro
nic
re-
nalo
rp
ulm
onar
yd
isea
se,
chro
nic
esse
ntia
lHT
bef
ore
pre
g-
nanc
y(p
ossi
bly
excl
uded
in
case
son
ly)
95/1
2810
.5/2
.3.0
2
(105
)H
unga
ryIn
cid
ent
seve
re
PE
Hea
lthy
NP
(n�
58)
&
heal
thy
P(n
�71
)
Not
spec
ified
,b
ut
mul
tipar
ous
imp
lied
BP
�16
0/90
mm
Hg
&p
ro-
tein
uria
(100
0m
g/24
h)
Non
eN
one
spec
ified
69/1
2918
.8/7
.0(P
)
&5.
2
(NP
)
�.0
5
(108
)Ja
pan
Inci
den
tP
IHN
orm
oten
sive
pre
gnan
t
cont
rols
Prim
ipar
ous
case
s&
con-
trol
s
One
orm
ore
ofth
efo
llow
ing:
(1)
30/1
5m
mH
gin
crea
se
inB
Pfr
omb
asel
ine
(2)
BP
�14
0/90
mm
Hg
or(3
)
pro
tein
uria
(1�
)
Non
eM
ultip
leb
irths
,re
nald
isea
se,
dia
be-
tes,
amni
otic
volu
me
abno
rmal
i-
ties,
pre
exis
ting
HT,
feta
lano
ma-
lies
71/1
090/
0N
A
(102
)Ita
lyP
EH
isto
ryH
ealth
yw
omen
with
nor-
mal
pre
gnan
cyhi
stor
y;
mat
ched
for
age,
gra-
vid
ity&
par
ity
Mul
tipar
ous
case
s&
con-
trol
s
Sus
tain
edH
T(B
P�
140/
90
mm
Hg)
&p
rote
inur
ia(�
0.3
g/l/2
4h)
Non
eN
ohi
stor
yof
HT
bef
ore
wee
k20
in
prio
rp
regn
anci
es,
dia
bet
es,
not
taki
nges
trop
roge
stin
icp
rep
ara-
tions
46/8
026
.1/3
.8�
.04
All
sub
ject
sw
ere
whi
te
(109
)U
KIn
cid
ent
PE
Age
-mat
ched
norm
oten
-
sive
Pco
ntro
ls(n
�
100)
&no
rmot
ensi
ve
NP
cont
rols
with
no
PE
hist
ory
(n�
100)
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
afte
r20
wee
ksw
ithat
leas
t25
mm
Hg
rise
ind
iast
olic
BP
&
pro
tein
uria
(�30
0m
g/24
h
or�
�d
ipst
ick)
;re
solv
ed
by
3m
onth
sp
ostp
artu
m
Non
eM
ultip
leb
irths
,co
ncur
rent
dia
bet
es,
rena
ldis
ease
,es
sent
ialH
T
283/
200
5.3/
5.5
(poo
led
cont
rols
)
�.0
5
(96)
Net
her-
land
s
PE
hist
ory
Age
&d
eliv
ery
dat
e
mat
ched
;no
rmot
en-
sive
pre
gnan
cyhi
stor
y
Prim
ipar
ous
case
s&
con-
trol
s
�30
/15
mm
Hg
rise
inB
Por
�B
P14
0/90
mm
Hg
&
pro
tein
uria
(�30
mg/
dL)
Fam
ilyhi
stor
yof
thro
mb
osis
orH
T,
bod
ym
ass,
smok
-
ing,
coag
ulat
ion
mut
atio
ns(P
ro-
thro
mb
inG
2021
0A)
Mul
tiple
pre
gnan
cies
,ch
roni
cH
T,
rena
ldis
ease
,d
iab
etes
,co
llage
n
vasc
ular
dis
ease
s,ca
ncer
,
thro
mb
osis
hist
ory
163/
163
9.8/
9.2
�.0
5
(92)
Not
e:
May
in-
clud
e
som
eof
the
sam
e
sub
ject
s
as(1
06)
Italy
Inci
den
tG
H
(with
&w
ith-
out
pro
tein
-
uria
)
Nor
mot
ensi
vegr
avid
wom
enm
atch
edon
ethn
icity
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
on2
occa
sion
s�
4h
apar
t&
with
(n�
70)
orw
ithou
t(n
�70
)p
rote
inur
ia(�
300
mg/
24h)
Mat
erna
lage
,p
arity
,
pro
thro
mb
in
A20
210
carr
ier
sta-
tus,
677T
MTH
FR
hom
ozyg
osity
Tran
sien
tH
T,d
iab
etes
,au
toim
mun
e
dis
ease
,ch
roni
cre
nalo
rp
ulm
o-
nary
dis
ease
&ch
roni
ces
sent
ial
HT
bef
ore
pre
gnan
cy
140/
216
7.9/
1.9
�.0
5
(91)
Isra
elIn
cid
ent
seve
re
PE
,A
P,
FGR
orst
ill-
birt
h
Age
&ge
ogra
phi
cally
mat
ched
norm
oten
sive
pre
gnan
cy
Mul
tipar
ous
case
s&
con-
trol
s
Sev
ere
PE
:B
P�
160/
110
mm
Hg,
pro
tein
uria
(�50
0
mg/
24h)
,H
ELL
Por
ecla
mp
sia
Non
e,b
utst
ratif
ied
by
dia
gnos
is
Var
ied
by
dia
gnos
is—
none
spec
ified
for
seve
reP
E
110/
110
For
seve
re
PE
(n�
34):
26.5
/6.0
�.0
5
52 Obstetrical and Gynecological Survey
subunit of the GP IIb/IIIa and �v�3 complexes thatbelong to a class of receptors that bind cell adhesionmolecules. The GP IIb/IIIa receptor is found only inplatelets and megakaryocytes and is important inplatelet aggregation. The �v�3 integrin is expressedby invading trophoblasts, suggesting it may be im-portant in placentation. Additionally, the �3 integrinhas been implicated in the failure of the cytotropho-blasts to adopt a vascular phenotype in women withPE (125).
A coding variant (C98T) in exon 2 of the GPIIIagene creates two antigenically distinct forms of themature GPIIb/IIIa antigen on platelets (Pl(A) anti-gens 1 and 2) (98). The variant (T98) has beenassociated with risk of premature acute coronarysyndromes and strokes in young white women (126).One association study has examine the role of thevariant in PE and found a statistically significantexcess of homozygotes (T98/T98) among whitenorthern European women with PE (98). These re-sults need to be confirmed in other studies and inother ethnic groups.
Matrix Metalloproteinase-1. Matrix metallopro-teinase-1 (MMP1) is produced by decidual, endothe-lial and trophoblastic cells and is involved in theprocess of interstitium remodeling and degradingcollagens I, II, III, VII, and X (127). Interstitiumremodeling occurs very early in pregnancy, beforetrophoblast invasion, and is a necessary part of thenormal arterial changes that occur during pregnancy.Among women with PE, the amount of MMP1 indecidual artery endothelial cells is reduced in com-parison to normotensive pregnant women, potentiallyexplaining the inhibited vascular invasion by cytotro-phoblasts in preeclamptics (128).
An insertion polymorphism (1G/2G) at position–1607 in the MMP1 gene creates an Ets binding siteand is associated with higher transcriptional activity(2G) (129). A single study has examined the possiblerole of this polymorphism in PE etiology, but foundno increase in PE, eclampsia, or PIH risk associatedwith the variant allele (127). However, because thesample size was small (58 cases), the failure to findan association may have been due to lack of power.Moreover, the study included primarily multiparouswomen who had a wide range of hypertensive preg-nancy disorders, creating a population potentially tooheterogeneous to find an effect. Lastly, fetal MMP1genotype could be important in the remodeling pro-cess, but was not considered in the single studyexamining MMP1 genotype and risk of PE.TA
BLE
4C
ont
inue
d
Ref
eren
ce
Cou
ntry
of
Stu
dy
Typ
eof
Cas
esTy
pe
ofC
ontr
ols
Par
ityP
ED
efin
ition
Cov
aria
tes
(Ad
just
men
ts)
Exc
lusi
ons
Cas
es/C
ontr
ols
FVL
Car
rier
Rat
e%
P
(94)
Isra
elIn
cid
ent
seve
re
PE
Age
&et
hnic
itym
atch
ed
wom
enw
ith�
1no
r-
mal
pre
gnan
cy
Mul
tipar
ous
case
s&
con-
trol
s
BP
�16
0/11
0m
mH
g&
pro
-
tein
uria
(�50
0m
g/24
h),
HE
LLP
orec
lam
psi
a
Not
spec
ified
His
tory
ofth
rom
bop
hilic
even
t,H
T
inea
rlyp
regn
ancy
,m
ultip
lep
reg-
nanc
ies
63/1
2624
.0/6
.30.
001
(un-
adj)
�.0
5(a
dj)
(110
)Io
wa
&
Nor
th
Car
o-
lina
Inci
den
tP
EH
isto
ryof
�2
norm
oten
-
sive
term
pre
gnan
cies
with
out
HT
orfa
mily
hist
ory
ofP
E
Mul
tipar
ous
case
s&
con-
trol
s
Mild
PE
:B
P�
140/
90m
mH
g
&p
rote
inur
ia(�
300
mm
/
24-h
or�
dip
stic
k)
Non
e,b
utst
ratif
ied
on
par
ity,
PE
seve
rity
&H
ELL
Pst
atus
Non
e(e
xcep
tas
liste
dfo
rco
ntro
ls)
281
/360
3.0/
2.4
(for
allP
E
case
s
com
-
bin
ed)
.53
(97)
U.S
.In
cid
ent
Sev
ere
PE
Nor
mot
ensi
veth
roug
hout
pre
gnan
cyw
ithno
his-
tory
ofP
E
Mul
tipar
ous
case
s&
con-
trol
s(im
plie
d)
BP
�16
0/11
0m
mH
g&
pro
-
tein
uria
(�30
0m
g/24
h),
ecla
mp
sia
orH
ELL
P
Non
eC
hron
icH
T,d
iab
etes
,p
reex
istin
g
rena
ldis
ease
,hi
stor
yof
thro
m-
boe
mb
olis
m,
mul
tifet
alge
stat
ion,
maj
orfe
talc
onge
nita
lano
mal
y
Mat
erna
l
Sam
ple
s:
Mat
erna
l
Sam
ple
s
110
/97
2.7/
1.5
�.0
5
Feta
l
Sam
ple
s:
Feta
l
Sam
ple
s:
75/9
22.
0/0
�.0
5
HT
�hy
per
tens
ion;
GH
�ge
stat
iona
lHT;
BP
�b
lood
pre
ssur
e;P
�p
regn
ant;
NP
�no
npre
gnan
t;A
P�
abru
ptio
pla
cent
ae;
FGR
�fe
talg
row
thre
tard
atio
n.
Molecular Epidemiology of Preeclampsia Y CME Review Article 53
TAB
LE5
Mo
lecu
lar
epid
emio
log
yst
udie
so
fth
eM
TH
FRC
677T
po
lym
orp
hism
and
risk
of
PE
Cas
es/C
ontr
ols
Ref
eren
ce
Cou
ntry
of
Stu
dy
Typ
eof
Cas
esTy
pe
ofC
ontr
ols
Par
ityP
ED
efin
ition
Cov
aria
tes
(Ad
just
men
ts)
Exc
lusi
ons
TT
Hom
ozyg
osity
%P
(106
)Ita
lyIn
cid
ent
PIH
Hea
lthy
par
ous
Pco
n-
trol
s;m
atch
edon
eth-
nici
ty
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
onat
leas
t2
occa
sion
s�
4h
apar
t(n
�51
)
Plu
sp
rote
inur
ia(�
300
mg/
24
hr)
(n�
45)
Non
eTr
ansi
ent
HT,
dia
bet
es,
chro
nic
re-
nalo
rp
ulm
onar
yd
isea
se,
chro
nic
esse
ntia
lHT
prio
rto
pre
gnan
cy(p
ossi
bly
excl
uded
in
case
son
ly)
94/1
2929
.8/1
8.6
.05
(114
)Ja
pan
PE
case
s“M
atch
ed”
heal
thy
Pco
n-
trol
s(n
�98
)&
nor-
mot
ensi
vehe
alth
yN
P
cont
rols
(n�
260)
Not
spec
ified
Acc
ord
ing
toA
CO
GN
one
Non
e67
/358
24.0
/11.
0
(all
con-
trol
s)
�.0
04
(109
)U
KIn
cid
ent
PE
Age
-mat
ched
norm
oten
-
sive
Pco
ntro
ls(n
�
100)
&no
rmot
ensi
ve
NP
cont
rols
with
no
PE
hist
ory
&ag
e�
50
atsc
reen
(n�
100)
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
afte
r20
wee
ksw
ithat
leas
t25
mm
Hg
rise
ind
iast
olic
BP
&
pro
tein
uria
(�30
0m
g/24
h
or�
�d
ipst
ick)
;re
solv
ed
by
3m
onth
sp
ostp
artu
m
Non
eM
ultip
leb
irths
,co
ncur
rent
dia
bet
es,
rena
ldis
ease
,es
sent
ialH
T
283/
200
11.0
/11.
5
(all
con-
trol
s)
�.0
5
(115
)P
enns
yl-
vani
a
Inci
den
tP
E(n
�99
)&
In-
cid
ent
GH
(n�
24)
Nor
mot
ensi
veP
cont
rols
Prim
ipar
ous
case
s&
con-
trol
s
GH
:B
P�
140/
90m
mH
gor
incr
ease
of�
30/1
5m
m
Hg
over
BP
at20
wee
ks
Non
e,b
utst
ratif
ied
by
dia
gnos
is
Chr
onic
HT,
rena
ldis
ease
,m
eta-
bol
icd
isea
se
123/
114
For
PE
vs.
Con
trol
s:
15.0
/12.
2
.21
PE
:G
H&
pro
tein
uria
(�50
0
mg/
24h
or�
�d
ipst
ick)
&
hyp
erur
icem
ia(�
5.5
mM
)
allr
esol
ving
afte
rp
reg-
nanc
y
*Not
e:al
so
foun
dno
diff
er-
ence
be-
twee
n
infa
nts
of
case
s&
cont
rols
(92)
Not
e:
May
in-
clud
e
som
eof
the
sam
e
sub
ject
s
as(1
06)
Italy
Inci
den
tG
H
(with
&w
ith-
out
pro
tein
-
uria
)
Nor
mot
ensi
vegr
avid
wom
enm
atch
edon
ethn
icity
Mul
tipar
ous
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
on2
occa
sion
s�
4h
apar
t&
with
(n�
70)
orw
ithou
t(n
�70
)p
rote
inur
ia(�
300
mg/
24h)
Mat
erna
lage
,p
arity
,
FVL
carr
ier
stat
us,
pro
thro
mb
in
A20
210
carr
ier
sta-
tus
Tran
sien
tH
T,d
iab
etes
,au
toim
mun
e
dis
ease
,ch
roni
cre
nalo
rp
ulm
o-
nary
dis
ease
&ch
roni
ces
sent
ial
HT
prio
rto
pre
gnan
cy
139/
216
24.5
(all
case
s)/
16.7
�.0
5
(91)
Isra
elIn
cid
ent
seve
re
PE
,A
P,
FGR
orst
ill-
birt
h
Age
&ge
ogra
phi
cally
mat
ched
norm
oten
sive
pre
gnan
cy
Mul
tipar
ous
case
s&
con-
trol
s
Sev
ere
PE
:B
P�
160/
110
mm
Hg,
pro
tein
uria
(�50
0
mg/
24h)
,H
ELL
Por
ecla
mp
sia
Non
e,b
utst
ratif
ied
by
dia
gnos
is
Var
ied
by
dia
gnos
is-n
one
spec
ified
for
seve
reP
E
110/
110
For
Sev
ere
PE
(n�
34):
20.6
/8.0
.05
(94)
Isra
elIn
cid
ent
seve
re
PE
Age
&et
hnic
itym
atch
ed
wom
enw
ith�
1no
r-
mal
pre
gnan
cy
Mul
tipar
ous
case
s&
con-
trol
s
BP
�16
0/11
0m
mH
g&
pro
-
tein
uria
(�50
0m
g/24
h),
HE
LLP
orec
lam
psi
a
Not
spec
ified
His
tory
ofth
rom
bop
hilic
even
t,H
T
inea
rlyp
regn
ancy
,m
ultip
lep
reg-
nanc
ies
63/1
2624
.0/1
0.0
.008
(una
dj)
�.0
5(a
dj)
54 Obstetrical and Gynecological Survey
TAB
LE5
Co
ntin
ued
Ref
eren
ce
Cou
ntry
of
Stu
dy
Typ
eof
Cas
esTy
pe
ofC
ontr
ols
Par
ityP
ED
efin
ition
Cov
aria
tes
(Ad
just
men
ts)
Exc
lusi
ons
Cas
es/C
ontr
ols
TT
Hom
ozyg
osity
%(C
ases
/
Con
trol
s)P
(117
)Fi
nlan
dP
E(n
�13
)&
Se-
vere
PE
(n�
100)
Hea
lthy
wom
enw
ith
hist
ory
of�
1no
r-
mot
ensi
vep
regn
an-
cies
Mul
tipar
ous
case
s&
con-
trol
s
PE
:B
P�
140/
90m
mH
gon
2oc
casi
ons
�6
hap
art
&
pro
tein
uria
(�30
0m
g/24
h
or�
dip
stic
k),
reso
lved
by
12w
eeks
pos
tpar
tum
Non
eR
enal
orau
toim
mun
ed
isea
se11
3/1
033.
0/7.
0.5
5
Sev
ere
PE
:�
160/
110
&p
ro-
tein
uria
(�20
0m
g/24
h),
reso
lved
by
12w
eeks
pos
tpar
tum
(118
)N
ethe
r-
land
s
PE
hist
ory;
con-
secu
tive
pa-
tient
s(n
�47
),
affe
cted
sib
-
pai
rfa
mili
es(n
�12
7)&
pa-
tient
sw
ith
know
nho
mo-
cyst
eine
stat
us
(n�
85)
Hea
lthy
Dut
chb
lood
don
ors
Not
spec
ified
for
case
s,un
-
know
nfo
r
cont
rols
Dia
stol
icB
P�
90m
mH
gon
atle
ast
2oc
casi
ons
�6
h
apar
tor
incr
ease
of20
mm
Hg
over
first
-trim
este
rle
v-
els
&p
rote
inur
ia(�
300
mg/
24h)
Str
atifi
edon
seve
rity
(PE
,H
ELL
Por
ecla
mp
sia)
;ad
-
just
edm
odel
with
hom
ocys
tein
e&
vita
min
leve
lsas
dep
end
ent
vari-
able
s&
geno
-
typ
esas
ind
epen
-
den
tva
riab
les
Non
e25
9/1
208.
5(c
onse
cu-
tive
case
s);
7.9
(sib
-pai
r
case
s)/9
.2
(con
trol
s)
.97
(con
-
secu
-
tive
case
s)
&0.
06
(sib
-
pai
r
case
s)
32.9
(hyp
erho
-
moc
ys-
tein
emia
)/
5.6
(nor
mal
hom
ocys
-
tein
e)
.001
(119
)A
ustr
alia
Sev
ere
PE
(n�
116)
&ec
lam
p-
sia
(n�
40)
case
s
Nor
mal
Pco
ntro
lsP
rimip
arou
s
case
s&
con-
trol
s
BP
�14
0/90
mm
Hg
orris
e
of25
/15
mm
Hg
from
bas
elin
eon
�2
occa
sion
s
�6
hap
art
&p
rote
inur
ia
(�30
0m
g/24
hor
��
dip
stic
k)
Non
eC
hron
icH
T,re
nald
isea
se,
met
a-
bol
icd
isea
se
156
/79
12.2
/13.
9�
.05
(110
)Io
wa
&
Nor
th
Car
o-
lina
Inci
den
tP
EH
isto
ryof
�2
norm
o-
tens
ive
term
pre
g-
nanc
ies
with
out
HT
orfa
mily
hist
ory
of
PE
Mul
tipar
ous
case
s&
con-
trol
s
Mild
PE
:B
P�
140/
90m
mH
g
&p
rote
inur
ia(�
300
mg/
24
hor
�d
ipst
ick)
Non
e,b
utst
ratif
ied
onp
arity
,P
Ese
-
verit
y&
HE
LLP
stat
us
Non
e(e
xcep
tas
liste
dfo
rco
ntro
ls)
281
/360
11.7
(all
PE
case
s)/1
1.4
.98
(97)
U.S
.In
cid
ent
Sev
ere
PE
Nor
mot
ensi
veth
roug
h-
out
pre
gnan
cyw
ith
Mul
tipar
ous
case
s&
con
BP
�16
0/11
0m
mH
g&
pro
-
tein
uria
(�30
0m
g/24
h),
Non
eC
hron
icH
T,d
iab
etes
,p
reex
istin
g
rena
ldis
ease
,hi
stor
yof
thro
m
Mat
erna
lSam
-
ple
s:
Mat
erna
lSam
-
ple
s
�.0
5
nohi
stor
yof
PE
trol
s(im
plie
d)
ecla
mp
sia
orH
ELP
boe
mb
olis
m,
mul
tifet
alge
stat
ion,
110
/97
9.0/
7.0
maj
orfe
talc
onge
nita
lano
mal
yFe
talS
amp
les:
Feta
lSam
ple
s:�
.05
75/9
29.
0/4.
0
GH
�ge
stat
iona
lhyp
erte
nsio
n;H
T�
hyp
erte
nsio
n;B
P�
blo
odp
ress
ure;
P�
pre
gnan
t;N
P�
nonp
regn
ant;
AP
�ab
rup
tiop
lace
ntae
;FG
R�
feta
lgro
wth
reta
rdat
ion.
Molecular Epidemiology of Preeclampsia Y CME Review Article 55
Endothelial Cell Health
Genes categorized as affecting endothelial cellhealth generally do so through their effect on oxida-tive stress. Specifically, small dense LDLs, which areincreased as part of the dyslipidemia present inwomen before the development of PE, are highlyoxidizable and can lead to membrane damage. Thus,genes involved in the production or metabolism ofreactive oxygen species (ROS) may lead to increasedoxidative stress and endothelial cell dysfunction.
Epoxide Hydrolase. Microsomal epoxide hydrolase(EPHX) is involved in the hydrolysis of certain oxidesto form less toxic products, but the process may alsoproduce toxic intermediates that could contribute to PEdevelopment (130). Two relevant polymorphisms havebeen described: Tyr113His in exon 3 has been associ-ated with decreased enzyme activity and His139Arg inexon 4 has been associated with increased enzymeactivity (130). The fast alleles that produce more activeenzymes, Tyr113 and Arg139, are hypothesized to in-crease the risk of PE by producing more toxic interme-diates and thus inducing endothelial cell damage. Allelefrequencies for the His139Arg polymorphism are rela-tively stable across populations but vary greatly bypopulation for the Tyr113His polymorphism. For ex-ample, the frequency of Tyr113/Tyr113 homozygotesranges from 26% to 69%, depending on the ethnicityand geography of the sample.
Thus far, only one study has addressed the possi-bility that one or both of these variants could play arole in PE development. Zusterzeel and colleagues(130) found no significant differences between casesand controls for genotypes at the 139 locus, however,at the 113 locus the fast genotype was two timesmore common in women with a history of PE than inwomen with a normotensive pregnancy history. Ad-ditionally, when genotypes at the two polymorphicsites were combined and categorized into high, inter-mediate, and low overall EPHX activity, Zusterzeelet al. (130) found that women with high EPHXactivity were over-represented in the PE cases com-pared with the controls.
A source of concern is the exclusion criteria usedin this study. Controls with known predisposing con-ditions (e.g., hypertension, renal disease, etc.) wereexcluded from the control population, although caseswith these conditions were not. In theory, bias awayfrom the null can occur when exclusions are madeamong the controls that are not made among thecases. Such bias could explain the associations seenin this study, therefore, more research on this gene is
needed before it can be dismissed or accepted as apotential candidate gene for PE.
Lipoprotein Lipase. Triglyceride and free fatty acidaccumulation in women with PE has led to speculationabout the potential role of the lipoprotein lipase (LPL)gene. LPL is responsible for mediating the clearance ofcirculating lipoprotein triglyceride and a decrease in itsactivity could lead to endothelial cell dysfunction. Fourrelatively common polymorphisms in the coding regionhave been identified thus far that are associated withalterations in plasma lipids: Asn291Ser (exon 6),Asp9Asn (exon 2), -93T 3G (promoter region), andSer447X (131). The Asn9 and -93G variants are instrong linkage disequilibrium and are inherited togetherin whites. Because the Ser291 and Asn9/-93G variantsresult in lower LDL activity and increased dyslipide-mia, they are expected to increase the risk of PE.Conversely, the X447 variant results in lower triglyc-erides and higher HDL concentrations and should,therefore, result in less risk of PE (132). Allele frequen-cies for the high-risk alleles were 1.5% for Ser291,0.7% for Asn9/-93G, and 9.4% for Ser447 in a randomsample of population controls from Pennsylvania (132).
To date, only two studies have examined the pos-sible role of LPL gene polymorphisms on the risk ofPE. In one well-designed study, the Ser291 andAsn9/-93G alleles were found statistically signifi-cantly more frequently among cases of PE thanamong either pregnancy or population controls, al-though the frequency of the X447 allele did not vary(132). Conversely, Kim et al. (133) failed to find astatistically significant difference in allele frequen-cies between cases of PE or their offspring comparedwith controls for the Asp9Asn, -93T 3G orAsn291Ser polymorphisms. However, among nullip-arous women with HELLP syndrome (n � 12), therewas a statistically significant increase in the fre-quency of the Ser291 allele compared with controls(8.3% vs. 1.5% respectively, P � .05) (133). Becauseboth study populations consisted entirely of whitesand patterns of linkage disequilibrium may differ byethnicity, more research should be conducted in otherethnic groups. However, the relative rarity of thesepolymorphisms suggests that they cannot possiblyaccount for a large proportion of PE cases.
Superoxide Dismutase. Superoxide dismutase(SOD) catalyzes the dismutation of superoxide intooxygen and hydrogen peroxide, thereby protectingagainst oxidative damage from superoxide radicals(134, 135). Decreased SOD activity results in in-creased levels of superoxides, which could reason-ably be expected to increase oxidative stress andpossibly result in PE.
56 Obstetrical and Gynecological Survey
There are three types of SOD in eukaryotic cells.The predominant type, encoded by the SOD1 gene, isfound in the cytosol and contains copper and zinc atthe catalytic site (CuZn-SOD), whereas the manga-nese-containing enzyme (Mn-SOD), encoded bySOD2, is found in the mitochondrial matrix (136). Athird unrelated gene, SOD3, encodes an extracellularenzyme also containing copper and zinc (137). Evi-dence suggests that trophoblasts from preeclampticplacentae generate statistically significantly more su-peroxide than trophoblasts from normal placentae(138). Likewise, SOD activity and relative mRNAexpression for CuZn-SOD were statistically signifi-cantly decreased in trophoblast cells from preeclamp-tic compared with normal placentae (138). Thus,increased superoxide generation seems to be associ-ated with decreased CuZn-SOD mRNA expressionand enzyme activity in trophoblast cells from pre-eclamptic placentae.
To date, only one study has examined the role ofthe SOD1 gene in relation to PE risk and no studieshave examined the role of SOD2 or SOD3. In a verysmall case-control study, Chen and colleagues (139)found that SOD activity was statistically significantlydecreased in women with PIH compared with preg-nant controls. They found no evidence of polymor-phism in EcoRI cutting sites in or near the CuZn-SOD gene; however, the possibility that other typesof polymorphisms in the gene might be responsiblefor the decreased SOD activity remains to be inves-tigated. In addition, 9 of the 14 women included inthe study did not have proteinuria, thus, they did notmeet the criteria for PE and the authors failed toexclude conditions known to be associated with anincreased risk of PE, creating a very heterogeneouspopulation. Thus, more epidemiological data areneeded before a role for the CuZn-SOD gene can beentirely dismissed and data need to be accumulatedto assess the role of the other SOD genes.
Long-Chain 3-Hydroxyacyl-CoA Dehydrogenas.Patients with long-chain 3-hydroxyacyl-CoA dehy-drogenase (LCHAD) deficiency have impaired oxi-dation of long-chain fatty acids and severe pregnancycomplications, including HELLP syndrome, havebeen reported in heterozygous carriers of a mutationin the LCHAD gene (140, 141). The mutation, aG3C substitution at position 1528 was shown to bedirectly responsible for the loss of LCHAD activity(142). Although mutation rates in subjects withLCHAD deficiency have been reported to be as highas 87% (142), the mutation seems to be very rare inthe general population (143).
The G1528C variant was studied in a Dutch pop-ulation, but not found to be associated with HELLPsyndrome (143). However, it has been suggested thatonly women carrying fetuses homozygous for theLCHAD mutation are at increased risk of HELLP.Unfortunately, Den Boer et al. (143) examined ma-ternal genotypes without consideration of fetal geno-types. Additionally, the lack of information on howthe cases and controls were selected for the studymakes these results difficult to interpret.
Apolipoprotein E. Apolipoprotein E (ApoE) is in-volved in the clearance of atherogenic remnants oftriglyceride-rich lipoproteins, and the ApoE gene hasthree common alleles �2, �3, and �4 (144). Choles-terol absorption efficiency in the intestines increasesin allelic order (�2 � �3 � �4) (145). Because theapoE �4 allele is an established risk factor for dys-lipidemia (146) and women with PE are known tohave alterations in their lipid profiles, the apoE �4allele would be expected to increase the risk of PE.
Nagy et al. (144) unexpectedly found a statisticallysignificant increase in PE risk associated with the �2allele and a nonsignificant reduction in �4 allelefrequency among preeclamptics compared withhealthy pregnant controls. In contrast, Makkonen andcolleagues (147) failed to find support for a role ofthe apoE gene in PE development. However, thestudy by Nagy and colleagues (144) was limited by afairly small sample size (52 healthy pregnant women,54 women with severe preeclampsia, and 101 healthynonpregnant women). More importantly, because therange of gestational ages in the normal, healthy preg-nancy group includes gestations that are before thegestational age at which PE is most commonly diag-nosed, we do not know if some of these controlwomen went on to develop PE. Additionally, bothstudies included a control group of healthy blooddonors, some of whom may have a history of PE. Ifsome women with PE were included in the controlgroups, it could obliterate a possible true associationbetween the ApoE �4 allele and PE as well as createspurious associations with other alleles. Thus, moreresearch is needed to clarify the possible role ofApoE genotype in PE development.
Immune Maladaptation
Immune function normally is suppressed duringpregnancy, ostensibly as an adaptive measure meantto protect the fetal allograft from maternal attack. Asmentioned previously, immune function seems to beabnormal in women who develop PE. Thus, genesthat encode the various aspects of the immune system
Molecular Epidemiology of Preeclampsia Y CME Review Article 57
may be good candidates for involvement in PEdevelopment.
Human Leukocyte Antigens. Human leukocyte an-tigens (HLA) are involved in immune tolerance andthus, deviation from normal expression may play arole in the immune maladaptation believed to beimportant in PE development. Specifically, HLA isthought to be involved in the rejection of the fetus bythe mother, or vice versa. To date, the literature inthis area is rather muddled with most studies failingto prove replicable (25). The usual difficulties instudying genetic susceptibility to PE are com-pounded by the fact that there are a large number ofantigens at each HLA locus, making it very difficultto study them effectively in small studies. In addi-tion, studies attempting to investigate numerous an-tigens suffer from the problem of multiple compari-sons and uninterpretable older studies in whichlaboratory methodologies were less reliable in distin-guishing between homozygotes and heterozygotes(25). Because comprehensive reviews have alreadybeen written to address the findings in the HLAsystem (25, 40, 148), we focus here on only a few ofthe more recent findings.
HLA-G. HLA-G is highly expressed in first trimes-ter but greatly reduced in third trimester cytotropho-blast cells suggesting that it may be important inplacental development (149) and in adequate tropho-blast invasion (150). HLA-G has also been shown tohelp regulate the release of cytokines and thus mightbe important in blocking a maternal immune re-sponse to the placenta and ensuring a viable preg-nancy (151, 152).
Despite evidence suggesting that HLA-G expres-sion in trophoblasts is reduced in preeclamptic preg-nancies (153), the results of both linkage and candi-date gene analyses fail to support the hypothesizedrole of HLA-G in genetic susceptibility to PE. Spe-cifically, Bermingham et al. (152) found no differ-ence between PE offspring and control offspring in awhite population for three polymorphisms studied:A3T substitution at codon 107, C3A substitution atcodon 110, and a 14-bp insertion/deletion polymor-phism in exon 8. An excess of heterozygotes for thedeletion polymorphism was found in PE offspring,but the relevance of this deviation has yet to bedetermined. Moreover, a well-designed study exam-ined the role of both maternally and fetally expressedHLA-G on the risk of PE, but found that neithershowed linkage to PE or eclampsia (E) (151).
Unfortunately, the Bermingham study failed to de-scribe the source population for its cases and con-trols, and therefore, it is impossible to assess whether
selection bias may be affecting the results. Addition-ally, proteinuria was not required for a PE diagnosis,suggesting that the study population may be hetero-geneous. Lastly, a relatively small sample size (n �68) could have limited their power to detect an effect.
HLA-DR. The HLA-DR antigens play a role in therecognition of self versus nonself and have been thefocus of several studies (154). To date, only the roleof maternal HLA-DR� has been examined; the pos-sible role of fetal HLA-DR� genotype PE risk hasnot be studied. The only study to examine this locusfailed to find linkage between maternal HLA-DR�and PE (155).
Among the more promising of the HLA-DR allelesis HLA-DR4, although the results are hardly conclu-sive. A study conducted by Kilpatrick et al. (156)found that the frequency of HLA-DR4 was statisti-cally significantly increased in preeclamptic womenas well as their babies compared with controls. Thestrongest finding was that HLA-DR4 sharing be-tween a preeclamptic mother and her fetus was sta-tistically significantly higher than between controlsand their babies (26% vs. 7.7%, respectively). None-theless, other studies found no association (42) orlinkage (155) between PE and HLA-DR4.
Other HLA. HLA-A, -B, and -C genes produceproducts that stimulate graft rejection (of fetus bymother or vice versa) and are blocked in placentaltrophoblast cells (157). In a relatively large prospec-tive study (n � 712 primigravids), women with theA23/29, B44, and DR7 haplotypes had a statisticallysignificant increase in incidence of PE (158). Despitethese highly suggestive findings, the frequency ofthese haplotypes in the general population are ratherlow (�5% of whites and �1% of blacks), ruling outthe possibility of a major contribution to disease.
TNF-�. Several lines of evidence support a role forTNF-� in the development of PE. First, PE has beenassociated with increased plasma and amniotic levelsof TNF-� and TNF receptors (159, 160). Second,TNF-� has been detected during fertilization and isthought to interact with other cytokines in early preg-nancy to promote growth and differentiation andnormal placentation (160, 161). Moreover, the blas-tocyst bears receptors for TNF-�, suggesting thatTNF-� is important in early embryo development(162). Third, an increased thromboxane-to-prostacy-clin ratio has been suggested to play a role in thepathophysiology of PE. Increased levels of TNF-�may contribute to the increased levels of thrombox-ane, which then leads to vasoconstriction and plateletaggregation (161, 163). Lastly, the endothelial celldamage observed in PE is thought to be a result of
58 Obstetrical and Gynecological Survey
attack by ROS. TNF-� can not only generate ROS,but may also interfere with the buffering capacity ofintracellular components so that the endothelium be-comes more susceptible to oxidant-mediated injury(163).
A substitution polymorphism at position –308 inthe promoter of the TNF-� gene produces two allelesknown as TNFA-1 (G) and TNFA-2 (A). TheTNFA-2 allele disrupts an AP-2 binding site, result-ing in six- to seven-fold higher levels of reportergene transcription in both mitogen-stimulated andunstimulated cells (164). Moreover, individuals ho-mozygous for the TNFA-2 allele have significantlyhigher constitutive and inducible levels of TNF-�secretion compared with TNFA-1 homozygotes, withheterozygous subjects having intermediate levels(165, 166). Thus, we expect women who are ho-mozygotes or heterozygotes for TNFA-2 to haveincreased risk of PE. Reported allele frequencies forthe TNFA-2 allele range from 16% in a Tunisianstudy (167) to 23% in a Dutch study (165).
Epidemiological studies have had mixed results,with one study suggesting that the TNFA-1 allele isincreased in PE (163), although another found nosignificant differences in allele frequencies betweenpreeclamptic women and normotensive gravid con-trols or published allele frequencies (161). The studyby Chen et al. (163) is limited by its small number ofPE cases (n � 14), the lack of exclusions for knownunderlying conditions (e.g., renal disease, diabetes,etc.) and the inclusion of multiparas. Moreover, thefinding of increased TNFA-1 frequency is counter-intuitive to what would be expected given our under-standing of the mechanisms involved and outlines theneed for additional research in this area.
Additionally, a second polymorphism exists at po-sition –238 in the promoter region, but does not seemto be associated with PE (168). The TNF-� gene isclosely linked to the gene for lymphotoxin-� (alsocalled TNF-�). The two polymorphisms describedfor the TNF-� gene in combination with two poly-morphisms in the TNF-� gene define five haplo-types, TNF-C, -E, -H, -I, and -P (169). Lachmeijer etal. (168) found an increased risk of PE or HELLPassociated with having at least one copy of the TNF-Ihaplotype. However, this association was only foundamong index cases, particularly index cases with PEonly, but not among their sisters, who met similardisease criteria. Other than suggesting that PE andHELLP are distinct diseases, these results remainpuzzling because any association with a haplotypeobserved in the index cases should have also been
seen among the similarly afflicted sisters of the indexcases.
Interleukin-1�. IL-1�, a cytokine produced by en-dothelial cells, monocytes and macrophages is in-volved in the initiation of a proinflamatory response.Both IL-1� and interleukin-1 receptor antagonist (IL-1RA) bind to the IL-1 receptor, the first initiating aproinflamatory response and the second terminatingthe inflammatory event. Although circulating levelsof IL-1� and IL-1RA have not been consistentlyelevated in preeclamptics, IL-1� expression in pla-centas of preeclamptic patients has been found to beupregulated (170).
Two genes within the interleukin beta cluster havebeen examined with respect to PE risk: the genesencoding IL-1� and IL-1RA. Two polymorphisms inthe IL-1� gene, one in the promoter at position –511(C3T) and another in exon 5 correlate with alteredIL-1� protein expression (171). Additionally, thesecond intron of the IL-1RA gene contains a 86 bprepeat polymorphism that varies between two and sixrepeats, with four repeats being the most common(IL-1RA*1) and 2 repeats (IL-1RA*2) being associ-ated with prolonged inflammatory response (172).Women who are homozygous for IL-1RA*2 wouldbe expected to be at increased risk of PE as wouldwomen with the IL-1� promoter variant (-511T) andthe IL-1� exon 5 variant (E2).
One study examined the role of all three polymor-phisms in relation to PE risk and found that none ofthe polymorphisms appeared to increase the risk ofPE (171). However, the power to detect an effect forthese variants was very low, thus a small to moderateeffect would likely go undetected in this study. Nev-ertheless, the results did suggest that disease severitymight be influenced by variations in these two genes.Specifically, preeclamptics with the IL-1RA*2 vari-ant had statistically significantly higher mean sys-tolic blood pressure upon admission compared withnormotensive pregnant controls (171). Moreover,when all three polymorphisms were combined, pre-eclamptic women with at least three mutant alleleshad statistically significantly higher mean systolicblood pressure than other preeclamptics (171).
Placentation
There are a large number of genes that regulate allaspects of placental growth, development, and func-tion. It has been suggested that these genes are goodcandidates for predisposing to PE due to their obvi-ous connection to placental function (22). Studiesconducted in mice have shown that mutations in
Molecular Epidemiology of Preeclampsia Y CME Review Article 59
placentation genes can affect signaling interactionsbetween embryonic and trophoblast tissues and leadto abnormal vascularization of the placenta (173).
Although functional data on the role of specificgenes in human placental development are very lim-ited, what is known seems to correspond well withthe findings in mice. Genes involved in the differen-tiation and maintenance of cells in the trophoblastlineage seem to be the best candidates for a role in PEdevelopment. Specifically, human homologues forMash2, which is involved in the maintenance oftrophoblast stem cells and Hand1, which promotesthe differentiation of trophoblast giant cells mayprove to be important in human placental develop-ment (173). Additionally, the chorionic trophoblastcells in Gcm1-deficient mice fail to undergo differ-entiation into syncytiotrophoblasts (174), whichcould theoretically lead to inadequate spiral arteryinvasion and eventually, PE. Of these genes, onlyHash2, the human homologue to Mash2, is known tobe polymorphic, containing a polymorphism in the 3'UTR of the gene. To date, no molecular epidemiol-ogy studies have been conducted to examine the roleof polymorphisms in placentation genes and the riskof PE but it is an area of research that deservesattention.
Other
IGF. Insulin-like growth factor-II (IGF-II) is in-volved in mammalian growth and has an importantinfluence on fetal cell division and differentiation(175). Moreover, trophoblast-derived IGF-II may beimportant for invasion and for both trophoblast anddecidual function (176). Beckwith-Wiedemann Syn-drome, a condition of prenatal overgrowth and pre-disposition to embryonic malignancies, has been as-sociated with both IGF-II overexpression and severe,early onset PE (175). This has prompted the hypoth-esis that genetic variation in IGF-II may be involvedin the development of PE by restricting intrauterinefetal growth.
An ApaI polymorphism in the 3' flanking region ofthe IGF-II gene produces two alleles, A and B. Allelefrequencies for the B allele was approximately 40%among an Irish population (152). Bermingham andcolleagues (152) found no association between theIGF-II polymorphism and PE using a mother-father-child trio design. However, this study suffers fromsome methodologic issues, as described earlier in thisarticle (HLA-G), which make the significance ofthese results difficult to interpret.
In addition to IGF-II, insulin-like growth factorbinding protein-1 (IGFBP1) and IGF-I have hypoth-esized etiological roles in PE. IGFBP1 is thought toinhibit cytotrophoblast invasion into the endometrialstromal multilayers and IGF-I is thought to be im-portant in fetal growth (177). Guidice and colleagues(177) found that in women with severe PE, circulat-ing levels of IGFBP-1 is significantly elevated, IGF-Iis significantly lower and IGF-II is no different incases versus premature labor controls. However, sev-eral concerns, including lack of control for potentialconfounders (e.g., intrauterine growth restriction)and the use of a potentially inappropriate comparisongroup (preterm labor controls might not be represen-tative of the base population from which the casesarose) may compromise the validity of these results.Thus, no definitive conclusions can be drawn fromthese data.
Mitochondrial DNA. The importance of energy forthe transport of nutrients across the placenta to thefetus has led some to hypothesize a role for reducedmitochondrial gene expression in the development ofPE (178). Many of the features of PE can be ex-plained by mitochondrial dysfunction, including va-soconstriction, platelet aggregation, disturbed iontransport, reduced prostaglandin synthesis, and hy-peruricemia (179). Moreover, uterine and placentaltissues obtained from women with PE showed en-gorgement in the endothelial mitochondria with lossof cristae and other mitochondrial changes sugges-tive of a systemic metabolic disorder (180).
However, the evidence for a mitochondrial contri-bution to PE development is mixed. Although oneEuropean study found no evidence of reduced respi-ratory chain enzyme activities in preeclamptic pla-centae (181), a very small study in Japan found areduced expression of cytochrome c oxidase andcytochrome oxidase subunit I (both of which areencoded by the mitochondrial genome) in placentaltissue from preeclamptic patients compared withwomen whose gestations were appropriate for theirgestational age (178). Another study examined twoNorwegian families with a high incidence of pre-eclampsia/eclampsia to determine if there were anymutations in their mitochondrial DNA (182). Theyfound two different single nucleotide polymorphisms(SNPs) in these families, but no causal relationshipcould be established because a high prevalence of agiven polymorphism within affected families doesnot necessarily imply causation. Additionally, theauthors included women who had PE/E in a secondor later pregnancy, indicating that the etiology oftheir PE may be different than primiparous PE. Al-
60 Obstetrical and Gynecological Survey
though it is conceivable that mitochondrial dysfunc-tion plays a role in a small subset of women with PE,it is unlikely to be involved in the majority of cases.
DISCUSSION
A common theme in almost all of the epidemio-logical evidence produced thus far is methodologicalproblems. A wide range of PE definitions is used,making it very difficult to compare studies. First, theinclusion of multiparous women in the study of adisease that is considered to be primarily one offirst-time mothers has created heterogeneous studypopulations, making it difficult to understand thegenetic factors that may be at play. Moreover, mul-tiparous women are more likely to have developedPE as a result of an underlying medical condition,which may or may not have a genetic basis but thatis brought out by the physical demands of pregnancy.These etiologic differences between parous andnonparous women can and probably do lead to spu-rious associations between PE and genes that areactually related to another disease.
Second, the failure to exclude women with essen-tial hypertension or hypertension before the 20thweek of gestation could lead to an inflated odds ratiofor genes, such as AGT, that are known to be asso-ciated with hypertension. Therefore, studies of PEshould exclude women with underlying conditionsthat resemble PE and either restrict enrollment tonulliparous women or stratify by parity at the anal-ysis stage. However, it should be noted that stratify-ing would substantially decrease the sample size andthus the power to detect an effect within a givenstratum.
The importance of fetal genotype, in addition tomaternal genotype, is fairly well recognized, yet theactualization of including fetal genotype assessmenthas, for the most part, not yet occurred. One of thebarriers to including fetal genotype is that, to look atgenotype-by-genotype interactions, a very large sam-ple size is needed. A large number of small studiessimply cannot address the question of maternal-fetalgenotypic interactions and, as such, large studiesshould be conducted on multiple genes. The inclu-sion of participants who are not white is essential,provided that large enough numbers of minority sub-jects can be recruited to provide a meaningfulanalysis.
With such an extensive list of candidate genes andmany others not yet studied, the challenge now be-comes determining which of them hold the mostpromise for future research. If our current under-
standing of the disease process is correct, then themost promising candidates may be those that areinvolved in placentation, vascular remodeling, andendothelial cell health. Placentation genes have notbeen included in any epidemiologic studies to dateand thus deserve special attention. For all of thesegenes, studies are needed to better characterize thesubpopulations that may be affected. Some genesthat have been associated with PE, such as those inthe thrombophilic pathway (prothrombin, FVL) maycharacterize a subpopulation at elevated risk, but donot seem to be independent risk factors for PE.MTHFR may be an important gene among folate-deficient women and, therefore, should be consideredwhen examining poor or undernourished popula-tions. Polymorphisms in LPL and LCHAD are toorare to account for a large number of PE cases, butmay be important in certain subsets of women. Evi-dence for an association between PE and genes in theblood pressure regulation pathway is fairly weak, butthese genes cannot be ruled out until an examinationof maternal-fetal genotypic interactions has beencompleted. Additionally, limited research on thegenes for GPIIIa, MMPI, EPHX, SOD, and ApoEhave had promising results and suggest that theymight be important in at least some populations.Because of the failure to include fetal genotype inmost molecular epidemiology studies, it seems pre-mature to rule out any of the potential candidatesbased on results obtained in maternal samples alone.
In conclusion, more molecular epidemiology stud-ies need to be conducted using larger sample sizeswhere genotype-by-genotype interactions can be ex-amined, especially maternal-fetal genotypic interac-tions. Additionally, placentation genes must be addedto the list of candidate genes.
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