Refined phenotyping identifies links between preeclampsia and related diseases in a Norwegian preeclampsia family cohort.

OBJECTIVE: Preeclampsia is a complex genetic disease of pregnancy with a heterogenous presentation, unknown cause and potential severe outcomes for both mother and child. Preeclamptic women have increased risk for atherothrombotic cardiovascular disease. We aimed to identify heritabilities and phenotypic correlations of preeclampsia and related conditions in the Norwegian Preeclampsia Family Biobank. METHODS: By applying a variance components model, a total of 493 individuals (from 138 families with increased occurrence of preeclampsia) were classified according to 30 disease-related phenotypes. RESULTS: Of parous women, 75.7% (263/338) had experienced preeclampsia and 35.7% of women with and 22.4% without preeclampsia delivered children small for gestational age (SGA). We identified 11 phenotypes as heritable. The increased occurrence of preeclampsia was reflected by the presence [heritability (H2r) = 0.60)] and severity (H2r = 0.15) of preeclampsia and being born in a preeclamptic pregnancy (H2r = 0.25). Other heritable phenotypes identified included SGA (H2r = 0.40), chronic hypertension (H2r = 0.57), severity of atherothrombotic cardiovascular disease (H2r = 0.31), BMI (H2r = 0.60) and pulmonary disease (H2r = 0.91). The heritable phenotype preeclampsia overlapped with SGA (P = 0.03), whereas pulmonary disease was phenotypically correlated with atherothrombotic cardiovascular disease (P < 0.01), SGA (P = 0.02) and BMI (P = 0.02). CONCLUSION: This is the first study identifying the H2r of a range of health-related conditions in preeclamptic families. Our study demonstrates how refinement of phenotypes leads to better H2r estimation and the identification of a biological relationship between preeclampsia and related traits.

ADAMTS13 gene variants and function in women with preeclampsia: a population- based nested case- control study from the HUNT Study.

INTRODUCTION: Known genetic variants with reference to preeclampsia only explain a proportion of the heritable contribution to the development of this condition. The association between preeclampsia and the risk of cardiovascular disease later in life has encouraged the study of genetic variants important in thrombosis and vascular inflammation also in relation to preeclampsia. The von Willebrand factor-cleaving protease, ADAMTS13, plays an important role in micro vascular thrombosis, and partial deficiencies of this enzyme have been observed in association with cardiovascular disease and preeclampsia. However, it remains unknown whether decreased ADAMTS13 levels represent a cause or an effect of the event in placental and cardiovascular disease. METHODS: We studied the distribution of three functional genetic variants of ADAMTS13, c.1852C>G (rs28647808), c.4143_4144dupA (rs387906343), and c.3178C>T (rs142572218) in women with preeclampsia and their controls in a nested case-control study from the second Nord-Trøndelag Health Study (HUNT2). We also studied the association between ADAMTS13 activity and preeclampsia, in serum samples procured unrelated in time of the preeclamptic pregnancy. RESULTS: No differences were observed in genotype, allele or haplotype frequencies of the different ADAMTS13 variants when comparing cases and controls, and no association to preeclampsia was found with lower levels of ADAMTS13 activity. CONCLUSION: Our findings indicate that ADAMTS13 variants and ADAMTS13 activity do not contribute to an increased risk of preeclampsia in the general population.

Preeclampsia and cardiovascular disease share genetic risk factors on chromosome 2q22.

OBJECTIVE: Four putative single nucleotide polymorphism (SNP) risk variants at the preeclampsia susceptibility locus on chromosome 2q22; rs2322659 (LCT), rs35821928 (LRP1B), rs115015150 (RND3) and rs17783344 (GCA), were recently shown to associate with known cardiovascular risk factors in a Mexican American cohort. This study aimed to further evaluate the pleiotropic effects of these preeclampsia risk variants in an independent Australian population-based cohort. METHODS: The four SNPs were genotyped in the Western Australian Pregnancy Cohort (Raine) Study that included DNA, clinical and biochemical data from 1246 mothers and 1404 of their now adolescent offspring. Genotype association analyses were undertaken using the SOLAR software. RESULTS: Nominal associations (P<0.05) with cardiovascular risk factors were detected for all four SNPs. The LCT SNP was associated with decreased maternal height (P=0.005) and decreased blood glucose levels in adolescents (P=0.022). The LRP1B SNP was associated with increased maternal height (P=0.026) and decreased maternal weight (P=0.044). The RND3 SNP was associated with decreased triglycerides in adolescents (P=0.001). The GCA SNP was associated with lower risk in adolescents to be born of a preeclamptic pregnancy (P=0.003) and having a mother with prior preeclamptic pregnancy (P=0.033). CONCLUSIONS: Our collective findings support the hypothesis that genetic mechanisms for preeclampsia and CVD are, at least in part, shared, but need to be interpreted with some caution as a Bonferroni correction for multiple testing adjusted the statistical significance threshold (adjusted P<0.001).

Validity of the diagnosis of pre-eclampsia in the Medical Birth Registry of Norway.

OBJECTIVE: Evaluating the validity of pre-eclampsia registration in the Medical Birth Registry of Norway (MBRN) according to both broader and restricted disease definitions. DESIGN: Retrospective nested cohort study. SETTING: Multicenter study. POPULATION: In this study, two cohorts of women with pre-eclamptic pregnancies registered in the MBRN were selected. Study group 1 contained 966 pregnancies from 1967 to 2002. Concomitant participation in the Nord-Trøndelag Health Study 2 was required. Study group 2 comprised 1138 pregnancies recorded in 1967-2005, examined as a pre-eclampsia biobank was established. METHODS: Diagnostic criteria vary. The broader criteria for pre-eclampsia, used by the MBRN, are one measurement of hypertension and proteinuria (Criterion A). Criteria used internationally today require two measurements of hypertension and proteinuria (Criterion B). The diagnostic validities in Study groups 1 and 2 were judged against medical records according to Criterion A and B, respectively. MAIN OUTCOME MEASURES: Positive predictive value (PPV) and trend analyses. RESULTS: The diagnosis was confirmed in 88.3% of pregnancies in Study group 1, and in 63.6% in Study group 2. PPV was high for Study group 1 throughout the period. For Study group 2, results improved significantly after 1986. CONCLUSIONS: This study ascertains high PPV of pre-eclampsia in the MBRN using broader traditional criteria, although the PPV decreases through assessment using restricted modern criteria. This illustrates how inclusion of direct measurements may improve registration of complex disorders defined by changing diagnostic criteria.

Genetic dissection of the pre-eclampsia susceptibility locus on chromosome 2q22 reveals shared novel risk factors for cardiovascular disease.

Pre-eclampsia is an idiopathic pregnancy disorder promoting morbidity and mortality to both mother and child. Delivery of the fetus is the only means to resolve severe symptoms. Women with pre-eclamptic pregnancies demonstrate increased risk for later life cardiovascular disease (CVD) and good evidence suggests these two syndromes share several risk factors and pathophysiological mechanisms. To elucidate the genetic architecture of pre-eclampsia we have dissected our chromosome 2q22 susceptibility locus in an extended Australian and New Zealand familial cohort. Positional candidate genes were prioritized for exon-centric sequencing using bioinformatics, SNPing, transcriptional profiling and QTL-walking. In total, we interrogated 1598 variants from 52 genes. Four independent SNP associations satisfied our gene-centric multiple testing correction criteria: a missense LCT SNP (rs2322659, P = 0.0027), a synonymous LRP1B SNP (rs35821928, P = 0.0001), an UTR-3 RND3 SNP (rs115015150, P = 0.0024) and a missense GCA SNP (rs17783344, P = 0.0020). We replicated the LCT SNP association (P = 0.02) and observed a borderline association for the GCA SNP (P = 0.07) in an independent Australian case-control population. The LRP1B and RND3 SNP associations were not replicated in this same Australian singleton cohort. Moreover, these four SNP associations could not be replicated in two additional case-control populations from Norway and Finland. These four SNPs, however, exhibit pleiotropic effects with several quantitative CVD-related traits. Our results underscore the genetic complexity of pre-eclampsia and present novel empirical evidence of possible shared genetic mechanisms underlying both pre-eclampsia and other CVD-related risk factors.

OP004. A SNP associated with susceptibility to preeclampsia near the inhibin, beta B gene, is also associated with cardiovascular disease risk traits.

INTRODUCTION: It is well established that preeclampsia (PE) increases later life cardiovascular disease (CVD) risk. Consequently, PE has started to gain a role as an early screening criterion for CVD. PE and CVD share several risk factors, pathological features and metabolic abnormalities. These common antecedents have drawn attention to the likelihood of shared genetic susceptibility. OBJECTIVES: Results from our previous PE GWAS identified a significant association with the rs7579169 SNP and maternal PE susceptibility (odds ratio 1.57). This SNP resides near the Inhibin, beta B (INHBB) gene on chromosome 2q14. Therefore, this study sought to interrogate this PE susceptibility SNP against several CVD risk traits in an effort to highlight additional empirical evidence of likely shared PE/CVD genetic mechanisms. METHODS: The rs7579169 SNP was genotyped in a large independent Australian cohort rich in quantitative CVD risk traits; The Western Australian Pregnancy Cohort (Raine) Study. This cohort comprises of fasting blood samples from 1246 mothers and 1461 adolescents (14- and 17-year-old) and clinical parameters pertaining, but not limited, to anthropometric measures of adiposity and lipid-related measures. Genetic association analyses of rs7579169 against the Raine CVD-related risk traits were performed using the software package R. All statistical analyses assumed an additive model of gene action. RESULTS: Significant associations (p<0.05) for rs7579169 with CVD-related risk traits were detected, both for the mothers and the adolescents. Specifically, the minor rs7579169-T allele (MAF 0.400) was found to be significantly associated with elevated levels of triglycerides, total and LDL cholesterol, a greater average waist:hip circumference ratio and a greater average hip circumference. CONCLUSION: We have previously identified rs7579169 located near the INHBB gene on chromosome 2q14 to significantly associate with maternal PE susceptibility. We have now demonstrated that this SNP is also significantly associated with several CVD-related risk traits in an independent Caucasian population. We hereby present additional empirical evidence of possible shared genetic risk factors underlying both PE and CVD related traits.

Genome-wide association scan identifies a risk locus for preeclampsia on 2q14, near the inhibin, beta B gene.

Elucidating the genetic architecture of preeclampsia is a major goal in obstetric medicine. We have performed a genome-wide association study (GWAS) for preeclampsia in unrelated Australian individuals of Caucasian ancestry using the Illumina OmniExpress-12 BeadChip to successfully genotype 648,175 SNPs in 538 preeclampsia cases and 540 normal pregnancy controls. Two SNP associations (rs7579169, p = 3.58×10(-7), OR = 1.57; rs12711941, p = 4.26×10(-7), OR = 1.56) satisfied our genome-wide significance threshold (modified Bonferroni p<5.11×10(-7)). These SNPs reside in an intergenic region less than 15 kb downstream from the 3' terminus of the Inhibin, beta B (INHBB) gene on 2q14.2. They are in linkage disequilibrium (LD) with each other (r(2) = 0.92), but not (r(2)<0.80) with any other genotyped SNP ±250 kb. DNA re-sequencing in and around the INHBB structural gene identified an additional 25 variants. Of the 21 variants that we successfully genotyped back in the case-control cohort the most significant association observed was for a third intergenic SNP (rs7576192, p = 1.48×10(-7), OR = 1.59) in strong LD with the two significant GWAS SNPs (r(2)>0.92). We attempted to provide evidence of a putative regulatory role for these SNPs using bioinformatic analyses and found that they all reside within regions of low sequence conservation and/or low complexity, suggesting functional importance is low. We also explored the mRNA expression in decidua of genes ±500 kb of INHBB and found a nominally significant correlation between a transcript encoded by the EPB41L5 gene, ∼250 kb centromeric to INHBB, and preeclampsia (p = 0.03). We were unable to replicate the associations shown by the significant GWAS SNPs in case-control cohorts from Norway and Finland, leading us to conclude that it is more likely that these SNPs are in LD with as yet unidentified causal variant(s).

Genetic and molecular functional characterization of variants within TNFSF13B, a positional candidate preeclampsia susceptibility gene on 13q.

BACKGROUND: Preeclampsia is a serious pregnancy complication, demonstrating a complex pattern of inheritance. The elucidation of genetic liability to preeclampsia remains a major challenge in obstetric medicine. We have adopted a positional cloning approach to identify maternal genetic components, with linkages previously demonstrated to chromosomes 2q, 5q and 13q in an Australian/New Zealand familial cohort. The current study aimed to identify potential functional and structural variants in the positional candidate gene TNFSF13B under the 13q linkage peak and assess their association status with maternal preeclampsia genetic susceptibility. METHODOLOGY/PRINCIPAL FINDINGS: The proximal promoter and coding regions of the positional candidate gene TNFSF13B residing within the 13q linkage region was sequenced using 48 proband or founder individuals from Australian/New Zealand families. Ten sequence variants (nine SNPs and one single base insertion) were identified and seven SNPs were successfully genotyped in the total Australian/New Zealand family cohort (74 families/480 individuals). Borderline association to preeclampsia (p = 0.0153) was observed for three rare SNPs (rs16972194, rs16972197 and rs56124946) in strong linkage disequilibrium with each other. Functional evaluation by electrophoretic mobility shift assays showed differential nuclear factor binding to the minor allele of the rs16972194 SNP, residing upstream of the translation start site, making this a putative functional variant. The observed genetic associations were not replicated in a Norwegian case/control cohort (The Nord-Trøndelag Health Study (HUNT2), 851 preeclamptic and 1,440 non-preeclamptic women). CONCLUSION/SIGNIFICANCE: TNFSF13B has previously been suggested to contribute to the normal immunological adaption crucial for a successful pregnancy. Our observations support TNFSF13B as a potential novel preeclampsia susceptibility gene. We discuss a possible role for TNFSF13B in preeclampsia pathogenesis, and propose the rs16972194 variant as a candidate for further functional evaluation.

The ERAP2 gene is associated with preeclampsia in Australian and Norwegian populations.

Preeclampsia is a heritable pregnancy disorder that presents new onset hypertension and proteinuria. We have previously reported genetic linkage to preeclampsia on chromosomes 2q, 5q and 13q in an Australian/New Zealand (Aust/NZ) familial cohort. This current study centered on identifying the susceptibility gene(s) at the 5q locus. We first prioritized candidate genes using a bioinformatic tool designed for this purpose. We then selected a panel of known SNPs within ten prioritized genes and genotyped them in an extended set of the Aust/NZ families and in a very large, independent Norwegian case/control cohort (1,139 cases, 2,269 controls). In the Aust/NZ cohort we identified evidence of a genetic association for the endoplasmic reticulum aminopeptidase 1 (ERAP1) gene (rs3734016, P (uncorr) = 0.009) and for the endoplasmic reticulum aminopeptidase 2 (ERAP2) gene (rs2549782, P (uncorr) = 0.004). In the Norwegian cohort we identified evidence of a genetic association for ERAP1 (rs34750, P (uncorr) = 0.011) and for ERAP2 (rs17408150, P (uncorr) = 0.009). The ERAP2 SNPs in both cohorts remained statistically significant (rs2549782, P (corr) = 0.018; rs17408150, P (corr) = 0.039) after corrections at an experiment-wide level. The ERAP1 and ERAP2 genes encode enzymes that are reported to play a role in blood pressure regulation and essential hypertension in addition to innate immune and inflammatory responses. Perturbations within vascular, immunological and inflammatory pathways constitute important physiological mechanisms in preeclampsia pathogenesis. We herein report a novel preeclampsia risk locus, ERAP2, in a region of known genetic linkage to this pregnancy-specific disorder.

Association between the candidate susceptibility gene ACVR2A on chromosome 2q22 and pre-eclampsia in a large Norwegian population-based study (the HUNT study).

Genome-wide scans in Icelandic, Australian/New Zealand and Finnish pedigrees have provided evidence for maternal susceptibility loci for pre-eclampsia on chromosome 2, although at different positions (Iceland: 2p13 and 2q23, Australia/New Zealand: 2p11-12 and 2q22, Finland: 2p25). In this project, a large population-based (n=65 000) nested case-control study was performed in Norway to further explore the association between positional candidate genes on chromosome 2q and pre-eclampsia, using single-nucleotide polymorphisms (SNPs). DNA samples from 1139 cases (women with one or more pre-eclamptic pregnancies) and 2269 controls (women with normal pregnancies) were genotyped using the Applied Biosystems SNPlex high-throughput genotyping assay. In total, 71 SNPs within positional candidate genes at 2q22-23 locus on chromosome 2 were genotyped in each individual. Genotype data were statistically analysed with the sequential oligogenic linkage analysis routines (SOLAR) computer package. Nominal evidence of association was found for six SNPs (rs1014064, rs17742134, rs1424941, rs2161983, rs3768687 and rs3764955) within the activin receptor type 2 gene (ACVR2A) (all P-values <0.05). The non-independence of statistical tests due to linkage disequilibrium between SNPs at a false discovery rate of 5% identifies our four best SNPs (rs1424941, rs1014064, rs2161983 and rs3768687) to remain statistically significant. The fact that populations with different ancestors (Iceland/Norway-Australia/New Zealand) demonstrate a common maternal pre-eclampsia susceptibility locus on chromosome 2q22-23, may suggest a general role of this locus, and possibly the ACVR2A gene, in pre-eclampsia pathogenesis.