SEARCH

SEARCH BY CITATION

Keywords:

  • ABO(H);
  • fetal growth restriction;
  • Lewis;
  • secretor;
  • von Willebrand factor

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References

Summary. Background: Fetal growth restriction (FGR) is associated with thrombosis of the placenta and an increased risk of subsequent vascular disease in the mother and fetus. The products of interactions between ABO(H), Lewis and Secretor genes are also associated with thrombosis and vascular disease risk. Objectives/Methods: A prospective case–control study of mothers with a severe FGR pregnancy (cases, n = 128; controls, n = 288) was performed to determine whether FGR is associated with particular maternal blood groups. Results: No association with ABO(H) status was observed, but FGR was more common in maternal secretors (odds ratio [OR] 1.70, 95% confidence interval [CI] 1.08–2.69) and consequently in those mothers expressing Le(b) on their red cells (OR 1.80, 95% CI 1.15–2.83), with a reduced risk in non-secretors and those expressing Le(a). Given the association between blood groups and both activated protein C resistance (APCR) and von Willebrand factor (VWF) levels, post hoc pilot studies on first-trimester APCR and VWF antigen levels and blood group genotypes were performed. No relationship with Lewis or Secretor was observed. Despite this, lower first-trimester VWF levels were observed in pregnancies subsequently complicated by FGR. Conclusions: This is the first study reporting a relationship between maternal Secretor/Lewis status and FGR. A link between blood groups and FGR is plausible, as both are associated with cardiovascular disease. We observed no relationship between Lewis/Secretor status and VWF or APCR, but this should be confirmed in a larger study. Thus, the mechanism whereby Secretor and/or Lewis influences FGR is unknown.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References

Fetal growth restriction (FGR) occurs in relation to a variety of disorders, but can be associated with thrombosis of the placental vasculature [1]. The products resulting from the complex interaction of the ABO(H), Lewis and Secretor genes are displayed on red cells and other cell types and as free substances in body secretions. There is convincing evidence that ABO(H) blood groups are associated with thrombotic risk, resulting, in part, from higher levels of von Willebrand factor (VWF) in those with blood groups other than O [2,3]. A role for Lewis/Secretor status in thrombosis has also been reported, but is more controversial [4–10], as is the link between Lewis or Secretor and VWF levels [11–14]. In addition to a potential link with thrombosis, isomers of the Lewis antigens play a role in tissue differentiation, inflammation [15], and infection risk [16], in particular functioning as a ligand for selectins [15]. Indeed, altered tissue expression of such isomers has also been associated with abnormal placental function [17]. It is therefore conceivable that these genes, which may alter the mother’s coagulation and/or inflammatory/infection response, could contribute to the development of FGR. To examine this hypothesis, Secretor, Lewis and ABO(H) genotypes and phenotypes were examined in a case–control study of FGR performed on a prospective cohort of pregnant women.

Methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References

Subjects for the current study were drawn from the Glasgow Outcome APC resistance and Lipid (GOAL) pregnancy study [18,19]. Ethical approval for the study was obtained from the local ethical committee at the Glasgow Royal Infirmary NHS Trust, Scotland. For the current investigation, FGR cases (n = 128) were women consecutively recruited to the GOAL study whose singleton pregnancy was delivered at ≥ 35 weeks of gestation and was complicated by severe FGR. Severe FGR was defined as a singleton pregnancy with a birthweight of ≤ 5th centile (normalized for maternal gestation, maternal parity, and fetal sex [20]). To try to maximize the uniformity of cases in the study, those women with FGR related to pre-eclampsia [21] were excluded. The genotype frequencies of this group were compared with those of a group of consecutively recruited singleton pregnancies from the same cohort, matched for age, parity, and smoking, who had uncomplicated pregnancies (n = 288). The number of subjects included was chosen to permit the study to have > 80% power (at P = 0.05) to determine an approximately two-fold increased FGR risk associated with positive Secretor status (i.e. the combined group of SeSe and Sese). The distributions of genotypes, phenotypes and allele frequencies in cases and controls were compared by the use of odds ratios (ORs) with 95% confidence intervals (CIs). Comparisons of normally distributed data were achieved by t-test, and those of non-normally distributed data by Mann–Whitney U-test (Prism 4; Graphpad Software Inc., La Jolla, CA, USA).

Sample handling [19], ABO(H), Lewis and Secretor genotyping [22–24] and activated protein C (APC) resistance (APCR) assessments [19] were performed as previously described. VWF antigen levels were determined, in duplicate, by ELISA (Corgenix UK, Peterborough, UK). APCR was assessed in cases and controls who were factor V Leiden-negative, and were sampled between 7 and 16 weeks of gestation (mean gestation 11 weeks; standard deviation [SD] 2 weeks). For APCR assessment, higher resistance to APC is indicated by a lower APC sensitivity ratio (APC:SR).

Results

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References

Blood groups and FGR

Descriptive information for both cases and controls is shown in Table 1. The groups were balanced with regard to their demographic characteristics. The overall cohort was in Hardy–Weinberg equilibrium with regard to the Secretor gene allelic frequencies.

Table 1.   Demographics of fetal growth restriction (FGR) subjects and controls
 FGR (n = 128)Controls (= 288)P
  1. The means (standard deviations) for age, body mass index, systolic blood pressure and diastolic blood pressure at first booking, maternal gestation at birth, birthweight centile and the number (%) of smokers, blood group O subjects and primigravid cases and controls are shown. The P-value for a two-sample t-test (for continuous variables) or Fisher’s exact test for each comparison is also shown.

Age (years)28 (6)28(6)0.84
Primigravida49 (38.3)105 (36.5)0.74
Smoker91 (70.1)194 (67.4)0.57
Body mass index (kg m−2)23.3 (4.4)24.3 (4.6)0.05
Booking systolic blood pressure108 (12)107 (10)0.28
Booking diastolic blood pressure64 (8)65 (8)0.10
O Blood group68 (53.1)154 (53.5)1.0
Gestation at birth40 (1.4)39 (1.4)0.002
Birthweight centile3 (1.6)50 (26.4)< 0.0001

As shown in Table 2, FGR was more common in secretors (OR 1.70, 95% CI 1.08–2.69) and correspondingly less common in non-secretors (i.e. sese genotypes). Consistent with being obligate secretors, those mothers who expressed Le(b) on their red cells had a similarly higher risk of FGR (OR 1.80, 95% CI 1.15–2.83), with the corollary of a reduced risk in those expressing Le(a) (Table 2). Interestingly, a significantly increased risk was also observed in those who were homozygous carriers of the Lewis gene (LeLe genotype, OR 3.46). However, this was accompanied by a wide 95% CI (1.10–10.90), and no relationship was observed when all Lewis carriers (i.e. the combined group of LeLe and Lele genotypes) were compared with non-carriers (lele), or when those secretors who were also carriers of LeLe were compared with non-secretor non-carriers (OR 2.97, 95% CI 0.35–25.30). For ABO(H), no significant relationship was observed between FGR and ABO(H) (Table 2), and no interaction between individual ABO(H) blood groups and Secretor/Lewis status was observed in relation to FGR (data not shown).

Table 2. Secretor, Lewis and ABO(H) genotypes and phenotype carriage in fetal growth restriction (FGR) subjects and controls
Genotype/phenotypeFGR +veControls +veFGR −veControls −veOR95% CI
  1. CI, confidence interval; OR, odds ratio. *Indicates a comparison of SeSe with sese only. Se genotype: PCR failure in two cases and five controls. †Indicates a comparison of LeLe with lele only. ‡Indicates statistically significant results. Le genotyping and phenotype: PCR failure (or no phenotype) in three cases and five controls. Non-O indicates a comparison of the combined group of A1A1, A1B, BB, A1A2, A2B, A1O1, BO1, BO2 and A1O2 with the combined group of O1O1, O1O2, O2O2, O1A2 and O2A2. No O indicates a comparison of the combined group of A1A1, A1B and BB with the combined group of O1O1, O1O2, O2O2, O1A2 and O2A2. ABO(H) genotyping: PCR failure in six cases and five controls.

SeSe*2352351121.410.76–2.63
SeSe/Sese91171351121.70‡1.08–2.69
sese35112911710.59‡0.37–0.93
LeLe72784153.46‡1.10–10.90
LeLe/Lele1212684151.690.55–5.21
lele4151212680.590.19–1.82
Le(b+)88161371221.80‡1.15–2.83
Le(a+)33107921760.59‡0.37–0.94
Le(a−b−)4151212680.590.19–1.82
Non-O47109751741.000.65–1.55
No O315751740.460.13–1.65

To investigate whether Secretor status influences fetal growth overall, the data were further explored to determine whether increasing birthweight is paralleled by a reduced OR of being a secretor (i.e. carrying either SeSe or Sese). On grouping of the normalized birthweights of the controls, and with the FGR group as the index, similar ORs for being a secretor were observed for the 6–25th (OR 0.47, 95% CI 0.26–0.88), 26–50th (OR 0.56, 95% CI 0.32–0.99) and 51–75th (OR 0.35, 95% CI 0.19–0.65) centile groupings. A non-significant result was evident for the 76–100th (OR 1.97, 95% CI 0.87–4.47) centile group comparison. Although the control group was not specifically designed for such an analysis, these results suggest that Secretor has its predominant influence on the risk of developing the pathologic state of severe growth restriction alone, rather than by influencing the whole spectrum of fetal growth.

Blood groups and APCR/VWF

Given the reported association of blood groups with both ‘acquired’ APCR [25] and VWF levels [2], post hoc studies to provide pilot data on the relationship between blood group genotypes and both the degree of ‘acquired’ APCR (in FV Leiden-negative subjects) and VWF levels were performed in those subjects where suitable maternal first-trimester plasma samples were available. Descriptive information on the subjects included in these analyses is shown in Table 3.

Table 3.   Demographics of fetal growth restriction (FGR) subjects and controls for (a) activated protein C resistance (APCR) assessment and (b) von Willebrand factor (VWF) assessment
(a)
 FGR (n = 86)Controls (n = 202)P
  1. The means (standard deviations) for age, body mass index, systolic blood pressure and diastolic blood pressure at first booking, maternal gestation at birth, birthweight centile and the number (%) of smokers, blood group O subjects and primigravid FGR cases and controls for the APCR (a) and VWF assessment (b) are shown. The P-value for a two-sample t-test (for continuous variables) or Fisher’s exact test for each comparison is also shown.

Age (years)28 (6)28 (6)0.93
Primigravida35 (40.7)72 (35.6)0.43
Smoker64 (74.4)135 (66.8)0.60
Body mass index (kg m−2)23.3 (4.4)24.0 (4.8)0.24
Booking systolic blood pressure108 (12)107 (10)0.66
Booking diastolic blood pressure64 (8)65 (8)0.30
O Blood group50 (58.1)109 (53.9)0.90
Gestation at birth39 (1.5)40 (1.3)0.0004
Birthweight centile3 (1.7)50 (26)< 0.0001
(b)
 FGR (n = 85)Controls (n = 205)P
Age (years)28 (6)28 (6)0.81
Primigravida34 (40.0)79 (38.5)0.89
Smoker58 (68.2)130 (63.4)0.50
Body mass index (kg m−2)23.4 (4.6)24.1 (4.6)0.20
Booking systolic blood pressure108 (13)108 (10)0.90
Booking diastolic blood pressure64 (9)66 (8)0.07
O Blood group45 (52.9)104 (50.7)0.80
Gestation at birth39 (1.5)40 (1.4)0.001
Birthweight centile3 (1.7)51 (25)< 0.0001

As expected [19], no effect of gestation was evident on APCR over this gestation interval. For all subjects, no significant relationships were observed between APC:SR and either Secretor genotypes (SeSe, n = 54, mean APC:SR 2.78, SD 0.43; SeSe/Sese, n = 169, mean APC:SR 2.77, SD 0.46; sese, n = 113, mean APC:SR 2.70, SD 0.38; all t-test comparisons, P > 0.05, six PCR failures), Lewis genotypes (LeLe, n = 166, mean APC:SR 2.77, SD 0.44; LeLe/Lele, n = 265, mean APC:SR 2.74, SD 0.44; lele, n = 17, mean APC:SR 2.76, SD 0.34; all t-test comparisons, P > 0.05, six PCR failures), or Lewis phenotypes (Le[a+], n = 106, mean APC:SR 2.69, SD 0.38; Le[a−b−], n = 17, mean APC:SR 2.76, SD 0.34; le[b+], n = 159, mean APC:SR 2.78, SD 0.47; all t-test comparisons, P > 0.05).

No relationship between sample gestation and VWF level was seen, and no significant differences in VWF antigen levels were observed when all secretors (SeSe/Sese, n = 169, median VWF 98 IU dL−1, interquartile range [IQR] 65–136) were compared with non-secretors (sese, n = 115, median VWF 101 IU dL−1, IQR 31–132, Mann–Whitney U-test, P = 0.60, six PCR failures), or when all Le(b+) subjects were compared with the remainder (Le[b+], n = 161, median VWF 87 IU dL−1, IQR 46–124; remainder, n = 124, median VWF 96 IU dL−1, IQR 66–132, Mann–Whitney U-test, P = 0.07, five phenotype failures). Similarly, when Lewis genotypes were compared, no significant differences in VWF antigen levels were seen (LeLe, n = 176, median 91 IU dL−1, IQR 51–126; LeLe/Lele, n = 269, median 91 IU dL−1, IQR 51–125; lele, n = 16, median 100 IU dL−1, IQR 76–185; all Mann–Whitney U-test comparisons, P > 0.05, five PCR failures).

Of interest, and despite the lack of an association of VWF with Secretor and Lewis groups, lower levels of VWF antigen were observed in those with FGR (median 61 IU dL−1, IQR 31–100) than in controls (median 99 IU dL−1, IQR 65–132, P < 0.001). To exclude undiagnosed VWF deficiency, the analysis was repeated, including only those with VWF levels above a quoted lower first-trimester reference limit for VWF antigen [26]. This produced similar results (FGR median 84 IU dL−1, IQR 52–114; control median 105 IU dL−1, IQR 72–138, P = 0.004).

Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References

As far as the authors are aware, this is the first study to report a relationship between maternal Secretor status and the occurrence of FGR. A link between blood groups and FGR is plausible, as both are associated with cardiovascular disease. In particular, a maternal family history of premature coronary heart disease is associated with the occurrence of FGR [27], and those with a personal history of FGR have a higher risk of poor cardiovascular health as adults [28]. Furthermore, women with an FGR pregnancy appear to have an increased future vascular disease risk [29–31]. Similarly, studies have linked Lewis/Secretor status with vascular disease, although the magnitude of the association is not yet clear. The majority of such studies have focused on Le(a−b−) subjects, comparing them with a combined group of Le(a+) and Le(b+) subjects. In these, Le(a−b−) has been associated with insulin resistance and other vascular risk factors in some (but not all [14]) studies in men [6–8], although not in women [8,14]. Furthermore, higher levels of VWF have been reported in group O subjects who are Le(a+) [11], although other studies have either observed no difference in VWF or FVIII levels between Le(a+) and Le(b+) subjects (irrespective of blood group [11]), or found higher VWF levels only in association with carriage of SeSe [23], or observed no relationship between FVIII or VWF antigen and Secretor status at all [12]. A higher level of cholesterol has also been reported in association with Le(a+) [32,33]. With specific regard to Lewis groups other than Le(a−b−), one uncontrolled study has reported a deficit of secretors (the majority of whom would be Le[b+]) in peripheral vascular disease [34], but other studies have shown no influence of Le(a+) on either myocardial infarction [9] or ischemic stroke [10] risk.

Despite the potential for a common link between FGR, blood groups, and thrombotic disease, we observed no association between ABO(H) genotype or phenotype and FGR. Lewis and Secretor are genetically independent, and although the vast majority of secretors will express Le(b) in their plasma, there will still be Le(a) in other secretions, although relatively little in the plasma. The current study indicates that non-Secretor status (and Le[a]) reduces the risk of FGR. The increased risk in association with homozygous Lewis carriage might suggest that there is a direct link between the amount of Lewis substance (predominantly Le[b]) and FGR risk. However, we did not observe a significant risk when all Lewis carriers, or only those LeLe carriers who were also secretors, were considered in isolation. This, however, may be a reflection of the numbers studied. Although allied Lewis substances are involved in cell interactions [16] and infection response [16], and may have altered placental expression in those with FGR [17], the mechanism whereby Secretor and/or Lewis influences FGR remains speculative.

We have previously reported an association between birthweights greater than the 90th percentile and the maternal carriage of FV Leiden mutation [18], indicating that large birthweights may also be associated with a maternal thrombotic tendency. Of interest, in the current study, we observed a non-significant positive OR for being a secretor for those with a birthweight ≥ 76th percentile when compared with FGR subjects. However, insufficient numbers of individuals of the highest birthweights were included in the current study to examine this fully, and the formal exclusion or confirmation of a significant association between very large birthweights and Secretor will require a study specifically designed to examine this birthweight group.

Changes in acquired ACPR in pregnancy do reflect fetal growth [35], although not the occurrence of growth restriction [19]. In the current study, no relationship between acquired APCR and Lewis or Secretor status was seen. In an attempt to further direct future research in this area, we performed a limited analysis that revealed a lower level of VWF in cases in the first trimester. However, despite the association of Secretor status with FGR, we did not observe a significant relationship between VWF and Secretor or Lewis status overall. Given this, the observation in one small study of no significant difference in FVIII levels (outwith pregnancy) in those with a history of isolated growth restriction [36], the post hoc nature of our pilot studies, and the age of our samples at the time of analysis, our observations on VWF require confirmation in studies with a larger sample size.

Acknowledgements

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References

We are indebted to M. Forgan, S. Wilson and J. Conkie for their technical assistance with genotyping and coagulation assays for the study. This research was supported by a grant from the Scottish Executive Health Department: Grant RO591CZG/1/83.

Disclosure of Conflict of Interests

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References

The authors state that they have no conflict of interest.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. Disclosure of Conflict of Interests
  9. References
  • 1
    Cox P, Marton T. Pathological assessment of intrauterine growth restriction. Best Pract Res Clin Obstet Gynaecol 2009; 23: 75164.
  • 2
    Wu O, Bayoumi N, Vickers M, Clark P. ABO(H) blood groups and vascular disease: a systematic review and meta-analysis. J Thromb Haemost 2008; 6: 629.
  • 3
    Clark P, Wu O. ABO blood groups and thrombosis: a causal association, but is there value in screening? Future Cardiol 2011; 7: 191201.
  • 4
    Djousse L, Karamohamed S, Herbert A, D’Agostino R, Cupples L, Ellison R. Fucosyltransferase 3 polymorphism and atherothrombotic disease in the Framingham Offspring Study. Am Heart J 2007; 153: 6369.
  • 5
    Hein HO, Suadicani P, Gyntelberg F. Lewis phenotypes, leisure time physical activity, and the risk of ischaemic heart disease: an 11 year follow up in the Copenhagen male study. Heart 2001; 85: 15964.
  • 6
    Hein S, Sorensen H, Saudicani P, Gyntelberg F. The Lewis blood group – a new genetic marker of ischaemic heart disease. J Intern Med 1992; 232: 4818.
  • 7
    Ellison R, Zhang Y, Myers R, Swanson J, Higgins M, Eckfeldt J. Lewis blood group phenotype as an independent risk factor for coronary heart disease (The NHLBI Family Heart Study). Am J Cardiol 1999; 83: 3458.
  • 8
    Clausen JO, Hein HO, Suadicani P, Winther K, Gyntelberg F, Pedersen O. Lewis phenotypes and the insulin resistance syndrome in young healthy white men and women. Am J Hypertens 1995; 8: 10606.
  • 9
    Saha N, Toh C, Ghosh M. Genetic association in myocardial infarction. Ethnicity, ABO, Rh, Le (a), Xg (a) blood groups, G6PD deficiency and abnormal haemoglobins. J Med Genet 1973; 10: 3405.
  • 10
    Clark P, Meiklejohn DJ, O’Sullivan A, Vickers MA, Greaves MJ. The relationships of ABO, Lewis and Secretor blood groups with cerebral ischaemia of arterial origin. J Thromb Haemost 2005; 3: 21058.
  • 11
    Orstavik K, Kornstad L, Reisner H, Berg K. Possible effect of secretor locus on plasma concentration of factor VIII and von Willebrand factor. Blood 1989; 73: 9903.
  • 12
    Schleef M, Erwin S, Dick A, Frank J, Schramm W, Spannagl M. Relationship between ABO and Secretor genotype with plasma levels of factor VIII and von Willebrand factor in thrombosis patients and control individuals. Br J Haematol 2005; 128: 1007.
  • 13
    Green D, Jarrett O, Ruth KJ, Folsom AR, Liu K. Relationship among Lewis phenotype, clotting factors, and other cardiovascular risk factors in young adults. J Lab Clin Med 1995; 125: 3349.
  • 14
    Cakir B, Heiss G, Pankow J, Salomaa V, Sharrett A, Couper D, Weston BW. Association of the Lewis genotype with cardiovascular risk factors and subclinical carotid atherosclerosis: the Atherosclerosis Risk in Communities (ARIC) study. J Intern Med 2004; 255: 4051.
  • 15
    Foxall C, Waltson S, Dowbenko D, Fennie C, Lasky L, Kiso M, Haseqawa A, As D, Brandly BK. The three members of the selectin receptor family recognize a common carbohydrate epitope, the sialyl Lewis (x) oligosaccharide. J Cell Biol 1992; 117: 895902.
  • 16
    Sheu B, Sheu S, Yang H, Huang A, Wu J. Host gastric Lewis expression determines the bacterial density of Helicobacter pylori in babA2 genopositive infection. Gut 2003; 52: 92732.
  • 17
    Minas V, Mylonas I, Scgiessl B, Mayr D, Schulze S, Friese K, Jeschke U, Makrigiannakis A. Expression of the blood-group-related antigens sialyl Lewis a, sialyl Lewis X and Lewis Y in term placentas of normal, preeclampsia, IUGR- and HELLP-complicated pregnancies. Histochem Cell Biol 2007; 128: 5563.
  • 18
    Clark P, Walker I, Govan L, Wu O, Greer I. The GOAL study: a prospective examination of the impact of factor V Leiden and ABO(H) blood groups on haemorrhagic and thrombotic pregnancy outcomes. Br J Haematol 2008; 140: 23640.
  • 19
    Clark P, Sattar N, Walker ID, Greer IA. The Glasgow outcome, APCR and lipid (GOAL) Pregnancy Study: significance of pregnancy associated activated protein C resistance. Thromb Haemost 2001; 85: 305.
  • 20
    Altman D, Coles E. Normograms for precise determination of birth weight for dates. Br J Obstet Gynaecol 1980; 87: 816.
  • 21
    Clark P, O’Sullivan A, Forgan M, Campbell DM, Greaves M, Walker ID, Greer IA, Vickers MA. ABO and secretor genotypes and the risk of pre-eclampsia. J Thromb Haemost 2005; 3(Suppl. 1): P0493.
  • 22
    Downing J, Darke C. A modified PCR-SSP method for the identification of ABO blood group antigens. Eur J Immunogen 2003; 30: 2958.
  • 23
    O’Donnell J, Boulton F, Manning R, Laffan M. Genotype at the secretor blood locus is a determinant of plasma von Willebrand factor level. Br J Haematol 2002; 116: 3506.
  • 24
    Vestergaard E, Wolf H, Orntoft T. Increased concentration of genotype-interpreted Ca 19-9 in urine of bladder cancer patients mark diffuse atypia of the urothelium. Clin Chem 1998; 44: 197204.
  • 25
    Clark P, Walker ID. The phenomenon known as acquired activated protein C resistance. Br J Haematol 2001; 115: 18.
  • 26
    Clark P, Brennand J, Conkie JA, McCall F, Greer IA, Walker ID. Activated protein C sensitivity, protein C, protein S and coagulation in normal pregnancy. Thromb Haemost 1998; 79: 116670.
  • 27
    Pell J, Smith G, Dominiczak A, Cobbe S, Dobbie R, McMahon A, Ford I. Family history of premature death from ischaemic heart disease is associated with an increased risk of delivering a low birth weight baby. Heart 2003; 89: 124950.
  • 28
    Barker D, Osmond C, Golding J, Kuh D, Wadsorth M. Growth in utero, blood pressure in childhood and adult life and mortality from cardiovascular disease. Br Med J 1989; 298: 5647.
  • 29
    Pell J, Smith G, Walsh D. Pregnancy complications and subsequent maternal cerebrovascular events: a retrospective cohort study of 119,668 births. Am J Epidemiol 2004; 159: 33642.
  • 30
    Berends A, de Groot C, Sijbrands E, Sie M, Benneheij S, Pal R, Hydanus R, Oostra BA, van Duijn CM, Steegers EAP. Shared constitutional risks for maternal vascular-related pregnancy complications and future cardiovascular disease. Hypertension 2008; 51: 103441.
  • 31
    Smith G, Harding S, Rosato M. Relation between infant’s birth weight and mother’s mortality: prospective observational study. Br Med J 2000; 320: 83940.
  • 32
    Langman M, Elwood P, Goote J, Ryrie D. ABO and Lewis blood groups and serum cholesterol. Lancet 1969; 2: 6079.
  • 33
    Wakely E, Langman MJ, Elwood P. Blood group A sub-groups and serum cholesterol. Cardiovasc Res 1973; 7: 67983.
  • 34
    Hall R, Bunch G, Humphrey C. The frequencies of ABO blood groups and secretors of ABH substances in peripheral atherosclerosis. Atheroscler 1971; 14: 2416.
  • 35
    Clark P, Walker I, Greer I. Acquired activated protein-C resistance in pregnancy and association with increased thrombin generation and fetal weight. Lancet 1999; 353: 2923.
  • 36
    Witsenburg C, Rosendaal F, Middeldorp J, van der Meer F, Scherjon S. Factor VIII levels and the risk of pre-eclampsia, HELLP syndrome, pregnancy related hypertension and severe intrauterine growth restriction. Thromb Res 2005; 115: 38792.