Apolipoprotein E4 and recurrent pregnancy loss: Is it time to draw a conclusion?

7 12 2009

To the Editor:

Lately, Bianca et al. revealed a negative association of apolipoprotein E4 (APOE4 ε) allele with recurrent pregnancy loss (RPL) (1), which was against the conclusion drawn by a previous study (2). The contradictory results lead us to believe that it is still too early to draw a conclusion about the association between APOE4 ε and RPL.

Although we agree that positive control groups should be included to reduce the influence of population-based polymorphism (3), we do think that a negative control group of sufficient sample size with healthy population is pivotal in their research.

Apolipoprotein E (apoE) is a glycosylated protein characterized by its wide tissue distribution and multiple functions. ApoE polymorphism has been extensively studied in a variety of diseases, such multiple sclerosis and Alzheimer’s disease. The potential effects of apoE polymorphism on human diseases seems educible considering its multiple properties, including immunomodulation and anti-oxidization (4).
ApoE genotype distribution may differ among populations, as reported by dozens of epidemiological studies. However, APOE4 ε allele frequency, especially the ratio of APOE4/4 genotype in healthy subjects, has been low (3, 5). The relatively low frequency of APOE4 ε allele in populations might influence the result interpretation by rendering the analyzers more apt to make type I errors. Thus a larger sample size in apoE polymorphism studies is a must. Moreover, the comparison of APOE4 ε carriers with healthy controls might not be able to exclude the influence of other APOE alleles, in that no evidence as yet has confirmed a similar or contrary effect among apoE isoforms.

The principal pitfall in the study by Goodman, et al was small sample size, which reduced the reliability of the conclusion. The enlarged sample size in the study by Bianca et al. might partially reduce the risk of type I error, which increased to some degree the reliability of the drawn conclusion, while lack of negative controls, on the contrary, might increase the risk of type II error.

In summary, it still seems too early, to draw a conclusion on the association of APOE4 ε and RPL. Further population-based, larger-scale studies, alongside with more detailed sub-classification of RPL, are to be implemented.

Hong-liang Zhang, M.D.a,b
Jiang Wu, M.D., Ph.D.a

aDepartment of Neurology
First Hospital of Jilin University
Changchun, China
bDepartment of Neurobiology
Care Sciences and Society
Karolinska Institute
Stockholm, Sweden

References
1. Bianca S, Barrano B, Cutuli N, Indaco L, Cataliotti A, Milana G, et al. No association between apolipoprotein E polymorphisms and recurrent pregnancy loss. Fertil Steril 2009 Published online. Doi:10.1016/j.fertnstert.2009.07.971.

2. Goodman C, Goodman CS, Hur J, Jeyendran RS, Coulam C. The association of apoprotien E polymorphisms with recurrent pregnancy loss. Am J Reprod Immunol 2009; 61: 34-8.

3. Gerdes LU, Klausen IC, Sihm I, Fægeman O. Apolipoprotein E polymorphism in a Danish population compared to findings in 45 other study populations around the world. Genetic Epidemiology 1992; 9: 155-167.

4. Kim J, Basak JM, Holtzman DM. The role of apolipoprotein E in Alzheimer’s disease. Neuron 2009; 63: 287-303.

5. Menzel HJ, Kladetzky RG, Assmann G. Apolipoprotein E polymorphism and coronary artery disease. Arteriosclerosis 1983; 3: 310-5.

Published online in Fertility and Sterility doi:10.1016/j.fertnstert.2009.12.014

The association of apolipoprotein E polymorphisms and recurrent pregnancy loss

To the Editor:

We read with interest the publication of Bianca et al. (1). While we agree with their findings, we disagree with their conclusion that their data are different from those previously reported by Goodman et al. (2) and that no association between Apo E polymorphisms and recurrent pregnancy loss exists.

Bianca et al. (1) reported the prevalence of Apo E3/3 and E4/4 genotypes among women with a history of recurrent pregnancy loss to be 14.3% and Goodman et al. (2) reported a not significantly different prevalence of 21.7% (p=0.18). Furthermore, Ergin et al. (3) reported a prevalence of Apo E3/3 and E4/4 genotypes among Turkish women experiencing recurrent pregnancy loss to be 19% (P=0.28 when compared with 14.3% of Bianca and P=0.71 when compared with 21.7% of Goodman). Thus the frequencies of Apo E3/3 and E4/4 genotypes are similar among the three populations from the United States, South America and Turkey.

Bianca et al. (1) compared their results in women experiencing recurrent pregnancy loss with women with a history of cardiovascular disease (CVD) and with women with a family history of inherited thrombophilia. The prevalences of Apo E3/3 and E4/4 genotypes among their positive control groups were 16% for CVD and 24% for women with a family history of thrombophilia. Based on no difference between their study patients and positive control groups, Bianca et al. (1) concluded that there was “no association between apolipoprotein E polymorphisms and recurrent pregnancy loss.” However, to come to that conclusion, appropriate negative controls are necessary. Bianca et al. (1) reported no negative controls. Negative controls were reported with the Goodman et al. (2) and Ergin et al. (3) studies. The prevalence of Apo E3/3 and E4/4 genotypes among women with at least two live births and no history of thrombophilia was 5% (2,3). When we looked at the frequency of Apo E3/3 and E4/4 genotypes among our positive controls of 59 individuals with a history of deep vein thrombosis (DVT), 10 or 17% displayed Apo E3/3 and E4/4 genotypes. Figure 1 summarizes the frequencies of Apo E3/3 and E4/4 genotypes among women with a history of recurrent pregnancy loss and their positive and negative controls. The only significant differences seen among the groups are between the women experiencing recurrent pregnancy loss and their fertile (negative) controls.

Figure 1 (Click on figure to enlarge)

In conclusion, Apo E4 polymorphisms may contribute to the thrombogenic risk factors contributing to recurrent pregnancy loss. However, we have previously demonstrated a cumulative effect of thrombogenic mutations on the risk of recurrent miscarriage (4).
Thus, inherited thrombophilia represents multiple gene defects rather than any single gene defect. The challenge is to develop of a panel of markers to identify risk factors for women experiencing recurrent pregnancy loss that would be most helpful in suggesting effective treatment.

Chelsi Goodman, B.S.
Rajasingam S Jeyendran, D.V.M., Ph.D.
Carolyn B Coulam, M.D.
Rinehart Center for Reproductive Medicine
Evanston, Illinois

References
1. Bianca S, Barrano B, Cutuli N, Indaco L, Cataliotti A, Milana G, et al. No associationbetween apolipoprotein E polymorphisms and recurrent pregnancy loss. Fertil Steril 2009 Published online. Doi:10.1016/j.fertnstert.2009.07.971.

2. Goodman C, Goodman CS, Hur J, Jeyendran RS, Coulam C. The association of apoprotien E polymorphisms with recurrent pregnancy loss. Am J Reprod Immunol 2009;61: 34-8.

3. Ergin E, Jeyendran RS, Ozornek H, Ozay A, Pillai MD, Coulam C. Apoprotein E codon 112 polymorphism is associated with recurrent pregnancy loss. Fertil Steril 2009;in press.

4. Coulam CB, Jeyendran RS, Fishel LA, Roussev RG. Multiple thrombophilic gene mutations rather than specific gene mutations are risk factors for recurrent miscarriage. Am J Reprod Immunology 2006;55:360-8.

Published online in Fertility and Sterility doi:10.1016/j.fertnstert.2009.12.016

The Authors Respond:

We thank Zhang and Wu and Goodman et al. for their comments regarding our paper on the negative association between apolipoprotein E (APOE) E4 allele and recurrent pregnancy loss (RPL) (1). ApoE genotype distribution may differ among populations and among different studies based on different genetic background and selected samples.

In our study, we compared the RPL group with two high-risk control groups using our database of molecular tests performed only for diagnostic use and not for research purposes. This is the reason that we do not have healthy controls. Moreover, to answer to the criticisms raised by Zhang and Wu and by Goodman et al. about the absence of a healthy population, we collected a control sample of 1,990 healthy residents of different Italian regions from Italian published studies on APOE polymorphism (2-5). We selected only Italian studies to reduce the possible allelic frequency variability among different populations. The presence of APOE4 allele was reported in 274 healthy subjects (13.76%; range, 9.1 – 21.2%), close to the 14.3% reported incidence in our RPL group and not statistically significantly different from our previously reported two high-risk control groups.

Data in the literature have demonstrated that APOE polymorphisms represent a genetic risk factor for cardiovascular and neurological disease, but a biological hypothesis for a placental vascular damage in presence of APOE4 polymorphism, to our knowledge, has never been reported. Moreover, we agree that further population-based, larger-scale studies in different populations worldwide are needed to add more data to define the possible association between RPL and APOE and the relative patient’s clinical management. We also agree that a cumulative effect of thrombogenic mutations on the risk of recurrent miscarriage exists; thus, inherited thrombophilia represents multiple gene defects rather than any single gene defect. Moreover, based on our previous data (1), also sustained by these additional results, we confirmed our point of view on the clinical management in women with recurrent pregnancy loss that should not be influenced by the presence or not of APO E polymorphisms.

Sebastiano Bianca, M.D., Ph.D.a
Barbara Barrano, M.D., Ph.D.a
Nunzio Cutuli, M.D.b
Lara Indaco, M.D.a
Antonella Cataliotti, M.D.a
Gabriella Milana, M.D.b
Chiara Barone, M.D.a
Giuseppe Ettore, M.D.c

aCentro di Consulenza Genetica e di Teratologia della Riproduzione Laboratorio di Citogenetica
Dipartimento Materno Infantile
ARNAS Garibaldi Nesima
Catania, Italy
bLaboratorio di Genetica Molecolare
AOU Policlinico
Catania, Italy.
cUOC Ginecologia e Ostetricia
Dipartimento Materno Infantile
ARNAS Garibaldi Nesima
Catania, Italy

References
1. Bianca S, Barrano B, Cutuli N, Indaco L, Cataliotti A, Milana G, et al. No association between apolipoprotein E polymorphisms and recurrent pregnancy loss. Fertil Steril 2009 Published online. Doi:10.1016/j.fertnstert.2009.07.971.

2. Margaglione M, Seripa D, Gravina C, Grandone E, Vecchione G, Cappucci G, et al. Prevalence of apolipoprotein E alleles in healthy subjects and survivors of ischemic stroke: an Italian Case-Control Study. Stroke. 1998 Feb;29(2):399-403.

3. Orsitto G, Seripa D, Panza F, Franceschi M, Cascavilla L, Placentino G, et al. Apolipoprotein E genotypes in hospitalized elderly patients with vascular dementia. Dement Geriatr Cogn Disord. 2007;23(5):327-33.

4. Corbo RM, Ulizzi L, Piombo L, Scacchi R. Study on a possible effect of four longevity candidate genes (ACE, PON1, PPAR-gamma, and APOE) on human fertility. Biogerontology. 2008 Oct;9(5):317-23.

5. De Blasi S, Montesanto A, Martino C, Dato S, De Rango F, Bruni AC, et al. APOE polymorphism affects episodic memory among non demented elderly subjects. Exp Gerontol. 2009 Mar;44(3):224-7.

Published online in Fertility and Sterility doi:10.1016/j.fertnstert.2009.12.015

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