Recurrent miscarriage: a defect in nature’s quality control?

Siobhan Quenby1, Gill Vince2, Roy Farquharson3 and John Aplin4,5

1 Department of Obstetrics and Gynaecology, 2 Department of Immunology, University of Liverpool, L69 3BX, 3 Liverpool Women’s Hospital, Crown Street, Liverpool L8 7SS and 4 Medical School and School of Biological Sciences, University of Manchester, Research Floor, St Mary’s Hospital, Manchester M13 OJH, UK


    Abstract
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
Recent data on recurrent miscarriage (RM) is discussed in the framework of the selection failure hypothesis which states, ‘Recurrent miscarriage is the result of failure of the prevention of ‘poor quality’ embryos implanting, allowing embryos that are destined to fail to implant and present clinically as recurrent miscarriage. Thus, recurrent miscarriage is a failure of nature’s quality control.’ The assumption that RM results from the maternal rejection of normal fetuses is challenged and evidence reviewed regarding the contribution of abnormal embryos and endometrial receptivity. Further research is needed to understand the mechanisms of maternal tract–embryo interaction and move towards improved management of recurrent pregnancy loss.

Key words: abnormal pregnancies/embryonic period of gestation/endometrial receptivity/recurrent miscarriage/recurrent pregnancy loss


    Introduction
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
For many years, it has been assumed that recurrent miscarriage (RM) results from the maternal rejection of normal fetuses. However, recent evidence from clinical, epidemiological and basic science has challenged this assumption. This new evidence will be reviewed and we will argue that a new paradigm is needed to explain RM.

The current definition of RM is three consecutive pregnancy losses before week 24 of pregnancy. This definition was derived from the following epidemiological data. The sporadic miscarriage rate in the general population is 15% (Warburton and Frazer, 1964Go; Poland et al., 1977Go; Stray-Pederson and Stray-Pederson, 1984Go) so that, by probability alone, the RM rate in fertile couples would be 0.153 = 0.3–0.4%. The actual prevalence of RM in all fertile couples of reproductive age is 0.5–1.0% (Tho et al., 1979Go; Alberman, 1988Go). Thus, it has been assumed that some couples are more prone to miscarriage than others. This hypothesis is supported by studies showing a positive correlation between the number of previous miscarriages and the miscarriage rate in the next pregnancy in women who have had at least three miscarriages (Parazzini et al., 1988Go; Regan et al., 1989Go; Quenby and Farquharson, 1993Go).

Epidemiologically, advanced maternal age is a strong risk factor for both spontaneous miscarriage and RM, implying that pregnancy abnormality is a significant contributory factor to miscarriage (Quenby and Farquharson, 1993Go; Nybo Andersen et al., 2000Go).


    The contribution of abnormal pregnancies to recurrent pregnancy loss
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
One of the most remarkable, and as yet unexplained, aspects of the first trimester of pregnancy is the fact that the majority (90%) of karyotypically abnormal pregnancies miscarry in the first trimester and the majority (93%) of karyotypically normal pregnancies continue (McFadyen, 1989Go). Hence, miscarriage can be viewed as a natural process of quality control. Many of the miscarriages from RM patients are of karyotypically abnormal fetuses. Chorionic villous karyotyping, performed on the next miscarried pregnancy following three or more pregnancy losses, revealed abnormality in 60% of cases (Stern et al., 1996Go). Conventional cytogenetic analysis depends on culturing cells prior to karyotyping. Miscarried tissue from women suffering RM has shown a 29–57% abnormality rate (Stern et al., 1996Go; Ogasawara et al., 2000Go; Carp et al., 2001Go; Stephenson et al., 2002Go). However, this technique is limited by external contamination, culture failure and selective growth of maternal cells (Goddijn and Leschot, 2000Go). Analysis of spontaneous miscarriages by comparative genomic hybridization, a technique that detects chromosomal imbalances without the need for culture, has suggested that culture techniques underestimate the incidence of chromosomal anomalies (Daniely et al., 1998Go; Fritz et al., 2001Go; Stephenson et al., 2002Go).

Recently, preimplantation genetic diagnosis (PGD) has been used to investigate women with RM, revealing that they produce more aneuploid embryos than normal women (Simón et al., 1998Go; Vidal et al., 1998Go; Pellicer et al., 1999Go; Gianaroli et al., 2000Go). When PGD is used to discard karyotypically abnormal embryos (known as aneuploidy screening), the miscarriage rate after IVF decreases (Munné et al., 1999Go). The majority of these aneuploid embryos are a result of maternal oocyte abnormalities (Cobo et al., 2001Go; Robinson et al., 2001Go). Ovarian hyperstimulation increases the rate of oocyte abnormality (London et al., 2000Go). Recent publications have also suggested an association between sperm chromosomal anomalies (and sperm DNA damage) and RM (Giorlandino et al., 1998Go; Rubio et al., 1999Go; Egozcue et al., 2000Go). Thus there may well be a paternal contribution to recurrent embryonic abnormality. Therefore, artificial reproductive technology has highlighted the importance of embryo quality in miscarriage. However, the question remains as to why couples who recurrently produce abnormal embryos do not present with infertility because of recurrent subclinical pregnancy loss.

Karyotyping does not detect pregnancies complicated by structural abnormalities in the presence of morphologically normal chromosomes, or many other types of abnormality. Newly introduced medical therapeutic abortion methodology has allowed detailed examination of undamaged first trimester pregnancies (Blanch et al., 1998Go). Severe structural abnormalities likely to be incompatible with survival into the second trimester were found in 34% of specimens examined (Blanch et al., 1998Go). Had these pregnancies continued they would have represented sporadic miscarriages. The structural abnormalities in our data (Blanch et al., 1998Go) and the karyotype abnormalities discussed above represent slightly different groups, but together they indicate that embryo abnormality is a prominent feature of miscarriage.

Screening for known causes of RM includes tests for antiphospholipid syndrome, endocrine abnormalities, parental chromosome abnormalities, uterine structural abnormalities, thyroid function, diabetes and thrombophilias (Li, 1998Go).


    The endometrium in recurrent miscarriage
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
Studies of abortion-prone mice have highlighted the importance of maternal leukocytes and the cytokine response to fetal trophoblast (Clark et al., 1999Go); however, these studies relate to the miscarriage of normal pregnancies in specific mouse strains. In humans the situation is different because the majority of miscarriages are of abnormal pregnancies. Although it is clear that endometrial differentiation is essential for the acquisition of a receptive state, the ranges that have been observed in endometrial parameters in fertile women (Serle et al., 1994Go; Meyer et al., 1999Go) imply that some considerable variation is compatible with successful implantation. There is accumulating evidence that the endometrium in humans may even be more receptive to implantation in women with RM than in normal controls.

Expression of the endometrial protein, MUC1, was found to be lower in the endometrium of women suffering from RM than controls (Aplin et al., 1996Go), suggesting that epithelial function may be compromised in some cases of RM. MUC1 is an anti-adhesion molecule that is thought to contribute to the barrier function of the endometrial surface, which resists implantation except during a specific time window in the mid-secretory phase (Bergh and Navot, 1992Go; Aplin, 2000Go). In-vitro implantation experiments have demonstrated that MUC1 is selectively removed from the human maternal epithelial cell surface in the immediate vicinity of an attaching embryo (Aplin et al., 2001Go; Meseguer et al., 2001Go). This appears to depend on an embryo to maternal signalling mechanism, the details of which have not been elucidated. However, any process that is initiated by a signal emanating from the embryo can be put to use in selection. It is stated that epithelial abnormality with impairment of barrier function could lead to a situation in which ‘poor quality’ embryos that are otherwise destined to fail, implant and present later as miscarriages (Aplin et al., 1996Go).

The most abundant leukocyte in the maternal decidua is the uterine natural killer cell, also known as the uterine large granular lymphocyte (LGL). This uterine-specific cell type is more abundant in decidua than in non-pregnant endometrium (Bulmer, 1996Go). Lachapelle and colleagues demonstrated that the endometrium of (non-pregnant) women with RM contains an increased proportion of CD16+ CD56dim LGL compared with normal fertile women in whom CD16- CD56bright LGL predominate (Lachapelle et al., 1996Go). Using immunohistochemistry, increased numbers of CD56+ LGL were found in the preimplantation endometrium of RM patients compared with controls (Clifford et al., 1999Go; Quenby et al., 1999Go). Furthermore, there were more CD56+ leukocytes in the endometrium of patients who subsequently miscarried than in those who had live births (Quenby et al., 1999Go). There is no general agreement as to the function of uterine CD56+ LGL in relation to pregnancy, but these observations suggest that they might facilitate the implantation of blastocysts, including those with abnormal karyotypes, leading to the clinical presentation of RM. The latter interpretation is supported by data showing that CD56+ LGL are more numerous in the decidua from chromosomally abnormal miscarriages than in chromosomally normal miscarriages (Yamamoto et al., 1999Go). However, until the phenotype of these cells is further defined, the possibility that RM is associated with an altered LGL phenotype cannot be refuted.

T helper and other immune cells can differentiate into subsets with distinctive patterns of cytokine release, and it has been proposed that type 1 responses [characterized by the production of interleukin (IL)-2 and interferon-{gamma}] are systemically suppressed in murine pregnancy and that local expression of type 2 cytokines (e.g. IL-4, IL-6, IL-10) in placental tissue might be beneficial for fetal survival (Wegmann et al., 1993Go). During human pregnancy, systemic maternal immune responses may be biased in favour of a type 2 profile (Wegmann et al., 1993Go; Marzi et al., 1996Go) and it has been suggested that RM is an abnormal type 1 response (Makhseed et al., 1999Go; Raghupathy et al., 1999Go, 2000Go). Peri-implantation endometrium was found to have a predominance of type 2 cytokines (Krasnow et al., 1996Go; Lim et al., 1996Go, 1998Go), and in pregnancy a 10-fold increase in decidual type 2 cytokine secretion occurred (Krasnow et al., 1996Go). We prospectively studied the early pregnancy cytokine profile of women suffering RM. This study was designed to elucidate whether a failure of the cytokine shift predated miscarriage and was therefore likely to be an aetiological factor in RM. Contrary to our hypothesis, the cytokine shift, which appears to characterize normal pregnancy, was accentuated rather than diminished in RM pregnant women (Bates et al., 2002Go). Therefore, the failure of type 1/type 2 shift seen by other authors (Raghupathy et al., 2000Go) may be a consequence rather than a cause of miscarriage. In our RM population, women appear immunologically more receptive to abnormal pregnancies.

There were more activated leukocytes in the decidua from the miscarriages of women with unexplained RM and a normal fetal karyotype than in decidua from women with RM and a trisomy 16 fetus (Quack et al., 2001Go). This suggests different cellular immunity associated with the miscarriage of karyotypically normal and abnormal pregnancies, and indeed a possible maternal immunological aetiology for the failure of chromosomally normal conceptuses. Our view is that T lymphocytes, LGL and other immune cells may be capable of expressing either a progestational or contragestational phenotype and, crucially, may be able to switch rapidly from the former to the latter in response to an abnormal gestation. Testing this proposal requires a better grasp of the functions of each category of cells so that specific phenotypic markers can be measured in healthy and RM tissue.

At present, we argue that the immunological data do not support a case for the endometrium being either hostile or defective in RM. They are equally consistent with a higher rate of abnormal embryos in RM women being further compounded by reduced endometrial selectivity, and thus presenting clinically as RM rather than infertility. Further evidence needs to be gathered to test this hypothesis in the future.


    Ultrasound perspective
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
The majority of miscarriages occur in the embryonic period (5–9 weeks gestation, 3–7 weeks post-ovulation) rather than the fetal period (>10 weeks gestation, and hence after the visualization of fetal heart activity) (Goldstein, 1994Go). Of miscarriages occurring prior to fetal heart visualization, 87% are karyotypically abnormal (Burgoyne et al., 1991Go). Branch and Silver suggested that fetal deaths are more likely to have a maternal aetiology, e.g. anti-phospholipid syndrome (APS), whilst embryonic deaths are more likely to be due to embryonic anomalies (Branch and Silver, 1996Go). Maternal causes were indeed more common amongst women suffering at least one second trimester loss, compared with women suffering only first trimester losses (Drakeley et al., 1998Go). The Liverpool Miscarriage Clinic has a pregnancy loss rate of 27% of which 23% are embryonic losses and 4.8% fetal losses (Bricker and Farquharson, 2002Go). Thus we suggest that the majority of losses in a miscarriage clinic setting are due to embryonic abnormality.

Improvements in the resolution of ultrasonography should enable many of the structural and karyotypic abnormalities seen macroscopically to be diagnosed in utero. Anembryonic pregnancies, seen as empty sacs on ultrasound, are clearly abnormal. Cystic hygromas, anencephaly, body stalk defects and even lethal dwarfism can also all be recognized prior to miscarriage with modern ultrasonography (Jurkovic and Jauniaux, 1995Go).

Clinical observation points to the causes of miscarriage being fetal rather than placental, as the ultrasound identification of embryo/fetal death often occurs many weeks prior to clinical signs of pregnancy failure due to placental malfunction. Pregnancies with placental mosaicism (containing both karyotypically normal and abnormal trophoblast cells) and a normal fetal karyotype are associated with intrauterine growth restriction rather than miscarriage (Wilkins-Haug et al., 1995Go). However, the proliferative and invasive characteristics of chromosomally abnormal trophoblast are unknown.


    The selection failure hypothesis
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
A new framework is needed in which to consider RM. The selection failure hypothesis was first suggested by Aplin et al. that recurrent miscarriage is the result of failure of the prevention of ‘poor quality’ embryos implanting, allowing embryos that are destined to fail to implant and present clinically as recurrent miscarriage. Thus, recurrent miscarriage is a failure of nature’s quality control (Aplin et al., 1996Go).

Miscarriage of an abnormal pregnancy in the first trimester incurs significant cost to maternal resources and often requires time for physical and mental recovery during which the individual is reproductively inactive. Therefore, in the context of a high rate of abnormal embryos seen in humans, active selection by the female tract in the early stages of pregnancy, that is at fertilization, gamete transport, implantation and corpus luteal rescue, is the best way for the biological system to maximize reproductive success.


    Clinical implications
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
Women with APS followed prospectively have been shown to have an excess number of embryonic deaths on aspirin when compared with aspirin and heparin treatment (Rai et al., 1997Go). It is unlikely that either APS or any of the other thrombophilias cause abnormal embryos. Therefore, it would be more effective to screen women who have miscarried one normal pregnancy for APS than those who miscarry three or four abnormal pregnancies. In the former case, treatment with aspirin and heparin in the second pregnancy should prevent the women having to suffer three miscarriages prior to the diagnosis of a treatable condition. In the latter case, recognition of the embryonic abnormalities causing the miscarriages would prevent unnecessary treatment with heparin and its possible side-effects of bruising, thrombocytopenia and osteoporosis.

At present, karyotyping the products of conception following miscarriage may be viewed in some hospitals as an unnecessary luxury. However, in the absence of karyotyping, it is assumed that women who repeatedly miscarry are losing normal pregnancies. Data detailed above have shown this unlikely to be the case. Women with RM are given expensive, intensive treatment (for example, heparin or i.v. immunoglobulin) to prevent miscarriage. Such treatment is not appropriate if the losses are due to karyotypic or structural abnormalities. Therefore, obtaining a karyotype after several pregnancy losses is of extreme importance.


    Implications for treatment trials
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 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
If new randomized controlled treatment trials are designed to prevent RM, it must be remembered that many pregnancies will be lost due to embryonic abnormalities. Thus large study groups are required, small benefits are difficult to detect, and trials may generate conflicting results depending on the numbers of couples that are losing abnormal pregnancies in each group. The selection failure hypothesis should overcome these problems by justifying inclusion of only those couples who have lost normal pregnancies in trials aimed at maintaining normal pregnancy. Similarly, any treatment aimed at improving the rate of normal pregnancy in couples suffering recurrent aneuploidy should include only couples known recurrently to lose abnormal pregnancies.


    Implications for future research
 Top
 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
Future research is needed to identify signals from the embryo that overcome the maternal barrier to implantation. It is possible that a trisomic embryo could over-produce these factors and thus breach the epithelium inappropriately. It is reasonable to postulate that some endometria are less discerning in embryo selection, through posing a more easily penetrated barrier (discussed above) or perhaps more robust production of embryotrophic growth factors (i.e. those that promote synthesis of embryo-derived signals). A more complete understanding of the molecules mediating maternal tract–embryo interaction is required to address these possibilities. It will also be important to identify mechanisms at the materno–fetal interface that select normal and reject abnormal pregnancies after implantation has begun.


    Conclusions
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 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
In order to improve both the management of and the research into recurrent pregnancy loss, it is important to differentiate between couples recurrently miscarrying abnormal pregnancies and those miscarrying normal pregnancies. Improving the rate of implantation of high quality embryos is the most appropriate objective for assisted reproductive technologies.


    Acknowledgements
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 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
We are grateful to Drs Daniel Brison and Sarah Robertson for their very helpful comments.


    Notes
 
5 To whom correspondence should be addressed. E-mail: john.aplin{at}man.ac.uk Back


    References
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 Abstract
 Introduction
 The contribution of abnormal...
 The endometrium in recurrent...
 Ultrasound perspective
 The selection failure hypothesis
 Clinical implications
 Implications for treatment...
 Implications for future research
 Conclusions
 Acknowledgements
 References
 
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