The presence of HLA-antibodies in recurrent miscarriage patients is associated with a reduced chance of a live birth

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Abstract

Anti-paternal HLA-antibodies are considered a harmless phenomenon during most pregnancies, whereas their role in recurrent miscarriage (RM) patients is disputed. In contrast to primary RM, patients with secondary RM have carried a fetus to term pregnancy prior to a series of miscarriages, which increases the chance that allogeneic fetal cells appear in the maternal circulation. This study investigates the frequency of HLA-antibodies in secondary RM, primary RM patients and parous controls and analyzes whether the presence of HLA-antibodies in early pregnancy is associated with pregnancy outcome. Sera from women with secondary RM (n = 56), primary RM (n = 13) and parous controls (n = 24) were tested for HLA-antibodies using an ELISA assay and complement dependent cytotoxicity. Samples were taken at gestational week 4–5 in 62 (90%) of the patients. HLA-antibodies were significantly more frequent in secondary RM patients with a boy prior to the miscarriages (62%) compared to secondary RM patients with a firstborn girl (29%, p = 0.03), primary RM patients (23%, p = 0.02) and parous controls (25%, p = 0.005). Forty-one percent of HLA-antibody positive pregnant RM patients had a live birth compared to 76% of HLA-antibody negative RM patients, p = 0.006 (adjusted OR: 0.22 (0.07–0.68), p = 0.008). In conclusion, HLA-antibodies are significantly more frequent in secondary RM patients with a firstborn boy than in other RM patients and controls. The presence of these antibodies in early pregnancy is associated with a reduced chance of a live birth. Further exploring this association may increase our understanding of maternal acceptance of the fetal allograft.

Introduction

Fetal acceptance by the maternal immune system remains an intriguing question and so does immune-mediated pregnancy losses (Laird et al., 2003). Although miscarriage is the most common complication of pregnancy, 40–80% of these losses are due to random chromosomal abnormalities incompatible with survival (Morales et al., 2008), which impedes the search for immune-mediated causes in humans. However, women with recurrent miscarriage (RM) are an ideal population for studying possible immune-mediated pregnancy losses as the frequency of abnormal embryonic karyotypes significantly decreases with the number of previous miscarriages (Ogasawara et al., 2000). Secondary RM, in contrast to primary RM, is preceded by a pregnancy lasting to the last trimester (Rai and Regan, 2006). Traffic of allogeneic fetal cells into the mother's circulation peaks in the last trimester (Adams et al., 2007, Huppertz et al., 2006), which may lead to activation of the maternal adaptive immune system (Verdijk et al., 2004). We have reported that the incidence of secondary RM is higher after delivery of a boy compared to a girl and the chance of a subsequent live birth is significantly reduced in patients with a firstborn boy compared to a girl (Christiansen et al., 2004, Nielsen et al., 2008) suggesting a role for abnormal immune responses against male-specific minor histocompatibility (H–Y) antigens (Nielsen et al., 2009).

The relevance of maternal immune responses against the paternal HLA-antigens of the fetus in pregnancy was first debated half a century ago (PAYNE and ROLFS, 1958, van Rood et al., 1958). Studies of anti-paternal HLA-antibodies in primary RM patients reported a low frequency of these antibodies, which fostered the hypothesis that lack of anti-paternal antibodies in these patients was caused by a failure to achieve an appropriate immunological response toward the fetus (McIntyre et al., 1984, Regan et al., 1991, Sargent et al., 1988, Tongio et al., 1972). No correlation between anti-paternal antibodies and pregnancy success was reported in a small prospective study including 12 primary RM and 6 secondary RM patients (Sargent et al., 1988). It therefore remains unknown whether HLA-antibodies play a role in patients with RM. In this study we investigated the frequency and specificity of HLA-antibodies in RM patients categorized as patients with primary RM, secondary RM with a firstborn boy, or secondary RM with a firstborn girl, as well as a cohort of women with a history of two prior uncomplicated pregnancies. Furthermore, we correlated the presence of HLA-antibodies in early pregnancy to the incidence of miscarriage or live birth.

Section snippets

Recurrent miscarriage

Patients with unexplained RM referred to the Danish Recurrent Miscarriage clinic from July 2004 to March 2008 were invited to participate in this study. RM was defined as three or more consecutive losses of intrauterine pregnancies before the 22nd gestational week. The primary RM diagnosis was used for patients with no history of a live or stillbirths while secondary RM was defined as RM subsequent to at least one pregnancy of minimally 22 weeks of gestation. The miscarriages were considered to

Results

A total of 93 serum samples were tested for antibodies to HLA-classes I and II antigens. The frequencies of HLA-classes I and II antibody positive individuals were compared between the subgroups of included women. The association between the presence of HLA-antibodies in early pregnancy and pregnancy success was tested in the RM patients who had serum samples taken in early pregnancy.

Discussion

In this study on the role of HLA-antibodies in patients with unexplained RM we found a significantly higher prevalence of HLA-antibodies in secondary RM patients with a boy prior to the miscarriages compared to secondary RM patients with a firstborn girl, primary RM patients and healthy parous women. In addition, a significantly reduced chance of a live birth in HLA-antibody positive RM patients was found.

The major limitation of this study is that almost all secondary RM and primary RM patients

Acknowledgements

This study was sponsored by the Dutch National Reference Center for Histocompatibility, the Netherlands Organization for Scientific Research (NWO) rewarded a travel grant for HSN; and the University Hospital Copenhagen Rigshospitalet rewarded a PhD-grant for HSN.

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