Hum. Reprod. Advance Access published online on October 16, 2008
Human Reproduction, doi:10.1093/humrep/den377
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Mannose-binding lectin-2 genotypes and recurrent late pregnancy losses
1 Fertility Clinic 4071, Rigshospitalet, Copenhagen University Hospital, Copenhagen, Denmark 2 Department of Clinical Immunology, Aalborg Hospital, Aarhus University, Aalborg, Denmark
3 Correspondence address. E-mail: obc{at}pregnancyloss.dk, rh08636{at}rh.dk
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
BACKGROUND: Low levels of mannose-binding lectin (MBL) predispose to various infectious and inflammatory disorders and have been reported to be associated with recurrent early miscarriages. Recurrent late pregnancy losses (RLPL) in the second trimester is a rare but devastating syndrome where maternal rather than fetal causes are likely to play a stronger role than in early recurrent miscarriage.
METHODS: We identified 75 patients with at least two late losses of pregnancies with apparently normal fetuses between gestational week 14 and 30 among patients with recurrent pregnancy losses referred to our clinic. Polymorphisms in the MBL2 gene associated with plasma MBL levels were investigated in all patients and in 104 women with two or more children and no miscarriages. The patients were divided into three groups: one with clinical signs of cervical insufficiency, one positive for the lupus anticoagulant (LAC) and an idiopathic group.
RESULTS: Among all patients with RLPL, 26.7% had MBL2 genotypes associated with MBL deficiency compared with 12.5% in controls [odds ratio (OR) 2.55; 95% confidence interval (CI) 1.17–5.52; P < 0.02]. Among patients with clinical signs of cervical insufficiency or the LAC, the frequency of genotypes associated with MBL deficiency was not significantly increased. However, among 38 patients with idiopathic RLPL, 36.8% carried low-producing MBL2 genotypes, which was significantly more than in controls (OR 4.08, 95% CI 1.70–9.83, P = 0.001).
CONCLUSIONS: MBL deficiency is strongly associated with idiopathic RLPL. This may point towards a role for excessive inflammatory disturbances as a cause of the syndrome.
Key words: cervical insufficiency/mannose-binding lectin/miscarriage/recurrent miscarriage/stillbirth
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Mannose-binding lectin (MBL) is a plasma protein produced in the liver. It is a member of the collectin family of proteins and binds to a series of specific repetitive carbohydrate structures on microbial surfaces and subsequently activates the complement cascade through the so-called MBL pathway or promotes phagocytosis of the micro-organisms by macrophages.
MBL levels are genetically determined by three major single-nucleotide polymorphisms (SNPs) in exon 1 of the MBL2 gene on chromosome 10q11 and by three SNPs in the promoter region of the gene. Combinations of these SNPs are associated with very low plasma MBL levels (MBL deficiency) that are present in 10–12% of the Caucasian population. This prevalence makes MBL deficiency the most frequent immunodeficiency known. Children and immunocompromised adults with MBL deficiency are at an increased risk of recurrent infections and several autoimmune diseases seem to exhibit a more severe course in individuals with MBL deficiency.
Several studies have reported that MBL deficiency is predisposing to recurrent miscarriage (Kilpatrick et al., 1995
; Christiansen et al., 1999; Kruse et al., 2002), but there is no complete consensus about this (Baxter et al., 2001). This lack of consensus may be caused by the fact that the majority of pregnancy losses among patients with recurrent miscarriage happen in the first trimester and at least half of these are due to fetal chromosome abnormality (Simpson, 2007), which obviously has nothing to do with MBL deficiency. Studies of the role of MBL deficiency in patients with recurrent early miscarriages are expected to show only weak associations due to this ‘contamination’ of the patient group with cases mainly caused by fetal aneuploidies. Such patients can rarely be excluded from studies since the chromosomal constitutions of embryos lost in early pregnancy remain unknown in most cases. To be sufficiently statistically powered, the relevant studies must therefore be very large to be capable of identifying significant associations (Kruse et al., 2002).
In theory, the association between MBL deficiency and recurrent late pregnancy losses (RLPL) may be stronger than the association with early recurrent miscarriage due to a series of features characterizing the former group, which will be discussed below.
Very few women experience pregnancy losses after gestational week (GW) 14—in one study, only 0.2% of all women in the population had experienced one and 0.02% had experienced two or more unexplained second trimester losses (Parle-McDermott et al., 2005). In contrast, patients with recurrent miscarriage have been reported to exhibit a much higher risk of late pregnancy losses (Drakeley et al., 1998). Fetuses miscarried after GW 14 are in most cases subjected to karyotyping and autopsy; however, aneuploidy or fetal malformations seem to be rare causes in this kind of pregnancy loss (Ancel et al., 2000; Simpson, 2007). Maternal disorders are therefore generally believed to be the dominant cause of sporadic late pregnancy loss and RLPL (Ball et al., 2006).
For many years, cervical insufficiency (CI) has been a recognized maternal cause of second trimester losses. The CI-like clinical picture is characterized by painless cervical dilatation in the second trimester (ACOG Practice Bulletin, 2003), generally in the absence of bleeding or clinical infection. At admission to hospital, the uterine cervix is typically found very shortened and dilated, often with a prolaps of the amniotic membranes followed by delivery of a fetus often exhibiting vital signs but dying within a few minutes due to extreme prematurity. Sometimes an emergency cerclage had been placed after cervical dilatation was revealed or a cerclage had been placed in the first trimester due to the reproductive history. It has been estimated that 5–8% of all second trimester miscarriages are due to CI (Drakeley et al., 1998; Ancel et al., 2000). Miscarriages under a CI-like picture are probably often caused by cervical weakness due to previous cervical tears or lack of cervical hydroxyproline (Petersen and Uldbjerg, 1996). They may also be caused by intrauterine infection, which is often not clinically apparent but can be demonstrated by the detection of signs of intra-amniotic inflammation (Lee et al., 2008) or histological chorioamnionitis and virus DNA in the placenta (Srinivas et al., 2006). Since MBL is a part of the first-line immune defence against infections, MBL deficiency may in theory play a role in CI-like late pregnancy losses.
Approximately one-third of second trimester pregnancy losses are associated with the presence of the antiphospholipid antibodies lupus anticoagulant (LAC) and anticardiolipin antibodies (ACA) (Drakeley et al., 1998), which are especially associated with second trimester losses (Bocciolone et al., 1994; Rey et al., 2003; Robertson et al., 2005). Antiphospholipid antibodies may induce thrombotic events in the placental vessels since losses in late pregnancy associated with these antibodies are normally associated with extensive placental infarctions (Out et al., 1991). MBL deficiency has been proposed to be associated with the production of antiphospholipid antibodies (Seelen et al., 2005; Font et al., 2007) and to predispose to thrombotic events (Limnell et al., 2002) and through these mechanisms may predispose to late pregnancy loss.
In the present study, we investigated polymorphisms in the MBL2 gene (previously shown to be associated with plasma MBL levels) in all patients with RLPL defined as two or more losses of apparently normal fetuses between GW 14 and GW 30 referred to our department in a 22 year period. The aim was to investigate whether MBL2 gene polymorphisms associated with low plasma MBL levels are more prevalent in patients with RLPL than in controls and furthermore to investigate which subsets of patients with RLFL may display the strongest association with MBL deficiency.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Patients and controls
Patients with recurrent pregnancy loss, defined as three or more confirmed losses of intrauterine pregnancies in the first trimester, or two or more losses in the second or third trimester, have been referred to the Danish recurrent miscarriage clinic since 1986. At the first consultation, a detailed reproductive history was recorded based on information from the patients themselves and copies of relevant patient files from hospitals and clinics. The uterine cavity was investigated by hysterosalpingography, hysteroscopy or hydrosonography; thyroid-stimulating hormone levels were measured and all couples were karyotyped. These investigations were normal in all the included patients.
All patients were screened for LAC until 1999 using a commercially available test (Viper Quick LA-test and Viper Quick LA-check; Organon Technics, Durham, NC, USA) and after 2000 by an in-house LAC test based on (i) lack of correction of a prolonged activated partial thromboplastin time after mixture of the sample with an equal amount of normal plasma and (ii) correction of prolonged Russels viper venom-induced coagulation time (IL test LAC screen) after addition of phospholipid (IL test LAC confirm). Furthermore, the majority of patients with first trimester miscarriages and almost all patients with second and third trimester losses were screened for ACA of immunoglobulin (Ig)M and IgG type by enzyme-linked immunosorbent assay (ELISA) tests. Patients with a positive ACA measurement were investigated again at least 6 weeks later and only if the levels were above the cut-off in both measurements, the patients were classified as ACA positive (Wilson et al., 1999). The ELISA tests for ACA measurement and the cut-off levels for positivity have changed during the 22 year period where patients have been referred to our clinics. In most of the patients, ACA has been measured using an ELISA assay (Varelisa; Elias, Freiburg, Germany) with cut-off levels 7.0 MPL-U/ml and 22.0 GPL-U/ml for IgM and IgG ACA, respectively, established from a local group of females of fertile age (Nielsen and Christiansen, 2005). Most of the remaining patients have been investigated using an ELISA method (Varilisa cardiolipin, Pharmacia) using reference serum from University of Louisville, USA, with cut-off levels 28 MPL-kU/l and 16 GPL-kU/l. EDTA blood was drawn from all patients for DNA preparation at the first consultation.
One thousand one hundred and forty patients had recurrent miscarriage with no or only one loss after GW 14. Seventy-nine patients had two or more losses detected after GW 14, where fetal biparietal diameter measured in utero by ultrasound, or fetal weight and length measured at autopsy, unambiguously showed that the fetus had died after GW 14. Two patients with only two late pregnancy losses were excluded from the study because in one case the fetus had a severe malformation and in the other case it was aneuploid. In two patients, we had no DNA left in the freezer and 75 patients consequently comprised the study group. All except three of these patients were Caucasians. In the study group, all fetuses lost after GW 14 had been macroscopic normal. In almost all cases, autopsy had been carried out and no organ malformation was revealed. In the 51 cases where karyotyping had succeeded (Tables I–III), the fetuses proved euploid.
|
|
The RLPL patients were divided into three groups. Group 1 (Table I) consisted of patients with RLPL under a CI-like clinical picture characterized by painless cervical dilatation followed by spontaneous mid-trimester loss, in the absence of significant bleeding or clinical signs of infection.
Group 2 comprised patients with a positive test for LAC irrespective of the clinical appearance of the late pregnancy losses (Table II).
|
Group 3 (Table III) comprised patients with a negative test for LAC and RLPLs without any clinical suspicion of CI. Typically, the pregnancy losses occurred under clinical pictures of intrauterine fetal death (IUFD) detected by ultrasonographic examination (often preceded by fetal growth retardation and oligohydramnios at previous examinations) or placental abruption (PA) with heavy vaginal bleeding, uterine pains and ultrasonographic signs of intrauterine haematomas. Group 3 will, in the following text, be referred to as having ‘idiopathic RLPL’ in accordance with Drakeley et al. (1998)
since the generally recognized causes, LAC and CI, were absent.
Controls were 104 women who had given birth to live born children at least twice and who had not experienced miscarriages or stillbirths. Control women were either staff members of the Department of Clinical Immunology and the Department of Obstetrics and Gynaecology, Aalborg Hospital, or consecutive women who delivered at the Department of Obstetrics at Aalborg Hospital in 1999. The median age of the controls was 32.3 years (range 26–47 years) and the median parity 2 (range 2–4) and all were Caucasians. Detailed description of the control group can be found in the previous publication (Kruse et al., 2002).
MBL2 genotyping
DNA was extracted by a salting-out method from EDTA blood sample. Six SNPs in the MBL2 gene were genotyped: two point mutations in the promoter region at position –550 (H/L variants) and –221 (X/Y variants); one point mutation in the 5' untranslated region at position +4 (P/Q) and three point mutations that are located at codons 52, 54 and 57 in exon 1 of the MBL2 gene, at nucleotides 223, 230 and 239, giving rise to three allelic variants called D, B and C, respectively. The presence of any of the alleles D, B and C will be referred to as 0 whereas the wild-type (normal) allele will be called A.
A real-time PCR technique was used to genotype the exon 1 of the MBL2 gene (codons 52, 54 and 57), where only one PCR reaction is sufficient to genotype one individual unambiguously (Steffensen et al., 2003). A real-time TaqMan assay combined with minor groove binder probes labelled with 6-carboxy-fluorescein was used for screening the promoter variants –550, –220 and +4 (Mølle et al., 2006).
Division into MBL2 genotype groups
On the basis of the genotypes, patients and controls were categorized into three groups: the high genotype group associated with high (500–5000 ng/ml) plasma MBL levels: HYA/HYA, LYA/HYA, LYA/LYA and HYA/LXA and LYA/LXA; the intermediate genotype group associated with intermediate (50–1200 ng/ml) plasma MBL levels: HYA/0, LYA/0 and LXA/LXA and the low MBL genotype group associated with low (0–200 ng/ml) plasma MBL levels: 0/0 and 0/LXA (Madsen et al., 1995).
Statistics
Differences in the prevalence of MBL2 genotypes between the groups were tested by the 
2 test. Differences in median values were tested by the Mann–Whitney test. A P-value of <0.05 was considered significant (two-sided test). Odds ratios (OR) and their 95% confidence intervals (CI) were calculated.
Ethics
The investigation of patients admitted before 2000 and all controls had been undertaken as a part of a previous study approved by the Ethics Committee of the County of Nordjylland. All patients and controls gave informed written consent. After 2000, MBL investigation was undertaken as a part of the routine investigation of patients admitted to our clinic and the patients had accordingly given their consent.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Clinical data and antiphospholipid antibodies
Twenty-eight of the RLPL patients belonged to Group 1 since they had CI-like losses, in 19 of their pregnancies an elective or emergency cerclage had been attempted without success (Table I). Six of the patients (21.4%) had had a previous live birth and 17 of them (60.7%) had had previous early miscarriages. One patient in Group 1 had insulin-dependent diabetes mellitus and one had a sarcoidosis-like lung disease, but none had experienced thromboembolic phenomena. Two (9.1%) of the patients in whom ACA measurements were undertaken were repeatedly ACA positive but with low or intermediate titres.
The nine RLPL patients who were LAC positive and thus belonged to Group 2 had only late losses of the IUFD type but six of them had also early miscarriages. None of these patients had experienced live births at the time of admission. In addition to LAC, all were also repeatedly positive for ACA with high titres (Table II). Four of the patients had had previous venous thromboembolic episodes and three had a diagnosis of systemic lupus erythematosus.
Thirty-eight patients had had idiopathic RLPL and comprised Group 3 (Table III). Twenty-five of these had experienced only late pregnancy losses of the IUFD type, eight had only losses of the PA type, four had experienced both IUFDs and PA-like losses and one had had a PA-like and a CI-like loss. Twenty-one of these patients (55.3%) had had one or more previous live births and 27 (71.1%) had experienced early miscarriages. None of the patients in Group 3 had had thromboembolic phenomena, but one had the autoimmune disease scleroderma. Six (20.0%) of the patients in whom ACA measurements were available were repeatedly ACA positive but with low or intermediate titres.
Table IV shows the median age and number of pregnancy losses, the frequency of previous live births and the frequency of positivity for ACA in patients with high/intermediate- and low-producing MBL2 genotypes, respectively. No differences in these parameters were found between the three study groups.
|
MBL2 genotypes
Overall, MBL2 genotypes associated with low MBL were found in 20 (26.7%) of the 75 patients with RLPL (Table IV); which is significantly higher than the 12.5% frequency in controls (OR = 2.55; 95% CI 1.17–5.52; P < 0.02).
The frequencies of MBL2 genotypes associated with high, intermediate and low plasma MBL levels, respectively, were compared between Group 1 patients and controls (Table V). Low-producing MBL2 genotypes were found in five patients (17.9%), a frequency that was increased, however not significantly, compared with controls (OR 1.52, 95% CI 0.49–4.70).
|
The frequencies of MBL2 genotypes were compared between patients in Groups 2 and 3 and controls, respectively (Table VI). The patients in Group 2 had a similar frequency of low-producing MBL2 genotypes as controls (11.1% versus 12.5%) with a corresponding OR of 0.88 (95% CI 0.10–7.58). In the 38 patients in Group 3, 36.8% carried low-producing MBL2 genotypes resulting in an OR 4.08 (1.70–9.83; P = 0.001) compared with controls.
|
From the OR = 4.08 and the 36.8% prevalence of low-producing MBL2 genotypes in Group 3 patients, it can be calculated that the population attributable risk of low-producing MBL2 genotypes for idiopathic RLPL is 0.28.
Pregnancy outcome after referral
Among the patients in Group 1, 19 became pregnant again after referral—eight (42%) of these pregnancies succeeded with a live birth. Two of the patients who gave birth were treated with an elective cerclage, three were treated with indometacin in the second trimester, two were treated with both elective cerclage and indometacin in the second trimester and one received i.v. Ig (IvIg) in the first trimester and indometacin in the second. Among the patients in Group 2, eight became pregnant again after referral. Four (50%) of these had a live birth after a combination of heparin and low-dose aspirin (LDA) and IvIg, one gave birth after IvIg alone and one gave birth after heparin and LDA alone.
Among the patients in Group 3, 30 became pregnant again and 22 (73%) had successful pregnancies. Several of these patients were treated in previous placebo-controlled trials of IvIg. Thirteen had a live birth after IvIg alone, three after a combination of IvIg and heparin and LDA, one after heparin and LDA and five after no treatment at all.
Among the patients in Group 3 who became pregnant, 15 out of 20 (75.0%) in the high/intermediate MBL group had a live birth compared with 7 out of 10 (70.0%) in the low MBL group. Among patients who became pregnant in all three groups, 26 out of 43 gave birth (60.4%) in the high/intermediate MBL2 genotype group compared with 10 out of 14 (71.4%) in the low MBL2 genotype group (not significant).
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Recurrent second trimester pregnancy losses (RLPL) is psychologically very devastating but fortunately a rare condition. Many patients with RLPL display clinical signs of CI (Group 1) or have LAC in the plasma (Group 2) and these women can be given an explanation and often offered treatments. However, in the majority of RLPL patients (idiopathic RLPL = Group 3), none of these risk factors can be detected and so far it has not been possible to give the patients any explanation for their losses.
This study supported our hypothesis that MBL2 genotypes associated with MBL deficiency are more strongly associated with RLPL (OR = 2.55) than with recurrent early miscarriages where we have previously found an OR = 1.33 using the same control group (Kruse et al., 2002).
Low-producing MBL2 genotypes were most strongly associated with a history of idiopathic RLPL (OR = 4.08), i.e. RLPL without clinical signs of CI in patients negative for LAC.
The population attributable risk of low-producing MBL2 genotypes in idiopathic RLPL reveals that 27.8% of all idiopathic RLPL cases would disappear, if these genotypes disappeared completely in the population.
We found a trend for low-producing MBL2 genotypes to be associated with RLPL under a clinical picture resembling CI (Table V). This could indicate that low MBL may be a contributing but not a major factor for pregnancy loss in this subgroup of RLPL patients.
The frequency of low-producing MBL2 genotypes was similar in LAC positive patients compared with controls (Table VI), which supports the hypothesis that LAC may be the only pathogenic factor in Group 2 patients. LAC is associated with an increased risk of thrombosis and is suggested to cause thrombosis in the placental vessels (Out et al., 1991) and subsequent placental infarctions and IUFD. This fits with the observation that RLPL patients with LAC exhibit a unique clinical picture: late IUFDs with no bleeding and often a history of maternal thromboembolic episodes (Table II). The correlation between ACA in the absence of LAC and the clinical appearance of late pregnancy losses is weak. ACA can be found in patients without signs of placental infarctions or fetal growth retardation (Table I), ACA without LAC is not or only weakly associated with thrombosis (Galli et al., 2003) and ACA is a very fluctuating antibody and tests needs standardization (Branch, 2004). Since we find LAC, but not ACA alone, a well-documented cause of pregnancy loss, we had classified LAC-positive but not ACA-positive patients into a separate group. In our study, low MBL seems not to predispose to the production of antiphospholipid antibodies, as suggested in some studies (Seelen et al., 2005; Font et al., 2007), since there was no difference in ACA positivity between those with and without genotypes associated with MBL deficiency (Table IV). Since the laboratory methods were modified and improved several times, the definition of positivity for ACA changed during the 22 year period where the patients had been referred to our clinic. The current criteria for the diagnosis of the antiphospholipid syndrome are two positive LAC tests and/or ACA tests on two or more occasions at least 12 weeks apart. For ACA, the recommended cut-off levels are >40 GPL or MPL units or >99th percentile measured by a standard ELISA assay (Miyakis et al., 2006).
The frequency of the low-production MBL2 genotypes 0/0 and 0/LXA among the controls was 12.5%, which is lower than the 17.6% frequency of the same genotypes found in 533 randomly selected Danes of both sexes (Hellemann et al., 2007). This discrepancy may be due to the exclusion of women with a history of miscarriage in our control group, which comprised exclusively women with normal pregnancies. Dahl et al. (2004) reported that MBL deficiency is associated with a borderline significantly increased risk of miscarriage in the background population, so a control group of women without miscarriages is expected to exhibit a decreased frequency of MBL2 low-production genes compared with unselected women. As mentioned in the previous section, the ranges of plasma MBL levels overlap between genotypes in the high, intermediate and low groups, e.g. an individual with an intermediate genotype may in fact have an MBL level below 100 ng/ml and would thus belong to the low MBL group in our previous study (Kruse et al., 2002). In the present study, we chose to investigate MBL genotypes instead of measuring the plasma protein because (i) we had DNA but no serum left from several of our patients and (ii) many of the previous plasma MBL measurements in the control women had been undertaken just after delivery (Kruse et al., 2002) where MBL levels are reported to be significantly increased from baseline (van de Geijn et al., 2007b). Using only the results based on genotyping would overcome the methodological problem that some of the available or previously tested plasma samples were taken before pregnancy and some in the puerperal period. However, the lack of data for plasma MBL levels in these patients may be a limitation of our study.
Our findings that MBL deficiency in mid-pregnancy is associated with pregnancy losses probably caused by malfunction of the placenta (placental insufficiency or abruption) rather than mechanisms associated with spontaneous preterm birth (cervical effacement and rupture of membranes) are consistent with findings in most studies of the effect of MBL deficiency in the third trimester.
The data concerning the association between MBL deficiency and preterm birth have been conflicting. One study reported that carriage of a low-production MBL2 polymorphism (codon 52 in exon 1) was significantly more frequent in infants born preterm than in infants born at term (Bodamer et al., 2006). One study (Annells et al., 2004) found that the presence of the codon 54 low-production MBL2 polymorphism was significantly associated with preterm birth, before 29 GW. On the other hand, maternal high-production MBL2 genotypes were found to be associated with a shorter gestational period than low-production MBL2 genotypes in primiparae (van de Geijn et al., 2008) and low MBL (<100 ng/ml) was not associated with an increased prevalence of preterm birth in patients with recurrent miscarriage (Kruse et al., 2002).
There is more consensus about the role of MBL deficiency for intrauterine growth retardation. The mean birthweight of children born to healthy women with the low-production MBL2 codon 54 polymorphism in exon 1 was decreased compared with women without the mutation (Megia et al., 2004) and MBL deficiency in patients with recurrent miscarriage was associated with a significant reduction of birthweight among children born >37 GW. These data support the theory that low MBL predisposes to impaired placenta growth and function but does not advance the time of delivery, which agrees with the findings in the present study.
The mechanisms through which MBL deficiency can contribute to placental insufficiency or abruption are unknown, but hypotheses can be put forward. Normal pregnancy has been proposed as being dependent on an adequate level of inflammation (Christiansen et al., 2006): excessive inflammation seems to be associated with pre-eclampsia (Borzychowski et al., 2005). The correct level of inflammation may be induced by apoptotic syncytiotrophoblast debris shed into the circulation from the placenta: if the shedding of debris becomes too excessive or if the clearance becomes deficient, too much inflammation may result. One function of MBL may be clearance of apoptotic cell material in the circulation (Ogden et al., 2001; Nauta et al., 2004; Stuart et al., 2005) and it is thus possible that pregnant women with low MBL may be at higher risk of excessive inflammatory responses due to failure of this clearance. MBL also seems to be a regulator of production of inflammatory cytokines such as tumour necrosis factor (TNF)-
, at least in vitro (Jack et al., 2001). Women with low MBL may be in a higher risk of excessive TNF- production, which may induce apoptosis of trophoblast (Kilani et al., 2007) and worsen the vicious circle of excessive trophoblast cell apoptosis and impaired clearance of apoptotic cells, leading to further inflammation. Another possible mechanism of late pregnancy loss in women with MBL deficiency may be impairment of clearance of micro-organisms at the fetomaternal interphase or in the amniotic fluid, which may predispose to cervical ripening or amniotic membrane rupture and thus extremely preterm birth (Lee et al., 2008).
Since 12.5% of multiparous women without miscarriages carry low MBL2 genotypes, low MBL on its own cannot cause RLPL. We have proposed (Christiansen et al., 2008) that recurrent pregnancy loss in most cases should be considered a multifactorial/polygenic disease at the level of the individual woman: each patient or patient couple must carry a series of genetic risk factors (which may be genetic polymorphisms related to immune response, thrombophilia and endocrine function) and be exposed to specific environmental factors before the syndrome develops. The present study shows that in patients with idiopathic RLPL, one of these genetic risk factors, responsible for 27.8% of the genetic background, is the genes associated with MBL deficiency and this suggests that the immunologic component is strong in this subset of RLPL patients. It also increases the chance that therapies which have proven efficient in the treatment of diseases characterized by excessive inflammation, such as IvIg, may be efficient in treating idiopathic RLPL. Indeed, among 17 women with idiopathic RLPL comprising a subset of the patients who were included in our two previous placebo-controlled trials of IvIg (Christiansen et al., 2002), seven of eight patients (87%) who were allocated to IvIg gave birth compared with only one of nine (11%) of those who received placebo (P < 0.01) (Christiansen et al., 2004). The active therapies with especially IvIg and heparin offered to most of the patients may be the reason that the live birth rates in subsequent pregnancies were similar in patients belonging to the high/intermediate MBL and low MBL groups. If the majority of the patients remained untreated, a decreased live birth rate in the low MBL group may have been found as reported in our previous study (Kruse et al., 2002).
In conclusion, the finding of a highly increased frequency of low-producing MBL2 genotypes among patients with idiopathic RLPL supports the hypothesis that the aetiology of this kind of RLPL is, to a high degree, immunological. In the future, research should be directed into the identification of other genetic biomarkers that, together with low MBL-associated genotypes, may constitute the complete genetic background for idiopathic RLPL, which may then no longer be called ‘idiopathic’.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
ACOG Practice Bulletin. Cervical insufficiency. Obstet Gynecol (2003) 102:1091–1099.[CrossRef][Medline]
Ancel P-Y, Saurel-Cubizolles M-J, Di Renzo GC, Papiernik E, Breart G, The Europop Group. Risk factors for 14–21 week abortions: a case–control study in Europe. Hum Reprod (2000) 15:2426–2432.
Annells MF, Hart PH, Mullighan CG, Heatley SL, Robinson JS, Bardy P, McDonald HM. Interleukins-1,-4,-6,-10, tumor necrosis factor, transforming growth factor-β, FAS, and mannose-binding protein C gene polymorphisms in Australian women: risk of preterm birth. Am J Obstet Gynecol (2004) 191:2056–2067.[CrossRef][Web of Science][Medline]
Ball E, Bulmer JN, Ayis S, Lyall F, Robson SC. Late sporadic miscarriage is associated with abnormalities in spiral artery transformation and trophoblast invasion. J Path (2006) 208:535–542.[CrossRef][Medline]
Baxter N, Sumiya M, Cheng S, Erlich H, Regan L, Simons A, Summerfield JA. Recurrent miscarriage and variant alleles of mannose binding lectin, tumour necrosis factor and lymphotoxin genes. Clin Exp Immunol (2001) 126:529.[CrossRef][Web of Science][Medline]
Bocciolone L, Meroni P, Parazzini F, Tincani A, Radici E, Tarantini M, Rossi E, Bianchi C, Mezzanotte C, D'Angelo A. Antiphospholipid antibodies and the risk of intrauterine late fetal death. Acta Obstet Gynecol Scand (1994) 73:389–392.[Web of Science][Medline]
Bodamer OA, Mitterer G, Pollak A, Mueller MW, Schmidt WM. Evidence for an association between mannose-binding lectin 2 (MBL2) gene polymorphisms and pre-term birth. Genet Med (2006) 8:518–524.[Web of Science][Medline]
Borzychowski AM, Croy BA, Chan WL, Redman CWG, Sargent IL. Changes in systemic type 1 and type 2 immunity in normal pregnancy and pre-eclampsia may be mediated by natural killer cells. Eur J Immunol (2005) 35:3054–3063.[CrossRef][Web of Science][Medline]
Branch DW. Antiphospholipid antibodies and fetal compromise. Thromb Res (2004) 114:415–418.[CrossRef][Web of Science][Medline]
Christiansen OB, Kilpatrick DC, Souter V, Varming K, Thiel S, Jensenius JC. Mannan-binding lectin deficiency is associated with unexplained recurrent miscarriage. Scand J Immunol (1999) 49:193–196.[CrossRef][Web of Science][Medline]
Christiansen OB, Pedersen B, Rosgaard A, Husth M. A randomized, double-blind, placebo-control-led trial of intravenous immunoglobulin in the prevention of recurrent miscarriage: evidence for a therapeutic effect in women with secondary recurrent miscarriage. Hum Reprod (2002) 17:809–816.
Christiansen OB, Nielsen HS, Pedersen B. Active or passive immunization in unexplained recurrent miscarriage. J Reprod Immunol (2004) 62:41–52.[CrossRef][Web of Science][Medline]
Christiansen OB, Nielsen HS, Kolte AM. Inflammation and miscarriage. Semin Fetal Neonatal Med (2006) 11:302–308.[CrossRef][Web of Science][Medline]
Christiansen OB, Steffensen R, Nielsen HS, Varming K. Multifactorial etiology of recurrent miscarriage and its scientific and clinical implications. Gynecol Obstet Invest (2008) 66:257–267.[CrossRef][Medline]
Dahl M, Tybjaerg-Hansen A, Schnohr P, Nordestgaard BG. A population-based study of morbidity and mortality in mannose-binding lectin deficiency. J Exp Med (2004) 199:1391–1399.
Drakeley AJ, Quenby S, Farquharson RG. Mid-trimester loss—appraisal of a screening protocol. Hum Reprod (1998) 13:1975–1980.
Font J, Ramos-Casals M, Brito-Zeron P, Nardi N, Ibanez A, Suarez B, Jiménez S, Tàssies D, Garcia-Criado A, Ros E, et al. Association of mannose-binding lectin polymorphisms with antiphospholipid syndrome, cardiovascular disease and chronic damage in patients with systemic lupus erythematosus. Rheumatology (2007) 46:76–80.
Galli M, Luciani D, Bertolini G, Barbui T. Anti-β2-glycoprotein I, antiprothrombin antibodies, and the risk of thrombosis in the antiphospholipid syndrome. Blood (2003) 102:211–225.
Hellemann D, Larsson A, Madsen HO, Bonde J, Jarløv JO, Wiis J, Faber T, Wetterslev J, Garred P. Heterozygosity of mannose-binding lectin (MBL2) genotypes predicts advantage (heterosis) in relation to fatal outcome in intensive case patients. Hum Mol Genet (2007) 16:3071–3080.
Jack DL, Read RC, Tenner AJ, Frosch M, Turner MW, Klein NJ. Mannose-binding lectin regulates the inflammatory response of human professional phagocytes to Neisseria menigititis Serogroup B. J Infect Dis (2001) 184:1152–1162.[CrossRef][Web of Science][Medline]
Kilani RT, Mackova M, Davidge ST, Winkler-Lowen B, Demianczuk N, Guilbert LJ. Endogenous tumor necrosis factor mediates enhanced apoptosis of cultured villous trophoblasts from intrauterine growth-restricted placentae. Reproduction (2007) 133:257–264.
Kilpatrick DC, Bevan BH, Liston WA. Association between mannan-binding protein deficiency and recurrent miscarriage. Hum Reprod (1995) 10:2501–2505.
Kruse C, Rosgaard A, Steffensen R, Varming K, Jensenius JC, Christiansen OB. Low serum level of mannan-binding lectin is a determinant for pregnancy outcome in women with recurrent spontaneous abortion. Am J Obstet Gynecol (2002) 187:1313–1320.[CrossRef][Web of Science][Medline]
Lee SE, Romeo R, Park C-W, Jun JK, Yoon BH. The frequency and significance of intraamniotic inflammation in patients with cervical insufficiency. Am J Obstet Gynecol (2008) 198:633.e1–633.e8.
Limnell V, Aittoniemi J, Vaarala O, Lehtimäki T, Laine S, Virtanen V, Palosuo T, Miettinen A. Association of mannan-binding lectin deficiency with venous bypass graft occlusions in patients with coronary heart disease. Cardiology (2002) 98:123–126.[CrossRef][Web of Science][Medline]
Madsen HO, Garred P, Thiel S, Kurtzhals JAL, Lamm LU, Ryder LP, Svejgaard A. Interplay between promoter and structural gene variants control basal serum level of mannan-binding protein. J Immunol (1995) 155:3013–3020.[Abstract]
Megia A, Gallart L, Fernandez-Real J-M, Vendrell J, Simon I, Gutierrrez C, Richart C. Mannose-binding lectin gene polymorphisms are associated with gestational diabetes mellitus. J Clin Endocrinol Metab (2004) 89:5081–5087.
Miyakis S, Lockshin M, Atsumi T, Branch D, Brey R, Cervera R, Derksen R, De Groot P, Koike T, Meroni P, et al. International consensus statement on an update of the classification criteria for the definite antiphospholipid syndrome (APS). J Thromb Haemost (2006) 4:295–306.[CrossRef][Web of Science][Medline]
Mølle I, Steffensen R, Thiel S, Peterslund NA. Chemotherapy-related infections in patients with multiple myeloma: associations with mannan-binding lectin genotypes. Eur J Haematol (2006) 77:19–26.[CrossRef][Web of Science][Medline]
Nauta AJ, Castellano G, Xu W, Woltman AM, Borrias MC, Daha MR, van Kooten C, Roos A. Opsonization with C1q and mannose-binding lectin targets apoptotic cells to dendritic cells. J Immunol (2004) 173:3044–3050.
Nielsen HS, Christiansen OB. Prognostic impact of anticardiolipin antibodies in women with recurrent miscarriage negative for the lupus anticoagulant. Hum Reprod (2005) 20:1720–1728.
Ogden CA, deCathelineu A, Hoffmann PR, Bratton D, Ghebrehiwet B, Fadok VA, Henson PM. C1q and mannose binding lectin engagement of cell surface calreticulin and CD91 initiates macropinocytosis and uptake of apoptotic cells. J Exp Med (2001) 194:781–795.
Out HJ, Kooijman CD, Bruinse HW, Derksen RH. Histopathological findings in placentae from patients with intra-uterine fetal death and anti-phospholipid antibodies. Eur J Obstet Gynecol Reprod Biol (1991) 41:179–186.[CrossRef][Web of Science][Medline]
Parle-McDermott A, Pangilinan F, Mills JL, Signore CC, Molloy AM, Cotter A, Conley M, Cox C, Kirke PN, Scott JM, et al. A polymorphism in the MTHFD1 gene increases a mother's risk of having an unexplained second trimester pregnancy loss. Mol Hum Reprod (2005) 11:477–480.
Petersen LK, Uldbjerg N. Cervical collagen in non-pregnant women with previous cervical incompetence. Eur J Obstet Gynecol Reprod Biol (1996) 67:41–45.[CrossRef][Web of Science][Medline]
Rey E, Kahn SR, David M, Shrier I. Thrombophilic disorders and fetal loss: a meta-analysis. Lancet (2003) 361:901–908.[CrossRef][Web of Science][Medline]
Robertson L, Wu O, Langhorne P, Twaddle S, Clark P, Lowe GDO, Walker ID, Greaves M, Brenkel I, Regan L, et al. Thrombophilia in pregnancy: a systematic review. Br J Haematol (2005) 132:171–196.[CrossRef][Web of Science]
Seelen MA, van der Bijl EA, Trouw LA, Zuiverloon TCM, Munoz JR, Fallaux van den Houten FC, Schlagwein N, Daha MR, Huizinga TW, Roos A. A role for mannose-binding lectin dysfunction in generation of autoantibodies in systemic lupus erythematosus. Rheumatology (2005) 44:111–119.
Simpson JL. Causes of fetal wastage. Clin Obstet Gynecol (2007) 50:10–30.[CrossRef][Web of Science][Medline]
Srinivas SK, Ma Y, Sammel MD, Chou D, McGrath C, Parry S, Elovitz MA. Placental inflammation and viral infection are implicated in second trimester pregnancy loss. Am J Obstet Gynecol (2006) 195:797–802.[CrossRef][Web of Science][Medline]
Steffensen R, Hoffmann K, Varming K. Rapid genotyping of MBL2 gene mutations using real-time PCR with fluorescent hybridisation probes. J Immunol Methods (2003) 278:191–199.[CrossRef][Web of Science][Medline]
Stuart LM, Takahashi K, Shi L, Savill J, Ezekowitz RAB. Mannose-binding lectin-deficient mice display defective apoptotic cell clearance but no autoimmune phenotype. J Immunol (2005) 174:3220–3226.
Van de Geijn FE, Dolhain RJEM, van Rijs W, Hazes LMW, de Groot CJM. Mannose-binding lectin genotypes and pre-eclampsia: a case–control study. Hum Immunol (2007) a 68:888–893.[CrossRef][Web of Science][Medline]
Van de Geijn FE, Roos A, de Man YA, Laman JD, de Groot CJM, Daha MR, Hazes JMW, Dolhain RJEM. Mannose-binding lectin levels during pregnancy: a longitudinal study. Hum Reprod (2007) b 22:362–371.
Van de Geijn FE, Dolhain RJEM, van Rijs W, Wilhelmsen SP, Hazes LMW, de Groot CJM. Mannose-binding lectin genotypes are associated with shorter gestational age. An evolutionary advantage of low MBL production genotypes? Mol Immunol (2008) 45:1514–1518.[CrossRef][Web of Science][Medline]
Wilson WA, Gharavi AE, Koike T, Lockshin MD, Branch DW, Piette JC, Brey R, Derksen R, Harris EN, Hughes GR, et al. International consensus statement on preliminary classification criteria for definite antiphospholipid syndrome: report of an international workshop. Arthritis Rheum (1999) 42:1309–1311.[CrossRef][Web of Science][Medline]
Submitted on July 30, 2008; resubmitted on September 17, 2008; accepted on September 22, 2008.
CiteULike 
Connotea 
Del.icio.us What's this?
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||