Recurrent Spontaneous Pregnancy Research Articles

Role of the CD40–CD40L expression level pathway in the diagnosis of unexplained recurrent pregnancy loss

Background Unexplained recurrent spontaneous pregnancy loss (URPL) lacks effective treatment and reliable early diagnosis and prediction. Immunologic dysfunction can be an underlying cause of recurrent pregnancy loss (RPL). Considering the regulatory role of CD40–CD40L in immune responses, we explored its clinical significance in URPL. Methods The 108 women with URPL who were treated in Hebei Yanda Hospital from January 2020 to December 2022 were selected as study subjects, and another 108 healthy women who were not pregnant and matched with the age and body mass index of the study group were selected as the control group. CD40 and CD4 + CD25 + Treg cells in peripheral blood mononuclear cells (PBMCs) and CD40L in peripheral blood platelets were measured by flow cytometry. The predictive value of CD40–CD40L in URPL for the risk of RPL was determined by receiver operating characteristic (ROC) curves. The correlations of CD40–CD40L with CD4 + CD25 + Treg cells and serum pro-inflammatory factors were assessed by Pearson’s analysis. Results CD40 on the surface of PBMCs and CD40L on the surface of platelets were up-regulated in URPL patients. CD40 in combination with CD40L had high predictive value for the risk of RPL in URPL patients. Peripheral blood CD40–CD40L was positively linked to IL-17 and IL-23, and negatively to CD4 + CD25 + Treg cells and IL-10 in URPL patients. Conclusions The CD40–CD40L pathway expression in peripheral blood can help predict the risk of RPL in URPL patients.

Comparison of Vitamin D Status between Healthy Pregnant Women and Women with Recurrent Early Spontaneous Pregnancy Loss

Introduction: Recurrent early spontaneous pregnancy loss is the most common negative outcome of pregnancy and identification of modifiable risk factors is potentially of great importance for public health. The immunological factors may play a role in failure of pregnancy in women with recurrent pregnancy loss. The role of this vitamin D has also been proved in the immune system. Aim of the Study: This study aim to determine serum vitamin D status among women with recurrent early spontaneous pregnancy loss and to compare the vitamin D level among them with healthy pregnant women. Methods: This cross sectional comparative study was done at the out patient department of obstetrics and Gynecology in Bangabandhu Sheikh Mujib Medical University during September 2018 to August 2019. The study comprises of 70 women carrying singleton fetus, gestational age between 8 to 12 weeks, in the age range of 18 to 35 years. The respondents were divided into two groups, 35 pregnant women in their early pregnancy loss with previous history of two or more early spontaneous pregnancy loss were considered as group I and 35 women in their early live pregnancy with previous history of one or more successful pregnancy and no history of any pregnancy loss were considered as group II. Fasting serum vitamin D level was measure by CMIA. Results: This study found that most the respondents were vitamin D deficient in both groups. In group I 17(48.6%) and in group II 10(28.6%) were vitamin D severe deficient. Again in group I 18(51.4%) and 24(68.6%) in group II were vitamin D deficient. The mean vitamin D level was 9.90±2.28 ng/ml in group I and 11.43±3.96 ng/ml in group II. The differences was statistically not significant (P>0.05) between two groups. There is a negative correlation (r-0.235; p 0.175) between serum vitamin D level and number of pregnancy loss. Which indicate increase number of pregnancy loss associated with decrease level of vitamin D. Conclusion: Vitamin D deficient in both groups, however the vitamin D level was comparatively low in women with recurrent early spontaneous pregnancy loss but the differences was not significant between two groups.

Prevalence and pregnancy outcome in antiphospholipid syndrome: an observational study

Antiphospholipid syndrome (APS) is an autoimmune disorder characterized by vascular thrombosis, both arterial and venous, and recurrent spontaneous pregnancy loss due to circulating antiphospholipid antibodies (aPL) including anticardiolipin antibodies (aCL) and/or anti-glycoprotein I antibodies (anti-GPI). This observational study was conducted among antenatal women at OBG Department, IMS, and SUM Hospital, Bhubaneswar, over a year from March 2020 to 2021. Of 1260 cases, 25 were diagnosed with ALPA syndrome, with a prevalence rate of 1.98%. The most common age was 26-30 years (40%) with a mean age of 28.83 ± 3.26 years. The primary cause was recurrent 1st-trimester spontaneous miscarriage. Beta 2 glycoprotein antibody was most prevalent (40%). Significant associations were found in hypertensive and FGR APLA patients compared to non-hypertensive and non-FGR APLA patients. Hypertension and FGR occurred in 24% and 20% of APLA patients, respectively. Preterm delivery rate was higher (52%) in APLA syndrome, without significant association. B2GP antibody was linked to the lowest live birth rate, and highest rates of preeclampsia, IUGR, and stillbirth. Screening for B2GP antibodies is recommended for FGR and early onset preeclampsia cases with/without previous spontaneous pregnancy loss.

The role of the HLA-complex genes in genesis of human idiopathic recurrent pregnancy loss

Aim.To study the immunogenetic prerequisites of idiopathic recurrent spontaneous pregnancy losses in humans of allo- and autoimmune genesis. Methods. PCR-SSP (polymerase chain reaction with sequence-specific primers). Results. A comprehensive analysis of the distribution and frequency of allelic variants of the HLA-DRB1, HLA-DQA1, HLA-DQB1 genes testifies to an increased risk of recurrent pregnancy losses with a total homology of 50 % or more in couples and presence of the DQ2.5 genotype in a woman. The OR indicates that the risk of idiopathic recurrent spontaneous pregnancy loss increases 2.68 times when the homology of the couples is 50 % or more for two loci (HLA-DRB1, HLA-DQA1) and 12.8 times when the homology is 50 % or more for by three loci (HLA-DRB1, HLA-DQA1, HLA-DQB1), and if the DQ2.5 genotype is present in women, this risk increases 4 times. Conclusions. HLA genotyping of married couples with a history of spontaneous pregnancy loss is important for preconceptional prevention of recurrent pregnancy losses.

A Case Report of Laparotomic Myomectomy After The Use of Gnrh Analogue in A Brazilian Woman

Uterine leiomyoma is the most common genital benign tumor in the female population. It is estimated that about 20 to 40% of women will have leiomyoma during their reproductive stage. IBGE (Brazilian Institute of Geography and Statistics) estimated the Brazilian population to be over 200 million Brazilians, of which half are women and approximately 50% are of reproductive age; hence, by calculating the prevalence of leiomyomatosis, there would be just over 6 million women with symptomatic fibroids in Brazil that require clinical or surgical treatment.[1-3-4] Due to its presence during the reproductive period, it can be caused by infertility or recurrent spontaneous pregnancy loss. Hence, myomectomy can be necessary when the fibroid itself causes distortion of the endometrial cavity, or competes with the embryo for the same place, or when conservative treatment was ineffective. This report describes a case of myomectomy which was performed after treatment with goserelin acetate with considerable reduction of fibroids volume, probably facilitating surgical approach, decreasing the blood loss and the risk of hysterectomy conversion. In addition, other GnRH analogue benefits before surgery such as doctor’s impression of decrease in size and better mobility of the tumor and patients feeling of decrease of the abdominal volume and the bleeding. This subjective approach can optimize the timing of surgery and establish the number of injections.

Immunologic deviations in recurrent spontaneous abortion mouse model.

Recurrent spontaneous abortion (RSA) or recurrent pregnancy loss (RPL) is an abnormality that has a great impact on women's quality of life. RSA is defined as at least three unexplained abortions (without prior live birth) occurring before the 20th week of pregnancy. The present review attempts to discuss immunologic deviations in mouse models of RSA. The mating of DBA/2J males with CBA/J female mice has provided specialists with a homologous model of RSA. Much of the research using the CBA/J × DBA/2J mouse model has shown immune system alteration results in rejection. The link between RSA and the immune system suggests new approaches to prevent RSA from an immunological perspective. Rejection in this model is linked with the changed immune system during pregnancy, including change in Th1/Th2 ratio and defects in T and NK cells function, and so forth. The use of animal models prone to RSA can help a lot to solve the remained mysteries. This study reviews the existing knowledge of immune system roles in the RSA mouse models.

Study on Anticardiolipin Antibodies in Women With Recurrent Abortion in Duhok Province, Kurdistan Region, Iraq

Recurrent abortion is a worldwide issue. Anticardiolipin antibodies (ACA) are found to be among the most important factors related to recurrent spontaneous early pregnancy loss. This study aimed to investigate the prevalence of anticardiolipin IgM and IgG antibodies in women with recurrent abortion in Kurdistan Region of Iraq. The present study was conducted in Duhok and Zakho cities for the period from April 2014 to May 2019. A total of 1230 women aged between 18 to 46-year-old were included in this study. ELISA was used for the detection of anticardiolipin antibodies. Among the studied subjects, the prevalence of anticardiolipin antibodies was 74 (6.02%) for ACA IgM and 56 (4.6%) for ACA IgG. It was also observed that 16 (1.3%) subjects were positive for both ACA IgM and IgG antibodies. Additionally, the highest anticardiolipin antibody positivity rates were recorded in patients older than 30-year-old (P<0.01). The IgM ACA positivity was higher in Zakho city 45 (7.2%) when compared to Duhok city 29 (4.6%) (P<0.04). In conclusion, anticardiolipin antibodies can have a positive association among women with recurrent abortion. Therefore, it is suggested that women with recurrent abortion should be screened for anticardiolipin antibodies; this could increase fetal survival by initiating early anticoagulant therapy when other causes of abortion are excluded.

Efficiency of organ-saving operations conducted about early postpartum uterine bleeding and their influence on the quality of women’s lives

Objective: to study the long-term eff ects of massive blood loss in parturition and the early postpartum period on the reproductive function and psychological status of women.Materials and methods: the study included 49 women who were imposed compressive hemostatic sutures in the uterus due to the development of early postpartum hemorrhage. It was carried out a retrospectiveanalysis of medical records, prospective survey - 46 women, tested scales postpartum depression - 46 women and assessment of quality of life using the SF-36 scales - 36 women.Results: the effi ciency of the compression joints method was 98 % (100 % exclusion criteria were coagulopathic obstetric bleeding), in 1 case the application of compression joints was ineff ective, which required uterus extirpation. In 14 women out of 48 (29.1 %), menstrual irregularities were revealed. In 28 (61.5 %) women, postnatal depression was confi rmed. In 30 out of 36 (87 %) women were found to decrease in quality of life. In 8 (16.7 %) women, a recurrent spontaneous pregnancy occurred within 2 years aft er suturing.Conclusion. Compression sutures remain an effective method for stopping postpartum uterine bleeding, allowing women to maintain fertility. However, some factors, including high blood loss, can signifi cantly reduce the quality of women’s lives.

The Association of Serum Vitamin D and Calcium Levels with Early Recurrent Spontaneous Pregnancy Loss

The Association of Serum Vitamin D and Calcium Levels with Early Recurrent Spontaneous Pregnancy Loss

Heterozygosity of maternal factor V G1691A (Leiden) and relationship with times of pregnancy loss among unexplained recurrent pregnancy loss women

Background: Recurrent pregnancy loss is defined by the consecutive loss of two or more pregnancies with the same partner. Recurrent pregnancy loss (RPL) or recurrent miscarriage (RM) affects from 1-5% of the reproductive age couples. This diagnosis is both emotionally challenging and confusing for most couples, as the definitive diagnosis using conventional evaluations is found in fewer than half of the couples experiencing repeated loss. The aim of this study was to evaluate the inherited heterozygosity of factor five Leiden (FVL) G1691A and Its relation to the time of recurrent spontaneous pregnancy loss. Materials and methods: Retrospective case- control study, in which women with RPL were compared to healthy women without any evidence of spontaneous abortion. This study was undertaken at Omdurman maternity hospital in Khartoum state, Sudan. The case group consisted of one hundred women who experienced at least three or more consecutive recurrent spontaneous pregnancy loss that occurred before 20 weeks of gestation and the control group consisted of ninety five healthy women without any history of adverse pregnancy outcome.Questionnaire and direct interview were used to collect information. Genotyping was based on polymerase chain reaction. Data were entered and analyzed by SPSS program version 17.0. Result: Heterozygosity for FVL alleles G/A was 8.0% in all cases and 6.4% was found in control group. Related to association with time of recurrent pregnancy loss our result shows three times (37.5%), four times (50.0%) and five times (12.5%). Conclusion: Heterozygosity of FV Leiden G1691A could be one reason for recurrent pregnancy loss and pregnancy complications among women with unexplained pregnancy loss. Our study showed that there is an association between heterozygosity for FVL G1691A and time of recurrent pregnancy loss.

Analysis of C3 Gene Variants in Patients With Idiopathic Recurrent Spontaneous Pregnancy Loss.

Miscarriage is the most common complication of pregnancy. Approximately 1% of couples trying to conceive will experience recurrent miscarriages, defined as three or more consecutive pregnancy losses and many of these cases remain idiopathic. Complement is implicated both in the physiology and pathology of pregnancy. Therefore, we hypothesized that alterations in the C3 gene could potentially predispose to this disorder. We performed full Sanger sequencing of all exons of C3, in 192 childless women, with at least two miscarriages and without any known risk factors. All exons carrying non-synonymous alterations found in the patients were then sequenced in a control group of 192 women. None of the identified alterations were significantly associated with the disorder. Thirteen identified non-synonymous alterations (R102G, K155Q, L302P, P314L, Y325H, V326A, S327P, V330I, K633R, R735W, R1591G, G1606D, and S1619R) were expressed recombinantly, upon which C3 expression and secretion were determined. The L302P and S327P were not secreted from the cells, likely due to misfolding and intracellular degradation. Y325H, V326A, V3301I, R1591G, and G1606D yielded approximately half C3 concentration in the cell media compared with wild type (WT). We analyzed the hemolytic activity of the secreted C3 variants by reconstituting C3-depleted serum. In this assay, R1591G had impaired hemolytic activity while majority of remaining mutants instead had increased activity. R1591G also yielded more factor B activation in solution compared with WT. R1591G and G1606D showed impaired degradation by factor I, irrespectively if factor H, CD46, or C4b-binding protein were used as cofactors. These two C3 mutants showed impaired binding of the cofactors and/or factor I. Taken together, several alterations in C3 were identified and some of these affected the secretion and/or the function of the protein, which might contribute to the disorder but the degree of association must be evaluated in larger cohorts.

Embryoscopy and karyotype findings of repeated miscarriages in recurrent pregnancy loss and spontaneous pregnancy loss.

The aim of the study was to assess cytogenetic and embryoscopic characteristics in subsequent miscarriages of spontaneous pregnancy losses (SPL) and recurrent pregnancy losses (RPL). A retrospective cohort of 75 women was affected by repeated pregnancy loss. Of those, 34 had SPL, 24 primary RPL, and 17 secondary RPL. Ploidy status and morphology was analyzed by transcervical embryoscopic examination of the embryo and cytogenetic analysis of the chorionic villi in subsequent miscarriages. Similar rates of recurrent ploidy status were observed between first and second miscarriage in SPL and RPL (82.4% recurrent ploidy status in SPL, p > 0.999; 73% recurrent ploidy status in RPL, p = 0.227). No difference was found regarding recurrent abnormal morphology between SPL and RPL (p = 0.092). However, secondary RPL resulted significantly more often in recurrent abnormal morphology compared to primary RPL (p = 0.004). High rates of recurrent normal/abnormal karyotypes were observed in all groups with a majority of embryos presenting with recurrent abnormal morphology. Secondary RPL presented significantly more often with recurrent abnormal morphology compared to primary RPL. These findings offer prognostic information for the affected patient and might impact treatment choice.

Subcutaneous Immunoglobulin Therapy in a Woman with Spontaneous Pregnancy Loss and Multiple Sclerosis

Background: Off-label use of intravenous immunoglobulin has been reported for the treatment of recurrent spontaneous pregnancy loss and selected cases of multiple sclerosis. However, the use of subcutaneous immunoglobulin has not been reported in these settings. Methods and results: We report on tolerability and clinical efficacy of subcutaneous immunoglobulin therapy in a woman with the unusual association of IgG4 subclass deficiency, C4 hypocomplementemia, localized scleroderma, unexplained spontaneous pregnancy loss and multiple sclerosis. Weekly infusions of subcutaneous immunoglobulin were self-administered at home (6 grams/week, Hizentra®) during the next pregnancy after abortion. Interferon beta-1a for the treatment of relapsing-remitting multiple sclerosis was stopped before pregnancy. During pregnancy only subcutaneous immunoglobulin was administered. The patient was found to have 4% of CD3+CD16/CD56+ (Natural Killer (NK) /T cells). The patient delivered a healthy baby (Apgar score 9 at 5 minutes) at 37 weeks gestation. During pregnancy and 3 months follow-up after delivery no multiple sclerosis flare-ups were observed. NK-T cell percentages remained stable. Conclusions: Our observation indicates that subcutaneous immunoglobulin therapy was well tolerated and efficacious in a selected case of unexplained abortion and multiple sclerosis.

NOD1 and NOD2 control the invasiveness of trophoblast cells via the MAPK/p38 signaling pathway in human first-trimester pregnancy

IntroductionNucleotide binding and oligomerization leucine-rich repeats (NLR) are recognized as members of pattern-recognition receptors that play important roles in host innate immune defenses. In this study, we have investigated the expression and regulation of nucleotide-binding oligomerization domains 1 and 2 (NOD1 and NOD2) on the invasiveness of primary human trophoblasts during the first trimester of pregnancy. MethodsThe expression of NOD1 and NOD2 by human first trimester villi from recurrent spontaneous abortion (RSA) patients and normal pregnancy was analyzed by immunochemistry, western blot and real-time RT-PCR, respectively. The effects of ligands on the regulation of their receptors and invasiveness of trophoblasts were examined. The ELISA was used to measure the secretion of matrix matallopeptidase-9 (MMP9) and matrix matallopeptidase-2 (MMP2) by trophoblasts. We also investigated the signaling pathways involved in invasion of trophoblasts. ResultsThe higher NOD1 and NOD2 were observed in villi from patients who experienced RSA compared with those who experienced a normal pregnancy. The ligands can up-regulate the expression of their receptors. After activation, NOD1 and NOD2 inhibited the invasion of trophoblast cells by decreasing MMP9 but not MMP2. After the activation of NOD1 and NOD2, the mitogen-activated protein kinase (MAPK)/p38 pathway was activated time dependently. Blocking this pathway with a specific inhibitor (SB203580) attenuated NOD1-and NOD2-regulated trophoblast invasions. DiscussionThese results suggest that abnormal elevated NOD1 and NOD2 expression in villi relate to pregnancy outcomes. Further, activation of NOD1 and NOD2 could have a bearing on our understanding of the invasive ability of trophoblasts.

To drive or be driven: the path of a mouse model of recurrent pregnancy loss

This review is an example of the use of an animal model to try to understand the immune biology of pregnancy. A well-known model of recurrent spontaneous pregnancy loss is put in clinical, historical, and theoretical context, with emphasis on T cell biology.

Immunology of Isolated and Recurrent Spontaneous Pregnancy Loss

Immunology of Isolated and Recurrent Spontaneous Pregnancy Loss

Candidate gene study of genetic thrombophilic polymorphisms in pre‐eclampsia and recurrent pregnancy loss in Sinhalese women

Genetic thrombophilias are known to contribute to adverse pregnancy outcomes. Studies in Western populations show that 5, 10-methylenetetrahydrofolate reductase (MTHFR) 677C>T and Factor V (F5) 1691G>A (Leiden) polymorphisms are commonly associated with pre-eclampsia and recurrent spontaneous pregnancy loss. The objective of this study was to investigate the association of MTHFR 677C>T (rs1801133); 1298A>C (rs1801131) and F5 1691G>A (rs6025); 4070A>G (rs1800595) polymorphisms with pre-eclampsia and recurrent pregnancy loss among Sinhalese women in Sri Lanka. Genotype and allele frequencies at each polymorphic site in the MTHFR and F5 genes and the haplotypes defined by them were determined in 175 Sinhalese women with pre-eclampsia, 171 normotensive controls, 200 Sinhalese women with two or more recurrent pregnancy losses and 200 controls with two or more living children and no pregnancy losses. Genotyping was done by polymerase chain reaction/restriction fragment length polymorphism. Odds ratios and χ(2) -testing were performed to compare genotype/haplotype frequencies at each polymorphic site for both cases and controls. The genotype frequencies at each polymorphic site in the MTHFR 677C>T; 1298A>C; F5 1691G>A and 4070A>G genes and the haplotypes defined by them were not significantly associated with either pre-eclampsia or recurrent pregnancy loss. There was no significant association of genetic thrombophilia with either early or late pregnancy losses. The MTHFR and F5 polymorphisms and the haplotypes defined by them were not significantly associated with either pre-eclampsia or recurrent pregnancy loss in this group of Sinhalese women.

Chromosomal aberrations in 2000 couples of Indian ethnicity with reproductive failure

Constitutional chromosomal aberrations contribute to infertility and repeated miscarriage leading to reproductive failure in couples. These aberrations may show no obvious clinical manifestations and remain undetected across multiple generations. However, infertility or recurrent spontaneous pregnancy loss, and/or genotypic/phenotypic aberrations may be manifested in the progeny during gametogenesis. The current study was a retrospective analysis to examine the chromosomal aberrations and prevalence in 2000 couples of Indian ethnicity with reproductive failure. Cytogenetic analysis via conventional G-band karyotyping analysis was carried out on phytohaemagglutinin stimulated peripheral blood lymphocytes, cultured in RPMI1640 medium. The chromosomes were enumerated as per International System for Human Cytogenetic Nomenclature at 500–550 band resolution, and recorded in the screening sheets. Chromosomal aberrations were detected in a total of 110 (2.78%) couples, with structural chromosomal aberrations in 88 cases including reciprocal translocations in 56 cases, Robertsonian translocations in 16 cases, inversions in eight cases, deletions in three cases, derivative chromosomes in five cases and numerical chromosome aberrations in 23 cases. The study emphasizes the importance of cytogenetic work up in both the partners associated with a history of reproductive failure. Genetic counselling with an option of prenatal diagnosis should be offered to couples with chromosomal aberrations.Alteration in the number or structure of chromosomes is associated with adverse obstetric outcome in the form of infertility or recurrent miscarriages. Most of these aberrations are balanced and hence, the person does not manifest any obvious clinical signs and symptoms. However, because of the formation of abnormal gametes (i.e. the eggs and sperm), these chromosomal aberrations result in infertility and recurrent spontaneous pregnancy losses. In the present study, we studied 2000 couples of Indian ethnicity with reproductive failure. Karyotyping was done on the blood sample of these couples using standard protocols. Chromosomal aberrations were detected in a total of 110 (2.78%) couples with structural chromosomal aberrations in 88 cases including reciprocal translocations in 56 cases, Robertsonian translocations in 16 cases, inversions in eight cases, deletions in three cases, derivative chromosomes in five cases and numerical chromosome aberrations in 23 cases. Thus, our study emphasizes the importance of cytogenetic work up in both the associated partners with history of reproductive failure which would help in better patient counselling and management.

Study of methyl transferase (G9aMT) and methylated histone (H3-K9) expressions in unexplained recurrent spontaneous abortion (URSA) and normal early pregnancy

We investigated the expression of methyl transferase G9a and methylated histone H3-K9 in fresh human decidual/endometrial tissue of 12 normal early pregnancies and 15 unexplained recurrent spontaneous abortions (URSA). The samples were obtained through dilatation and curettage and collected as per strict inclusion-exclusion criteria. The tissue was subjected to immunohistochemical analysis (IHC), western blotting (WB) and RT-PCR analysis. The results demonstrated methyl transferase G9a to have a lower expression in abortions when compared with that in normal pregnancy (P < 0.05). The sensitivity of RT-PCR, IHC and WB were respectively 66.67, 75 and 71.43%, while specificity of the same were 66.67, 60 and 78.92%, respectively. Methylated histone H3-K9 was significantly lower (P < 0.0001) in URSA tissues than in controls. This study suggests that methylation may cause URSA and indicates the need for further work to explore the role of methylation in URSA and its possible prevention through locally acting methylating/demethylating agents.

Role of anti-human lymphocyte culture cytotoxic antibodies in recurrent spontaneous pregnancy loss women

BACKGROUND:Recurrent spontaneous pregnancy (RSA) is defined as a sequence of three or more consecutive spontaneous abortions. One of the major causes of RSA is immunological where alloimmune antibodies develop towards human leucocyte antigen (HLA) antigens. Earlier research had suggested that anti-HLA antibodies are produced in normal women; studies have been reported that normal pregnant women develop anti-HLA antibodies, mostly after 20–28 weeks of gestation.AIM:To evaluate the role of anti-HLA antibodies in RSA patientsMATERIALS AND METHODS:A total of 80 randomly selected couples with unexplained three or more RSA and control group of 50 normal pregnant women were screened for anti-HLA A and B antibodies. The anti-HLA antibodies were analyzed following the standard two-stage NIH microlymphocytotoxicity assay.RESULTS:In our study group a high frequency of anti-HLA antibodies among women with RSA (26.25%) was detected compared to normal pregnant women (8.0%). Most of the sera showed HLA-A and HLA-B antibodies which had high titer, up to a dilution of 1: 4096.CONCLUSION:This incidence of high anti-HLA antibodies in RSA women during early weeks of gestation may explain the recurrent pregnancy loss.