Recurrent pregnancy loss (RPL), also called recurrent abortion (RA), or a recurrent spontaneous miscarriage, is practically defined as three or more consecutive pregnancy losses of clinically recognized pregnancies up to 20 weeks of gestation without taking into account ectopic, molar and biochemical pregnancies. This definition applies to indications for clinical treatment in many developed countries. The reason was a decrease in the number of babies born in each family. The incidence of RPL should be approximately 1 in 300 pregnancies, based on the frequency of sporadic pregnancy loss. RPL brings not only grief for the patient, but also physical damage to female reproductive organs and affects 2-5% of all couples.
In addition, regardless of how early pregnancy occurs the loss of a child is almost the same as a child’s loss to the patient. Therefore, RPL has a serious impact on all associated mental and physical feelings. RPL only unites these feelings and can lead to increased stress, anger, frustration, feelings of loneliness and despair, emptiness and a sense of self-worth and failure. Although no specific data determined the probability of finding an etiology for RPL in a population with two or three miscarriages, it was suggested that the risk of a subsequent miscarriage in the following pregnancies is 30% after two losses, compared to 33% after three losses among patients without a history of live birth.
Diagnostic testing and therapeutic intervention for RPL as a clinical subject are based on an understanding of the increased risk for subsequent fetal loss and the underlying etiology of the disorder. Thus, a significant role for evaluation after two losses, in patients without prior live births, is strongly recommended in general. If embryonic cardiac activity was identified before loss, a prophylactic evaluation should be considered in women over 35 years of age, or the couple had infertility treatment. There are few etiologies common to the RPL.
Approximately 2-4% of RPL is associated with parental balanced structural chromosome rearrangement, most often balanced mutual or Robertsonian translocations. These structural rearrangements have already been approved for treatment with preimplantation genetic diagnosis (PGD) using assisted reproductive technology (ART) to avoid subsequent miscarriage. There are other structural anomalies associated with RPL, such as chromosome inversions, insertions and mosaicism. There may be some association with RPL, including some individual gene defects, such as cystic fibrosis or sickle cell anemia. Parental karyotyping should be performed to evaluate RPL with genetic counseling. Appropriate genetic counseling is indicated in all cases of RPL associated with pathologies of parental chromosomes.
The final treatment may require ART with PGD, depending on the specific diagnosis. Preimplantation genetic screening (PGS) is explained in detail in the following paragraphs. The use of donor gametes for treatment can be suggested in cases of genetic abnormalities that always lead to embryonic aneuploidy, such as Robertson’s translocations involving homologous chromosomes.
Approximately 10-15% of the causes of RPL are derived from the anatomical malformation of female reproductive organs and are generally believed to cause miscarriages due to an insufficient vascular network of the endometrium, which causes abnormal and inadequate placenta.
There are some alleged causes, which are congenital abnormalities of the uterus, intrauterine adhesions and uterine myomas or polyps. These anomalies are considered potential causes of RPL through interruption of vascular nutrition in the endometrium.
The most closely related to RPL is the uterine septum, a congenital abnormality of the uterus, with a 76% risk of spontaneous loss of pregnancy among affected patients. Other Müllerian abnormalities, including unicornuate, didelphic and bicornuate uteri, were associated with a smaller increase in RPL risk. The uterus may or may not cause the RPL. The presence of intrauterine adhesions, sometimes associated with Asherman’s syndrome, can significantly affect the placentation and lead to an early pregnancy. Intramuscular fibroids more than 5 cm, as well as sub-mucosal fibroids of any size can be associated with RPL.
Myomectomy should be considered in cases of sub-mucosal fibroids or any type of fibroid more than 5 cm, especially in patients with infertility treatment. It is noteworthy that the improvement in fertility rates was shown with myomectomy from 57 to 93%. Myomectomy can be performed with open laparotomy, laparoscopy or hysteroscopy. Congenital anomalies caused by prenatal exposure to diethylstilbestrol (DES) are well known in the case of RPL. Nevertheless, the impact of the DES administration becomes less clinically significant, since most affected patients go beyond the reproductive age. Anatomical abnormalities of the uterus should be assessed using either office hysteroscopy, or hysterosalpingography (HSG). If these anomalies are identified, resection of intrauterine adhesions or intrauterine partitions should be performed hysteroscopically. Successful resection of the hysteroscopic septum brings almost normal pregnancy results to patients, with a delivery time of about 75%, and a fertility rate of about 85%.
Endocrine disorders, such as luteal insufficiency, polycystic ovary syndrome (PCOS), diabetes mellitus, thyroid disease and hyperprolactinaemia, can be associated with RPL by about 17-20%. Luteal failure was defined as insufficient production of progesterone by the corpus luteum and insufficient maturation of the endometrium for implantation. However, a definite effect of luteal insufficiency on RPL is controversial, and endometrial biopsies for diagnosing luteal insufficiency are less satisfied. Insulin resistance with resultant hyperinsulinemia may play a role in RPL of patients complicated with PCOS, as well as in type 2 diabetes mellitus, as treatment of these patients with an insulin sensitizer, metformin, reduces the rate of spontaneous pregnancy loss.
There is evidence of PCOS, at least 40% of women with RPL. Poorly controlled Type I diabetes mellitus is also associated with an increased risk of spontaneous abortion. Untreated hypothyroidism is clearly associated with spontaneous abortion and RPL, but the relationship between antithyroid antibodies and RPL in patients with euthyroidism is currently being studied. There is evidence that euthyroid women with antithyroid antibodies, especially those undergoing infertility treatment, are likely to become clinically hypothyroid when they reach pregnancy.
Because the results of pregnancy in these women can improve with early (perhaps prenatal) replacement of the thyroid hormone, currently similar approaches are being studied among women with RPL. Evaluation of endocrine disorders should include measurement of the level of thyroid-stimulating hormone (TSH). Other tests that may be indicated based on a patient’s presentation include testing insulin resistance, testing the ovarian reserve, serum prolactin in the presence of irregular menstruation, testing antithyroid antibodies and, very rarely, endometrial biopsy of the luteal phase. Recently, the popularity of therapy with insulin sensitizers for the treatment of RPL, which occurs in the presence of PCOS, has gained popularity.
The role of infectious diseases in the RPL has not yet been clarified, but a frequency of 0.5-5% is proposed. There are some potential infectious diseases, such as Listeria monocytogenes, Toxoplasma gondii, rubella, herpes simplex virus (HSV), measles, cytomegalovirus and coxsackie viruses. Infectious diseases can cause pregnancy loss by the following mechanisms, such as direct infection of the uterus, fetus or placenta, placental insufficiency, chronic endometritis/endocervicitis, amnionitis or intrauterine different infection. Infections of mycoplasma, ureaplasma, Chlamydia trachomatis, L. monocytogenes and HIV presumably play a role in the RPL. Chronic infection is the most significant risk of secondary relapse of the RPL for the acute stage in a patient with weakened immunity. Assessment for chronic infections can be justified for these patients. In general, the prevention of infectious diseases is not necessary, but favorable for the patient RPL, to alleviate their anxiety.
Alloimmune (Histocompatible) Disorder
It is reasonable to assume that there are immunological events that must occur so that the mother can carry the fetus throughout the pregnancy without rejection, because the fetus is not genetically identical to the mother. Therefore, in these immunological mechanisms, there may be deviations that can lead to both sporadic and recurrent pregnancies. Despite the intense interest in this potential etiology for the RPL, there is no consensus on appropriate diagnostic work or therapy. It has been shown that therapy such as paternal leukocyte immunization, intravenous immune globulin, third party donor immunization and infusion of trophoblast membranes, do not significantly improve fertility rates and are only available for use in some areas.
Antiphospholipid Antibody Syndrome (APS)
Antiphospholipid antibody syndrome (APS) is characterized by the presence of at least one clinical and one laboratory criterion. If you need details of APS, please refer to the literature. APS has a strong connection with the RPL, but especially in the second trimester. The ratio of APS to RPL has not been clarified yet, but it is thrombophilia, which is the most commonly acquired risk factor. There is a risk of thrombophilia with a prevalence of 3-5% in the general population. The RPL assessment associated with APS should include at least testing of antibodies against anticardiolipin and lupus erythematosus. Treatment recommendations include low-dose aspirin (LDA: 81-100 mg/day) and/or low molecular weight heparin. LDA should begin before conception or with a positive pregnancy test, but heparin should begin with a positive pregnancy test. Heparin is safe during pregnancy because of a large complex of molecules that do not cross the placenta.
This problem occurs mainly among Caucasian people. Inherited and combined inherited/acquired thrombophilia are common with more than 15% of the white population carrying the inherited thrombophilic mutation. Factor V Leiden mutation is the most common. This is a mutation in the promoter region of the prothrombin gene and a mutation in the gene coding for methylenetetrahydrofolate reductase (MTHFR). These common mutations are associated with mild thrombotic risks, and this remains controversial for homozygous MTHFR mutations associated with vascular disease in general. The potential relationship between RPL and heritable thrombophilia is based on a theory that disrupts the development and functioning of the placenta.
This mechanism causes venous and/or arterial thrombosis, and then causes miscarriage. Loss of pregnancy by this type of thrombophilia occurs for more than 10 weeks of pregnancy, and not until 10 weeks of pregnancy, because maternal blood begins to flow in the interspecies spaces of the placenta after about 10 weeks of pregnancy. Of course, the transfer of nutrition from maternal blood to the fetal tissue depends on the uterine blood flow, regardless of the age of gestation.
Thus, thrombotic events occurring in any gestational age play the role of thrombophilia in the loss of pregnancy. Hereditary thrombophilia associated with RPL include hyperhomocysteinemia resulting from mutations of MTHFR, activated protein C, associated with factor V Leiden mutations, protein C and protein deficiencies, mutations of the prothrombin promoter and antithrombin mutations.
Acquired thrombophilia associated with RPL include hyperhomocysteinemia and resistance to active protein C. Certain causal relationships between these inherited and acquired states have yet to be discarded. Nevertheless, testing of factor V Leiden mutations, protein S levels, prothrombin promoter mutants, homocysteine levels, and resistance to global activated protein C are suitable targets for treatment selection. After determining the diagnosis, appropriate therapy should be initiated for inherited or acquired thrombophilia. Specific for an individual disorder should be performed as follows: the addition of folic acid for patients with hyperhomocysteinemia and preventive anticoagulation in cases of isolated defects without a personal or family history of thrombotic complications. Therapeutic anticoagulation should be performed in cases of combined thrombophilia defects.
Alcohol, Smoking, and Caffeine
Patients are often particularly concerned about the possibility that the effects of the environment can lead to losses in pregnancy, because of its tendency to the emergence of responsibility and guilt. Association RPL and professional and/or environmental impacts of organic solvents, medicines, ionizing radiation and toxins have been suggested, as always, but it is difficult to make firm conclusions from the fact that they tend to retrospective and mixed with alternative or additional environmental influences.
Bad habits, such as smoking, alcohol and caffeine, are the three main themes that are widely used and can be changed. Maternal alcoholism (or frequent use of stupefying amounts of alcohol) is known with a higher incidence of spontaneous miscarriage, but the association with a more moderate ingestion remains controversial. Smoking cigarettes, apparently, can increase the risk of spontaneous abortion, based on the intake of nicotine, a potent vasoconstrictor that is known to reduce uterine and placental blood flow.
Least, the relationship between smoking and pregnancy. As for the consumption of caffeine, there is some evidence that caffeine, even in amounts equal to three to five cups of coffee a day, can increase the risk of spontaneous pregnancy. The relationship between consumption of caffeine, alcohol and nicotine with RPL is even weaker than their association with sporadic loss.
Direct and indirect interventions for patients with RPL are described in previous sections. However, when all known and potential causes of RPL are identified, almost half of the patients will be left without a definitive diagnosis. Optimal management of these patients is often as fuzzy as the etiology of their RPL. It has been shown that progesterone is useful for reducing the incidence of miscarriages among women who have experienced at least three lesions. Treatment with a low dose of aspirin (LDA) has also been investigated as a potential therapy for unexplained RPL. It has been proven that one use before and during pregnancy increases the birth rate among women with previous miscarriages exceeding 13 weeks of pregnancy. In fact, the most effective therapy for patients with unexplained RPL is often the simplest: antenatal counseling and psychological support. It was shown that these remedies have the following indicators of the success rate of pregnancy in 86% compared with the success rate of 33% in women who received no additional antenatal care.
Aneuploid Embryo and RPL
Although the exact causes of RPL have not been elucidated and are still inexplicable or idiopathic, some reasons can be explained by various factors, such as described above. The quintessential possibility of these causes for idiopathic RPL is that these couples produce more aneuploid embryos, which leads to a higher miscarriage.
Widely reported on the role of chromosomal abnormalities in miscarriage, while 50-70% of miscarriages of the first trimester were attributed to aneuploidy. In addition, it has been demonstrated that analysis of malignant neoplasms of the fetus can explain 80% of unexplained RPL in elderly women. Many authors confirmed a higher level of aneuploidy in patients with RPL.
Preimplantation genetic screening (PGS) was proposed as a method of reducing miscarriage by selecting embryos of euploids for transmission because of the prevalence of aneuploidy in losses in the first trimester and an increase in the prevalence of aneuploidy in the RPL population. Existing standards of care for patients with unexplained RPL, supported by the American Society of Reproductive Medicine (ASRM), are expectant management. However, the emotional trauma that may accompany clinical miscarriages and the perceived urgency for conception, felt by many patients with RPL, leads them to alternative treatment options, including assisted reproductive technologies, and in particular to in vitro fertilization (IVF) and PGS.
Thus, PGS for indicating idiopathic RPL is that euploidy embryos can be selected for embryo transfer, which leads to a decrease in the rate of pregnancy loss in patients with idiopathic RPL. All studies using PGS for this indication evaluated that the frequency of miscarriages after this procedure showed a decrease. Again, it is widely recognized that aneuploidy is the most common genetic abnormality in embryos and the most common cause of miscarriage. Regardless of how well the patient is treated, if the implanted embryo was aneuploidy, it never works.
Is PGS Useful for Patients With RPL?
The term “aneuploidy” was used to describe the loss or enhancement of the genetic material of the chromosome (s) from the time of the first patient with aneuploidy. Since then, it has been proven that aneuploidy is a very common cause, accounting for at least 15-20% of all clinical pregnancies. Most aneuploid embryos will never lead to clinical pregnancy and live birth, which makes aneuploidy the main cause of miscarriage, but some of them are compatible with live birth, which makes aneuploidy the main cause of congenital malformations and mental retardation. Aneuploidy has been identified as an important factor contributing to the failure of the IVF cycle, especially the failure of implantation and/or spontaneous abortion in the area of assisted reproductive technology (ART). However, recent advances in reproductive medicine and molecular cytogenetics have completely changed the treatment protocol for infertile couples suffering from repeated aneuploidic losses.
Genetic testing, such as chorionic villus sampling, amniocentesis and a NIPT (noninvasive prenatal test) from maternal blood, was prenatally available. When these methods are applied, if adverse results are detected, further restoration of the live fruit will still be necessary. Fluorescence in situ hybridization (FISH) was the main methodology of PGD or PGS over the past two decades, and anorexia, discovered using FISH technology with reference to infertility, was presented at the beginning. Despite the confirmation of the high rate of aneuploidy, both with repeated failures of IVF and with miscarriages, the improvement of the results of IVF with PGS for FISH has not been successfully demonstrated. These earlier studies were routinely performed using the FISH estimation of embryos of the cleavage stage and usually tested only 7-12 chromosomes. In one meta-analysis, four observational studies were evaluated in which fertile patients with RPL underwent biopsy at two stages of cleavage by 1-2 cells and compared with patients with the natural concept of RPL.
All four studies conducted FISH screening for 3-9 chromosomes. The incidence of spontaneous abortion (SABR) ranged from 0 to 10% (an average of 9%) in patients with RPL with PGS compared with 14-52% (averaging 28%) with a natural concept (P = 0.0013).
After that, the technology of the CGH array appeared in the analysis of all 24 chromosomes, in contrast to FISH, which allowed obtaining more accurate results when aneuploidy was detected. There are several methods of complex chromosomal screening (CCS), including a single nucleotide polymorphism (SNP) array, CGH and a quantitative polymerase chain reaction (PCR). Comparison of FISH with the SNP array showed up to 60% false positive rate with FISH. When FISH was compared to CCS, it was found that mosaicism was three times more common in FISH. Therefore, the European Society for Human Reproduction (ESHRE) recently recommended replacing this method with complex screening methods. In conclusion, PGS by CCS should be applied to RPL patients in the modern era of ART.
The Use of PGS for Patients with RPL
It is difficult to find the ideal control group for RPL studies to determine whether PGS is beneficial to reduce miscarriage. The question is whether to compare the RPL couple with other PGS-passing couples, with or without infertility, or only with the RPL history in PGS, and found that PGS using FISH significantly reduces the level of miscarriages, from 36% of the expected rate to 13 %. Patients who were offered PGS but rejected it had a 44% incidence of miscarriage, which is also another way of comparing patients with RPL using PGS with appropriate controls. This positive effect of PGS on RPL was observed in both fertile and infertile patients with RPL undergoing IVF.
However, these studies used FISH, evaluated a limited number of chromosomes, and used day biopsies of three embryos, indicating that it could adversely affect the potential for implanting a biopsy embryo, whereas blastocyst biopsy does not appear harmful. The clinical efficacy of IVF and PGS compared to survival, which is the current standard of treatment for the treatment of patients with RPL, has not been investigated by longitudinal prospective studies or randomized clinical trials. In addition, IVF-PGS is an expensive treatment option, and it is necessary to examine the cost-effectiveness of IVF-PGS compared to the expected management.
However, a recent follow-up report demonstrated the positive effect of PGS on patients with RPL. He concluded that patients with RPL initiating PGS have a significantly higher level of LBR compared to the expected control without significant differences in the incidence of miscarriage. The level of miscarriage is likely to be lower if all patients with IVF intending PGS will complete the cycle as expected, as aneuploidy is the common cause of miscarriage of the first trimester. Of course, further research is needed to investigate the cost-effectiveness of this treatment strategy for fertile patients with RPL.
Rationale for Applying PGS to RPL
It is widely recognized that aneuploidy is the main cause of failure of implantation and miscarriage in both fertile and infertile couples seeking to achieve pregnancy. Cytogenetic analysis of previous miscarriages is an important component in evaluating couples with a history of pregnancy loss, as it can lead to subsequent treatment. In addition, the PGS field for screening aneuploidy also provided an opportunity to understand cell division errors, which eliminated the potential failure of implantation due to aneuploidy. In addition, the use of PGS has positively affected the success rates of IVF in some cases, worldwide is approaching transmission with one embryo and reduces the number of deliveries after ART. The most common numerical chromosomal abnormality (NCA) occurs both before implantation after IVF, and after implantation (D&C) in infertile population pursuing pregnancy.
Despite the fact that the frequency of cases of monosomy and trisomy was observed before implantation, it was rarely observed in these embryos implanted. These results protect PGS technology as an advantageous facilitator, which helps circumvent the inheritance of aneuploidy. Performing PGS will significantly reduce the incidence of NCA, thereby reducing the likelihood of failure of implantation and/or miscarriage after IVF. Given this knowledge, one can expect that, after the IVF cycle, the implantation and pregnancy rates for one transfer will be transferred, by eliminating the unnecessary transfer of embryos, preventing both the rejection of implantation and early miscarriages.
Many researchers analyzed the incidence of NCA among human fetuses by analyzing chorionic villi after miscarriage and reporting a prevalence of NCA in the range of 40 to 80%. However, as a rule, these messages are biased because of the high rate of spontaneous reduction of chromosomal abnormal pregnancies before pregnancy was clinically recognized, as well as the absence of patients who increasingly opt for cytogenetic analysis of their POPs. It is noteworthy that when these results were compared with tissue harvested after D&C, monosomy was rarely observed, and trisomes most often appeared on chromosomes 22, 16, 21, 15 and 19 (the order reflects the frequency).
NCA are present at a high frequency, taking root from early development. The previous molecular genetic analysis of chromosomal abnormalities arising from miscarriages showed that most aneuploid events occur during female meiosis, usually because of non-disection in the first meiotic division. Direct observation of female meiotic divisions (by means of polar bodies analysis) and early embryonic stages showed that a wide spectrum of aneuploidia is present before implantation. Historically, most studies have focused on trisomy, especially those that are compatible with live births.
On the basis of these analyzes, three “rules” of human non-disection were formulated: first, regardless of the specific chromosome, most trisomies occur during oogenesis; secondly, for most chromosomes, maternal MI errors are more common than those of maternal meiosis II (MII); and, thirdly, the proportion of cases of maternal origin increases with age. PGS continues to evolve, but the current PGS method does not detect definite polyploidy. The effectiveness of genomic technologies could reveal abnormal embryos, which otherwise might seem morphologically normal. PGS helps the decision-making process before embryo transfer, revealing any NCA that potentially avoids early pregnancy loss.
The discrete application of embryonic screening can maximize implantation and fertility and minimize the frequency of miscarriages associated with chromosomal abnormalities. Moreover, understanding the etiology of reproductive losses can ease the patient’s sense of guilt or irresponsibility in treating infertility. Nevertheless, further research can serve as a guide for optimizing or even improving the quality of oocytes, such as pronuclear transmission, spindle maternity transfer or nuclear genome transfer, as well as non-invasive methods for identifying reproductive-competent embryos to improve clinical outcomes.
The etiology of recurrent pregnancy loss is not well identified, but there are some patients with effective treatment. However, any treatment cannot overcome the aneuploid embryo,
Chromosomal errors, aneuploidy, in human embryos are the main cause of failure of ART, miscarriage, obstetric complications, stillbirth and infertility and lead to the birth of affected children,
An accurate technology for the detection of chromosomally normal embryos of euploids exists and is now available for clinical use,
Recent technologies such as the CGH array, qPCR and NGS, can detect all 24 chromosomes with high accuracy and should be used for PGS in patients suffering from IVF failure and/or pregnancy recurrence,
PGS can shorten the time before birth, choosing only embryo euploids and reducing the incidence of miscarriage,
PGS allows a single transfer of embryos of euploids, which maximizes the probability of a child’s birth without the risk of multiple pregnancies,
Any treatment plan for RPL cannot prevail over aneuploid embryos,
Patients do not have embryo transfer, because embryos from euploids prefer to know reality. They can move on in the future, rather than waiting for a failure or a possible miscarriage or the birth of the affected child,
Yes, PGS is permissible to prevent recurrent pregnancy loss; however, ART is vital for a patient who thought naturally.