Article
Insertion/deletion polymorphism in intron 16 of ACE gene in idiopathic recurrent spontaneous abortion: case-control study, systematic review and meta-analysis

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Abstract

The insertion/deletion (I/D) polymorphism in intron 16 of the angiotensin I-converting enzyme gene (ACE) has been extensively studied as a predisposing factor for idiopathic recurrent spontaneous abortion (IRSA). A case-control study including 149 women with ≥3 spontaneous abortions and 149 controls was performed to test the association of ACE I/D polymorphism with IRSA. A systematic review was conducted of previous case-control studies, with strict selection criteria for meta-analyses. We also aimed to evaluate the potential differences in summary estimates between studies defining IRSA as ≥2 and ≥3 spontaneous abortions. Genotyping was performed by PCR, and systematic review conducted using PubMed and Scopus. There was no association of the polymorphism with IRSA in Slovenian women. Sixteen case-control studies, showing substantial differences regarding IRSA definition and selection criteria for women were identified. Meta-analysis was performed and included four studies defining IRSA as ≥2 spontaneous abortions and the current study, which defined IRSA as ≥3 spontaneous abortions. Based on random effects model, meta-analysis conducted on 1192 patients and 736 controls showed no association with IRSA under dominant(DD+IDvsII) and recessive(DDvsID+II) genetic models. Well-designed studies are needed to evaluate the role of ACE I/D polymorphism in IRSA defined as ≥3 spontaneous abortions.

Introduction

The renin-angiotensin system (RAS) is an important regulator of human pregnancy, during which the maternal and fetal circulating RAS interact with various tissue RAS, including those of the ovaries and fetoplacental unit (Irani, Xia, 2011, Lumbers, Pringle, 2014). At the systemic level, the RAS is involved in the regulation of blood pressure and volume, haemostasis and homeostasis of water and electrolytes, whereas at the fetoplacental unit it influences implantation, placentation, angiogenesis and uteroplacental blood flow (Irani, Xia, 2011, Lumbers, Pringle, 2014, Nielsen et al, 2000). Therefore, abnormalities in the RAS have been suggested as a potential contributing mechanism to different pregnancy complications, including idiopathic recurrent spontaneous abortion (IRSA).

According to the evidence-based guidelines for the investigation and treatment of RSA, the condition is characterized by three or more (≥3) consecutive spontaneous abortions (Jauniaux et al., 2006), although many scientific groups include couples with two or more (≥2) spontaneous abortions in their research. The aetiology of RSA cannot be determined in approximately 60% of cases. Considering that the expression of genes encoding RAS components, including renin, angiotensinogen (AGT), angiotensin I-converting enzyme (ACE), AGT II type 1 and 2 receptors (AGT1R, AGT2R), is in part influenced by gene variations (Jeunemaitre, 2008), a large number of studies investigated their association with IRSA. The insertion/deletion (I/D) of a 287 bp sequence in intron 16 of the ACE gene is the most commonly tested variation; however, there is no conclusive evidence on its role in the pathogenesis of IRSA.

Angiotensin I-converting enzyme is a zinc metallopeptidase that converts the inactive angiotensin I to active angiotensin II (ANG II), and in addition cleaves bradykinin and several other peptides (Masuyer et al., 2014). Although the ACE I/D polymorphism in intron 16 is associated with alterations in circulating and tissue concentrations of ACE, evidence suggests that it is not a functional polymorphism, but is linked with other, yet unknown, intragenic functional variations (Sayed-Tabatabaei et al., 2006). Nevertheless, the DD genotype is accompanied by increased serum ACE and plasminogen activator inhibitor-1 (PAI-1) concentrations compared with the II genotype (Rigat et al, 1990, Tiret et al, 1992).

According to the first genetic-association study, the assumption for a possible association between the ACE I/D polymorphism in intron 16 and IRSA was based on the well-known roles of ANG II in the control of vascular tone and fibrinolysis, which, if disturbed, might lead to abnormal development of placental vasculature and disturbances of haemostasis (Fatini et al., 2000). Afterwards, a series of studies in different populations continued the search for a potential association, but results did not confirm one. A qualitative and quantitative synthesis of results was performed in three studies; however, we observed that the process of study selection was not performed in accordance with the proposed eligibility criteria (Su et al, 2013, Wang et al, 2013, Yang et al, 2012). Therefore, we performed a case-control study to test the association of ACE I/D polymorphism in intron 16 with IRSA in Slovenian women and conducted a thorough systematic review of previous case-control studies. Finally, based on precisely chosen inclusion criteria for meta-analyses, we aimed to evaluate whether there is a difference in summary estimates between studies in which IRSA is defined as ≥2 and ≥3 spontaneous abortions.

Section snippets

Subjects

In order to test the genetic association between ACE I/D polymorphism in intron 16 and IRSA, a case-control study in Slovenian women was conducted. One hundred and forty-nine women with a history of ≥3 consecutive spontaneous abortions of unknown aetiology before the 22nd week of gestation, with the same partner, and 149 control women were included in the study. Exclusion criteria for IRSA women were: endocrine disorders, antiphospholipid syndrome (APS), autoimmune or systemic diseases, venous

Case-control study

The power of the present study was 100% to detect a twofold increase in ACE I allele frequency. No statistically significant differences were found in the distribution of genotype and allele frequencies between IRSA and control women or between women with primary and secondary IRSA (Table 1). Genotype frequencies were in HWE in all groups. Nor did the study determine statistically significant associations of ACE I/D polymorphism in intron 16 with IRSA under any genetic model (Table 2).

Systematic review

The

Discussion

In the present study an extensive examination of the role of ACE I/D polymorphism in intron 16 in IRSA women was performed through a case-control study, systematic review and meta-analysis.

The results of the case-control study suggest that ACE I/D polymorphism in intron 16 is not associated with IRSA in Slovenian women. Although it was not possible to include this study in a meta-analysis of studies defining IRSA as ≥3 spontaneous abortions, the results are comparable with those obtained in the

Acknowledgements

This study was supported by research grants “Genetic factors in the aetiology of idiopathic recurrent spontaneous abortion” (University of Rijeka, Croatia, number 13.06.1.3.32) and “Gynecology and Reproduction: Genomics and Stem Cells” (Slovenia, number P3—0326).

Nina Pereza, MD, PhD is a postdoctoral researcher at the Faculty of Medicine, University of Rijeka. Her main interest of research is reproductive genetics, especially genetic causes of idiopathic recurrent spontaneous abortion.

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