Pregnancy is an acquired hypercoagulable state and women with a prior tendency to thrombosis may develop clinical symptoms of placental vascular complications such as preeclampsia, intrauterine growth restriction, and fetal death for unknown causes, that impact the maternal-fetal morbidity and mortality [¹–⁴]. Recurrent pregnancy loss is an important obstetric complication with a prevalence of 1–5% [⁵]. Inherited thrombophilia has been postulated as a cause of recurrent pregnancy loss, although the association between inherited thrombophilia and recurrent pregnancy loss has not been conclusively established. Some studies have demonstrated an association between recurrent pregnancy loss and prothrombotic states rendered by some genetic single nucleotide polymorphisms (SNPs), such as factor V Leiden G1691A (FV Leiden), prothrombin G20210A (FII G20210A), and methylenetetrahydrofolate reductase C677T (MTHFR C677T), and activated protein C resistance (APC resistance) [⁶–¹⁴], whereas others have reported lack of any association [¹⁵–¹⁹]. In addition, a retrospective cohort study showed that women with deficiencies of antithrombin (AT), protein C (PC), or protein S (PS) have an eightfold increased relative risk of thrombosis during pregnancy compared to controls [²⁰].

Most of the studies on thrombophilia and recurrent pregnancy loss have been conducted in Caucasian populations [²¹, ²²]. Therefore, the association between these thrombophilias and recurrent pregnancy loss is almost unknown in triethnic populations such as the Colombian population whose genetic mixture is approximately 70% Caucasian, 15% Amerindian, and 15% African [²³, ²⁴]. We conducted this study to evaluate the association between inherited thrombophilia and recurrent pregnancy loss in a group of patients from Colombia.

Demographics of the women from the case and the control groups are shown in Table 1. In 93 cases and 206 controls, the frequency of thrombophilia-associated SNPs (as shown in Table 2) for both FV Leiden and FII G20210A heterozygous carriers was one patient and one control (1.1% versus 0.5%, resp.; OR 2.25; 95% CI, 0–83). Homozygous carriers of the MTHFR 677T allele were detected in 12 patients and 30 controls (12.9% versus 14.9%, resp.; OR 0.85; 95% CI, 0.4–1.8). APC resistance was found in 5 of 87 patients versus 8 of 187 controls (5.7% versus 4.3%, resp.; OR 1.36; 95% CI, 0.4–4.8) of which only one women in each group was attributable to FV Leiden. Additionally, none of the patients and only two of the controls had <50% AT activity (0 versus 1%, resp.; OR 0; 95%, CI 0–9.2), and 2 of 91 patients versus 3 of 195 controls showed <50% in PC activity (2.2 versus 1.5%, resp.; OR 1.44; 95% CI, 0.2–10.8), as shown in Table 3. Since the determinations of PS levels were nonreproducible in stored samples, these measurements were excluded from analysis in this study. No association was observed between recurrent pregnancy loss and inherited thrombophilia by either SNP genotypes (FV Leiden, FII G20210A, and MTHFR C677T) or functional phenotypes (APC resistance, PC deficiency, and AT deficiency).

The present study is the first to report the frequency of inherited thrombophilia (thrombophilic polymorphisms and functional tests) in a group of Colombian patients with recurrent pregnancy loss. Thrombophilia-associated SNPs such FV Leiden and FII G20210A were very low in both patients and controls, and although the frequency of MTHFR C677T SNPs was high in the patients, differences were not significant. Other inherited thrombophilias such as PC and AT deficiency are not commonly found in patients who had already experienced recurrent pregnancy loss. Only three patients with PC deficiency and no patients with AT deficiency were found among 93 cases. Overall, we found a low frequency of inherited thrombophilia in our patient population unlike Caucasian populations; therefore, it is unlikely that the thrombophilic alterations studied play a role in the pathogenesis of recurrent pregnancy loss in this Latin American population.

The low frequency of FV Leiden in our study agrees with two previous reports on the Colombian population. Camacho Vanegas et al. tested 150 healthy individuals from Bogota (30–50 years old) using PCR-RFLP and found no evidence of FV Leiden [³⁵]; however, the allelic frequency of the MTHFR C677T SNP was 0.487, one of highest reported in the literature (70/150 heterozygous CT and 38/150 homozygous TT) [³⁵]. The other study was performed by Varela et al. and used PCR-sequence specific oligonucleotide probes in 495 Colombian blood donors from banks in four cities, reporting a prevalence of FV Leiden of 1.9% (7/370) in individuals from Bogota, while it was 0% in 125 individuals from the other three cities (Medellín, Bucaramanga, and Barranquilla) [³⁶]. In addition, the presence of inherited thrombophilia in 100 Colombian patients with deep venous thrombosis and in a control group (n = 114) was evaluated with the following results in patients versus controls: 10% versus 0.9% were heterozygous for FV Leiden, 3% versus 0% were heterozygous for FII G20210A (one patient was homozygous), 24% versus 19.3% were homozygous for MTHFR 677T, and 25.2% versus 5.3% had APC resistance (these thrombophilias reached statistical significance, except for the MTHFR polymorphisms) [³⁷]. During this research, the first homozygous family for the G20210A prothrombin polymorphism reported in Latin America, although only some carriers developed venous thrombosis [³⁸].

The FV Leiden, as genetic factor involved in the etiology of thrombosis, has motivated research of this abnormality as a risk factor for recurrent pregnancy loss, as reviewed by Krabbendam et al. [³⁹]; however, this association has not been easy to confirm in multiple studies, making it necessary to reevaluate the experimental design and population size. For example, Nurk et al. [⁴⁰] evaluated FV Leiden in 5874 women and found that this polymorphism is a risk factor for pregnancy complications and correlated with adverse outcomes. On the other hand, some researchers have shown no association between inherited thrombophilia and recurrent pregnancy loss [⁴¹–⁴⁵], possibly because either the only evaluated SNP was MTHFR C677T [⁴³, ⁴⁴] or the sample sizes were small, that is, 40 patients and 20 controls [⁴²] and 55 women with recurrent pregnancy loss and 50 controls [⁴¹]. Additionally, in a Malasyan population, the identification of FV Leiden and FII G20210A in women with recurrent pregnancy loss was also low [⁴⁶].

Our results are in contrast with previous recurrent pregnancy loss studies in Latin American populations. Brazilian women carrying FV Leiden had a 4.9 times higher risk of recurrent pregnancy loss than noncarriers [⁹], and women heterozygous for FV Leiden from Uruguay had a 5 times higher risk of recurrent pregnancy loss than noncarriers (CI 95%, 1.5–21) [⁴⁷]. Therefore, our study highlights special genetic characteristics in this triethnic population from Colombia compared to populations with a higher percentage of Caucasians [²³, ²⁴]. Our results confirm the differences in the genetic background between Caucasian and non-Caucasian populations, in one extreme populations such as the Japanese that virtually lacks FV Leiden carriers [⁴⁸] and somewhere in the spectrum of thrombophilia frequency, our triethnic population with low frequency but identifiable thrombophilia-associated SNPs. These results provide evidence of how a genetic component such as polymorphisms in genes associated with thrombosis and its interaction with diverse environmental factors such as food, geographical location, medical resources, public health, and the age of the woman could lead to variable manifestations of complex diseases such as recurrent pregnancy loss.

In the current study, the wide confidence intervals suggest that the estimated sample size of 300 individuals was insufficient to detect differences between patients with recurrent pregnancy loss and control women for whom the power of the study decreased from an estimated 80% to 30% for detections of an OR of 2.0 with a frequency of 0.5% for FV Leiden in the controls. Nevertheless, our results are valuable because they confirm that the frequency of these inherited thrombophilias in our populations is low; therefore, we believe that there is no justification to recommend testing inherited thrombophilias in the initial evaluation of patients with recurrent pregnancy loss. To detect an association between thrombophilias and recurrent pregnancy loss, it would be necessary to increase considerably the sample size but with unlikely clinical significance.

These authors contributed equally to this study.

This work was supported by Grant 1115-04-11916 from Colciencias and Universidad de Antioquia.