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Year : 2011  |  Volume : 54  |  Issue : 4  |  Page : 752-755
Coagulation inhibitors and activated protein C resistance in recurrent pregnancy losses in Indian women

1 Department of Pathology and Obstetrics, Lady Hardinge Medical College and Associated Hospitals, (Opposite Shivaji Stadium), New Delhi, India
2 Department of Gynaecology, Lady Hardinge Medical College and Associated Hospitals, (Opposite Shivaji Stadium), New Delhi, India

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Date of Web Publication6-Jan-2012


Background: Thrombophilias, both acquired and inherited, have been investigated in the etiopathogenesis of unexplained recurrent pregnancy loss. Aim: To study coagulation inhibitors and activated protein C resistance (APCR) in recurrent pregnancy losses (RPL) occurring in second and third trimesters. Materials and Methods: A total of 30 pregnant women (group A) with two or more recurrent unexplained fetal loses were evaluated for APCR, protein C deficiency, protein S deficiency, antithrombin deficiency, and antiphospholipid antibodies (APLA). Thirty age-matched controls were taken (group B) comprising of pregnant women with at least one live issue. Statistical Analysis: Comparisons between two group frequencies and group means were made using Chi square test and Student's t test, respectively. Results: Protein C and protein S levels were reduced in group A compared with group B and the difference was statistically significant (P=0.005 and P=0.032, respectively). The mean value of antithrombin was slightly reduced in group A compared with group B. APCR was observed in 16.6% cases and 3.3% controls. However, the difference was not statistically significant. APLA was observed in 20% cases and none of the controls. Of these, lupus anticoagulant was positive in 16.6% cases and anticardiolipin antibodies in 10% cases. Combined defects were seen in seven patients. Conclusion: There is a significant risk of RPL in pregnant women with thrombophilias. Therefore, screening for thrombophilias may be justified in pregnant women with unexplained recurrent fetal wastage, especially in second and third trimester.

Keywords: Antiphospholipid antibodies, activated protein C resistance, recurrent pregnancy loss, recurrent abortions, thrombophilias

How to cite this article:
Jyotsna P L, Sharma S, Trivedi SS. Coagulation inhibitors and activated protein C resistance in recurrent pregnancy losses in Indian women. Indian J Pathol Microbiol 2011;54:752-5

How to cite this URL:
Jyotsna P L, Sharma S, Trivedi SS. Coagulation inhibitors and activated protein C resistance in recurrent pregnancy losses in Indian women. Indian J Pathol Microbiol [serial online] 2011 [cited 2023 Feb 1];54:752-5. Available from:

   Introduction Top

Recurrent pregnancy loss is a heterogeneous condition with several etiological factors. Nearly 70% of spontaneous abortions in the first 12 weeks are due to chromosomal anomalies. However, losses due to cervical incompetence and hematologic disorders are common after twelve weeks. [1]

Pregnancy is a hypercoagulable state and women with thrombophilias are at increased risk for thrombosis during pregnancy and adverse maternal and fetal sequelae. The hemostatic system plays an important role in the success of pregnancy and the process of implantation and placentation. Implantation of the fertilized egg into the uterine decidua establishes a contact between the fetus, the placenta, and the maternal circulation. This contact is crucial for the success of pregnancy. Prothrombotic changes and thrombosis may interfere with these processes leading to miscarriage.

Severe pregnancy complications such as severe preeclampsia, intrauterine growth retardation (IUGR), abruptio placentae, and still-birth have been shown to be associated with thrombophilias, both acquired and inherited. [2],[3],[4],[5],[6],[7],[8],[9],[10],[11] The present study was conducted to study natural and acquired coagulation inhibitors and activated protein C resistance (APCR) in recurrent pregnancy losses (RPL).

   Materials and Methods Top

The study group comprised of thirty pregnant women, with a history of two/more unexplained fetal losses (spontaneous abortions and/or fetal deaths) occurring after the first trimester. The control group comprised of 30 age-matched pregnant women with two or more live issues without any prior history of abortions and/or fetal deaths. A detailed clinical history was taken and physical examination conducted. Patients with any apparent cause of fetal wastage, like genital tract/fetal malformations, infectious causes, ABO and Rh incompatibility, and diabetes mellitus were ruled out using relevant tests like ultrasonography, Venereal Disease Research Laboratory (VDRL) test, TORCH test (Toxoplasma, other infections, Rubella, Cytomegalovirus, Herpes simplex), blood group, and blood glucose tests.

Complete blood counts, screening coagulation tests like Prothrombin Time(PT) and Activated Partial Thromboplastin Time (APTT), tests for natural and acquired coagulation inhibitors, i.e., protein C, protein S, anti-thrombin, antiphospholipid antibodies (APLA), and APCR were performed. Platelet-poor plasma was prepared by centrifuging the blood samples at 2 500 g for 15 to 20 minutes. Estimation of protein C, protein S, APCR, and lupus anticoagulant (LA) was performed using clot-based assays, while the semiquantitative determination of APLA classes (IgG/IgM) was done using ELISA kit and antithrombin was measured using chromogenic assay (DIAGNOSTICA STAGO, France). In all cases showing LA positivity and/or high APLA values, the test was repeated after an interval of 12 weeks.

Statistical analysis was performed using SPSS software. Comparisons between two group frequencies were made using Chi square test. For the comparison of group means, Student's t test was applied.

   Results Top

The mean age of group A was 24.9 years and that of group B was observed to be 25 years. Most of the subjects in group A had at least two recurrent second trimester abortions and/or stillbirths. The subjects in group B had no pregnancy loss. There was no significant difference between the 2 groups with respect to the routine hematological parameters.

PT and APTT values did not show significant difference between the two groups.

Protein C, Protein S, and Antithrombin

The mean value of protein C in group A (69.95 ± 18.33%) when compared with group B (81.33 ± 9.90%) was found to be significantly low (P=0.005). Protein C levels were lowered in 33.3% of group A subjects as compared with 3.3% in group B subjects. The mean value of protein S in group A (74.43 ± 19.8%) was significantly lower than that of group B (83.05 ± 7.44%) (P=0.032). The protein S levels were lowered in 23.3% of group A subjects, while the corresponding values in group B fell within reference range. The mean value of antithrombin in group A (90.4 ± 15.39%) was slightly lower than that of group B (96.9 ± 8.79%). However, the difference was statistically significant. This could be because the single subject of group A showed antithrombin deficiency had a very low level (20% of PNP) [Table 1].
Table 1: Comparison of natural coagulation inhibitors

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Activated Protein C Resistance

APCR was observed in 5 of 30 (16.6%) subjects in group A, while 1 of 30 (3.3%) subjects in group B showed APCR. Although APCR was present more in patients than in controls, the difference in the group frequencies compared using the Chi-square test was not found to be statistically significant (P=0.195).

Antiphospholipid Antibodies

APLA were positive in 6/30 cases in group A (20%) and none in controls. The difference was statistically significant ( P=0.024). Of these, LA was found in 5/30 cases (16.6%), while high titers of anticardiolipin antibodies (aCL) were observed in 2/30 subjects (10%) in group A. One of these patients concomitantly had both LA and high titers of aCL. All patients with APLA were tested again after an interval of 12 weeks and showed persistent positivity.

Combined Defects

Combined defects were seen in seven patients. Four patients showed combined deficiencies of protein C and protein S. One patient each showed protein C deficiency with APCR, APCR with APLA, and protein C deficiency with APLA, respectively.

   Discussion Top

Maternal thrombophilia has recently been identified as a major cause of adverse pregnancy outcome, including recurrent pregnancy loss, still-birth, severe pre-eclampsia, placental abruption, and intrauterine growth restriction. [2],[3],[4],[5],[6],[7],[8],[9],[10],[11]

The most common inherited disorders are deficiencies of antithrombin, protein C, protein S, APCR due to factor V Leiden(FVL) mutation, a function-enhancing mutation of the prothrombin gene (G20210A), and thermolabile mutation for methylene-tetrahydrofolate reductase C677T. The antiphospholipid antibody syndrome (APS) is the most common cause of acquired thrombophilias. [4] It is associated with complications spanning all trimesters of pregnancy. [12],[13],[14]

The results obtained in this study for protein C and S were comparable with those by de Vries et al.[15] who demonstrated deficiencies of protein C and protein S in patients with complicated pregnancies (intrauterine fetal death, small-for-gestational age, and/or placental abruption). Gris et al.[16] also found an increased prevalence of protein C and protein S deficiency in patients with at least one late still-birth. However, both the above series were unable to identify antithrombin deficiency.

Preston et al.[17] found a significant increase in miscarriages and still-births in patients with protein C, protein S, and antithrombin deficiency. Sanson et al.[18] also found a greater risk for fetal loss after 8 weeks in patients with these deficiencies. However, other studies on women with RPL could not find an increased incidence of protein C/protein S deficiency. [19]

There are conflicting reports on the association of APCR with RPL. APCR can be inherited or acquired. Inherited APCR is most commonly due to factor V Leiden mutation or other factor V polymorphisms. It is said to be the most common inherited form of thrombophilia. First described in 1993, Ridker et al.[20] did a survey on the ethnic distribution of factor V Leiden mutation in 4 047 American men and women. Their data indicated a higher prevalence of factor V Leiden among Caucasians (5.27%) than minority Americans (0.45% in Asian Americans and 1.23% in African Americans).

Preston et al.[17] found a modest increase of still-births, but not of miscarriage in women with factor V Leiden mutation. Rai et al. [21] found a significantly higher prevalence of APCR in women with recurrent second trimester miscarriage, thereby suggesting APCR as a mechanism of fetal loss in the second trimester. Dizon-Townson et al.[22] found no association between factor V Leiden and RPL in their studies.

Rai et al.,[21] on the other hand, demonstrated acquired APCR, but not factor V Leiden was more common in early/late RPL, thereby concluding that acquired but not congenital APCR is associated with RPL. In the present study, APCR was found in 5/30 (16.6%) cases as compared with 1/30 (3.3%) controls and the difference was not statistically significant ( P = 0.195). APCR (due to factor V Leiden) may not be an important cause of RPL in Asian/Indian population as demonstrated by Kazumasa [23] and Biswas et al.[24] The APCR positivity obtained in the present study may be due to acquired APCR. Vora et al.[25] found that the risk of fetal loss with protein S deficiency was the highest risk observed for any heritable thrombophilia, followed by protein C, factor V Leiden, endothelial protein C receptor, antithrombin III deficiency, and beta448 fibrinogen polymorphism, in a study on Indian patients. Ayadurai et al.[26] found that thrombophilia was identified in more than one-fourth of the RPL subjects. APCR not caused by factor V Leiden mutation was the most common thrombophilia marker in Malaysians, whereas in Caucasians, it was APCR due to FVL mutation. Larger studies are required to determine the role of APCR in Indian patients with RPL.

Recurrent pregnancy loss is one of the defining diagnostic criteria of APS. APS is the most common cause of acquired thrombophilias. It is associated with complications that span all trimesters of pregnancy, including RPL, severe pre-eclampsia, IUGR, and placental abruption. Various studies have shown LA positivity in RPL ranging from 9 to 17%, while the frequency of aCL in RPL ranged from 11 to 42%. [27],[28],[29],[30] The results obtained in the present study were similar to those by Narayan et al.[31]

In conclusion, this study suggests that a significant number of Indian patients with RPL show thrombophilic defects. Hence, all patients with unexplained RPL should be investigated for thrombophilic defects. More prospective studies are required to establish the causal link between thrombophilia and adverse pregnancy outcome and assess the effectiveness of thromboprophylaxis in pregnant women with RPL.

   References Top

1.Hatasaka HH. Recurrent miscarriage: Epidemiologic factors, definitions, and incidence. Clin Obstet Gynecol 1994;37:625-34.   Back to cited text no. 1
2.Gerhard A, Scharf RE, Beckmann MW, Struve S, Bender HG, Pillny M, et al. Prothrombin and factor V mutations in women with a history of thrombosis during pregnancy and the puerperium. N Engl J Med 2000;342:374-80.  Back to cited text no. 2
3.Greer IA. The challenge of thrombophilia in maternal-fetal medicine. N Engl J Med 2000;342:424-5.  Back to cited text no. 3
4.Girling J, de Swiet M. Inherited thrombophilia and pregnancy. Curr Opin Obtet Gynecol 1998;10:135-44.  Back to cited text no. 4
5.Arias F, Romero R, Joist H, Kraus FT. Thrombophilia: A mechanism of disease in women with adverse pregnancy outcome and thrombotic lesions in placenta. J Maten Fetal Med 1998;7:277-86.  Back to cited text no. 5
6.Ray JG, Laskin CA. Folic acid and homocyst(e)ine metabolic defects and the risk of placental abruption, pre-eclampsia and spontaneous pregnancy loss: A systematic review. Placenta 1999;20:519-29.  Back to cited text no. 6
7.Van Pampus MG, Dekker GA, Wolf H, Huijgens PC, Koopman MM, von Blomberg BM, et al. High prevalence of hemostatic abnormalities in women with a history of severe preeclampsia. Am J Obstet Gynecol 1999;180:1146-50.  Back to cited text no. 7
8.Wang J, Trudinger BJ, Duarte N, Wilcken DE, LiWang X. Elevated circulating homocysteine levels in placental vascular disease and associated pre-eclampsia. Br J Obstet Gynaecol 2000;107:935-8.  Back to cited text no. 8
9.Kahn SR. Severe preeclampsia associated with coinheritance of factor V Leiden mutation andprotein S deficiency. Obstet Gynecol 1998;91:812-4.  Back to cited text no. 9
10.Van der Molen EF, Verbruuggen B, Novakova I, Eskes TK, Monnens LA, Blom HJ. Hyperhomocysteinemia and other thrombotic risk factors in women with placental vasculopathy. Br J Obstet Gynaecol 2000;107:785-91.  Back to cited text no. 10
11.Goddijn-Wessel TA, Wouters MG, van de Molen EF, Spuijbreek MD, Steegers-Theunissen RP, Blom HJ, et al. Hyperhomocysteinemia: A risk factor for placental abruption or infarction. Eur J Obstet Gynecol Reprod Biol 1996;66:23-9.  Back to cited text no. 11
12.Rai R, Regan L. Antiphospholipid syndrome and pregnancy loss. Hosp Med 1998;59:637-9.  Back to cited text no. 12
13.Backos M, Rai R, Baxter N, Chilcott IT, Cohen H, Regan L. Pregnancy complications in women with recurrent miscarriages associated with antiphospholipid antibodies treated with low dose aspirin and heparin. Br J Obstet Gynaecol 1999;106:102-7.  Back to cited text no. 13
14.Kupferminc MJ, Eldor A, Steinman N, Many A, Bar-Am A, Jaffa A, et al. Increased frequency of genetic thrombophilia in women with complications of pregnancy. N Engl J Med 1999;340:9-13.  Back to cited text no. 14
15.De Vries JIP, Dekker GA, Huijgens PC, Jakobs C Blomberg BME, et al. Hyperhomocysteinaemia and protein S deficiency in complicated pregnancies. Br J Obstet Gynecol 1997; 104:1248-1254.  Back to cited text no. 15
16.Gris JC, Quere I, Monpeyroux F, Mercier E, Ripart-Neveu S, Tailland ML, et al. Case-control study of the frequency of thrombophilic disorders in couples with late fetal loss and no thrombotic antecedent. Thromb Haemost 1999;81:891-9.  Back to cited text no. 16
17.Preston FE, Rosendaal FR, Walker ID, Briet E, Berntorp E, Conard J, et al. Increased fetal loss in women with heritable thrombophilia. Lancet 1996;348:913-16.  Back to cited text no. 17
18.Sanson BJ, Friederich PW, Simioni P, Zanardi S, Hilsman MV, Girolami A, et al. The risk of abortion and stillbirth in antithrombin-, protein C -, and protein S-deficient women. Thromb Haemost 1996,75:387-88.  Back to cited text no. 18
19.Yamada H, Kato EH, Kobashi G, Ebina Y, Shimada S, Morikawa M, et al. Recurrent pregnancy loss: Etiology of thrombophilia. Semin Thromb Haemost 2001;27:121-9.  Back to cited text no. 19
20.Ridker PM, Miletich JP, Hennekens CH, Buring JE. Ethnic distribution of factor V Leiden in 4047 men and women. Implications for venous thromboembolism screening. JAMA 1997;277:1305-7.  Back to cited text no. 20
21.Rai R, Shlebak A, Cohen H, Backos M, Holmes Z, Marriott K, et al. Factor V Leiden and acquired activated protein C resistance among 1000 women with recurrent miscarriage. Hum Reprod 2001;16:961-5.  Back to cited text no. 21
22.Dizon-Townson DS, Kinney S, Branch DW, Ward K. The factor V Leiden mutation is not a common cause of recurrent miscarriage. J Reprod Immunol 1997;34:217-23.  Back to cited text no. 22
23.Kazumasa H. The factor V Leiden mutation in Japanese couples with recurrent spontaneous abortion. Hum Reprod 1999;14:1872-4.  Back to cited text no. 23
24.Biswas A, Choudhry P, Mittal A, Meena A, Ranjan R, Choudhry VP, et al. Recurrent abortions in Asian Indians: No role of factor V Leiden Hong Kong/Cambridge mutation and MTHFR polymorphism. Clin Appl Thromb Hemost 2008;14:102-4.  Back to cited text no. 24
25.Vora S, Shetty S, Salvi V, Satoskar P, Ghosh K. Thrombophilia and unexplained pregnancy loss in Indian patients. Natl Med J India 2008;21:116-9.  Back to cited text no. 25
26.Ayadurai T, Muniandy S, Omar S. Thrombophilia investigation in Malaysian women with recurrent pregnancy loss. J Obstet Gynaecol Res 2009;35:1061-8.  Back to cited text no. 26
27.Miyakis S, Lockshin MD, Atsumi T, Branch DW, Brey RL, Cervera R, et al. International consensus statement on an update of the classification criteria for definite antiphospholipid syndrome(APS). J Thromb Haemost 2006;4:295-306.  Back to cited text no. 27
28.Petri M, Golbus M, Anderson R, Whiting-O'Keefe Q, Corash L, Hellmann D. Antinuclear antibody, lupus anticoagulant, and anticardiolipin antibody in women with idiopathic habitual abortion. A controlled, prospective study of forty-four women. Arthritis Rheum 1987;30:601-6.  Back to cited text no. 28
29.Barbui T, Cortelazzo S, Galli M, Parazzini F, Radici E, Rossi E, et al. Antiphospholipid antibodies in early repeated abortions: A case-controlled study. Fertil Steril 1988;50:589-92.  Back to cited text no. 29
30.Stephenson MD. Frequency of factors associated with habitual abortion in 197 couples. Fertil Steril 1996;66:24-9.  Back to cited text no. 30
31.Narayan S, Bhattacharya G, Sharma S, Baliga S. A study of Lupus anticoagulant in unexplained fetal wastage. Indian J Pathol Microbiol 2000;43:123-6.  Back to cited text no. 31
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Sunita Sharma
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DOI: 10.4103/0377-4929.91507

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