Patterns of coagulation profiles observed in different trimesters of pregnancy

INTRODUCTION

The coagulation system undergoes significant changes during pregnancy; an increase in levels of some of the clotting factors, decreased concentrations of some of the natural anticoagulants and diminishing fibrinolytic activity, thus producing a state of hypercoagulability.^1-3 This phenomenon, supposedly due to hormonal changes, helps in maintaining placental function during pregnancy, and protects from fatal hemorrhage during delivery.^4,8

The risk of venous thromboembolism (VTE) is high during pregnancy due to both physiologic changes of pregnancy and the additional impact of the inherited and acquired thrombophilias.^2,5,6 The overall rate of venous thromboembolic events in pregnancy is 200 per 100,000 deliveries.^2,7,8 The main risk appears to occur in the postpartum period where the incidence increases about 3-fold and is estimated at 500 per 100,000.^2,7 These events are deep vein thrombosis as opposed to the more serious pulmonary embolism.^2,10

Inherited and acquired thrombophilia contribute further to an increased predisposition to thrombotic events.^2,9 The overall impact of inherited and acquired thrombophilia is low in the non-pregnant population, and the majority of patients never experience a thrombotic event.^2,8-10 During pregnancy, however, the increased risk of thrombosis in patients with inherited and acquired thrombophilia can be substantial and warrants consideration, especially as thrombosis is one of the leading causes of mortality during pregnancy.^2,7,9

Positive efforts have been made in reducing maternal death from postpartum hemorrhage (PPH) over years.⁹ Other causes of maternal death are frequently undiagnosed or under-diagnosed.⁷ Most of these deaths result from thrombo-embolic disease in pregnancy.^7,13,11 Approximately 50% of women with venous thrombo-embolic (VTE) disorders detected during pregnancy or puerperium have an underlying hereditary thrombophilia in addition to the acquired risk.⁶ Several investigators have reported an association between thrombophilia and adverse pregnancy outcomes caused by uteroplacental thrombosis. Other groups, however, have failed to confirm this association.^7,8,16,12

Protein S (PS) deficiency and protein C deficiency (PC) have been found in the general population with pregnant women not excluded and antithrombin III (ATIII) deficiency is associated with recurrent miscarriages.^5,17,13

Several reports have regarded coagulation profiles as markers of thromboembolic disorder in pregnancy.^10,14,15 A study done in Kano Nigeria showed that platelet count, and factor VIII concentrations are increased during pregnancy with significant reduction of protein C.³ However, there is paucity of information on the level of natural coagulation inhibitors during pregnancy from this environment. This research work aimed at determining the level of natural coagulation inhibitors in pregnant women in the three trimesters of pregnancy.

METHODS

Study area

This study was conducted at the ante-natal clinic, booking clinic and labor ward of Ladoke Akintola University of Technology Teaching Hospital, Ogbomoso, Oyo State, Nigeria.

The hospital serves as a research and training institution for undergraduate and postgraduate medical programs as well as a major referral centre. The strategic location of the teaching hospital in Ogbomoso enables it to serve the people of Oke-Ogun, Oyo town and its environs, Ogbomoso and its environs, some parts of Osun as well as Kwara state with qualitative health care delivery.

Study design

The study was a comparative cross-sectional hospital-based study. Pregnant women between ages of 15-49 years with serum or ultrasound confirmation of pregnancy were included in the study after obtaining informed consent.

Sample size

The sample size for patients was calculated using the statistical formula comparative study.¹⁶ The minimum sample size derived was 24.55. To enhance the power of this study, a total sample size of 75 was considered for both subjects and controls. Therefore, a total sample size of 150 was used. From this, 75 normal pregnant women were subdivided into three groups; 25 in their first trimester, 25 in their second trimester, 25 in their third trimester and 75 age-matched non-pregnant women as control.

Procedure

A semi-structured questionnaire was used as the study instrument to obtain information on socio-demographic characteristics and clinical history. After obtaining informed consent, 5mls of venous blood was collected from each subject into EDTA bottle. The specimens were centrifuged, the plasma was separated into aliquot bottles and stored at a temperature of -30⁰C for analysis of protein C, protein S, antithrombin III assays. The enzyme-linked immunosorbent assay (ELISA) method was used for the quantitative determination of protein C, protein S antigens and human antithrombin III.

Data analysis

The statistical analysis was carried out using Statistical Package for Social Science (SPSS) version 24. A multivariate analysis was done. Mean and standard deviation were used to report baseline continuous variable while frequency and proportions were used to report categorical variables. Pearson’s chi-squared, student T-test were used to compare variables as appropriate.

RESULTS

One hundred and fifty (150) questionnaires were administered in all; seventy-five for subjects which were subdivided into twenty-five pregnant women each in their first trimester, second trimester and third trimester respectively (twenty-two of the subjects were primigravida) and seventy-five controls who were equally age matched healthy non-pregnant women.

Table 1 shows the demographic profile of the participants. The mean ages of the subjects and control group were 32.6±4.6 and 34.5±6.9 years respectively. The majority of participants in both groups were married (control: 84%, cases: 98.7%).

t: t-statistic, p value < 0.05 indicates significance.

Table 2 shows that m the majority of participants in the subjects and control group have had a history of two to three pregnancies before (study: 56%, control: 58.7%) respectively and 37.7% of the subjects and 34.5% of the control group had previous miscarriages respectively. Two (3.8%) of the subjects and none of the control group had a history of intrauterine fetal death (p=0.155). There was no significant difference in the history of intrauterine growth restriction between the subjects and the control group (1.9 versus 3.7%; p=0.560).

Table 3 depicts that there was no significant difference in family history of deep venous thrombosis, pulmonary embolism, bleeding disorder and sudden death during pregnancy between the two groups (p value >0.05).

There was no significant difference in the mean protein C between the subjects in the first trimester and the control group, p value =0.4890. Antithrombin III was significantly higher among subjects in the first trimester than the controls (603.52±56.43 ng/ml and 554.16±54.65 ng/mL respectively), p value =0.0004. Similarly, protein S was significantly higher among subjects in the first trimester than the controls (5.77±0.69 ng/ml and 4.78±0.65 ng/ml respectively), p value <0.0001. However, plasma level of natural coagulation inhibitors among subjects in the second trimester did not differ significantly from controls. Antithrombin III was significantly lower among subjects in the third trimester than the controls (530.00±48.13 ng/ml and 554.16±54.65 ng/ml respectively), p value =0.0413. There was also a downward trend in the natural coagulation inhibitors as pregnancy advanced in this study.

Table 5 shows that protein C, protein S and antithrombin III all declined with gestational age. The mean protein C decreased from the first to the third trimester (first: 1094.1±276.7 pg/ml, second: 1064±241.2 pg/ml, third: 1043.8±218.0 pg/ml). These differences were not significant (p=0.769). The mean protein S among subjects in the first trimester was 5.7±0.7 ng/ml and 4.8±0.7 ng/ml in both the second and third trimesters. These differences were statistically significant (p=0.001). The mean antithrombin III decreased from the first trimester to the third trimester (first: 603.5±56.4 ng/ml, second: 570.8±57.6 ng/ml, third: 530.0±48.1 ng/ml). These differences were statistically significant (p=0.001).

*F: F statistic, p value < 0.05 indicates significance.

DISCUSSION

The mean age of the pregnant women was 32.6±4.6 years while the mean age of participants in the control group was 34.5±6.9 years and the majority of the participants were aged between 30-34 years. This finding is similar to the age range found in the work done by Oladosu et al who reported a mean age of 29.9±5.2 years.¹⁷ Okwesili et al however reported a lower mean age (25.52±4.92 years) in a study done in Sokoto Nigeria, this may be a result of the practice of early marriage among women in the northern part of Nigeria. Age is an important factor in the likelihood of occurrence of thrombophilia and pregnancy complications; the older age group is associated with higher risk.

Most of the participants (70.7%) in this study were Christians which reflected the dominance of Christians in Ogbomoso. 98.7% of the participants in the study group were married while 84.0% in the control group were married. This finding is similar to a study done by Okeyinka Y who reported that 78.2% of women in Ogbomoso were married.¹⁸

As expected, the dominant ethnic group of participants (study group-93.3%, control group 100%) was Yoruba since this study was conducted in the South Western part of Nigeria, a Yoruba-dominated population. This study revealed that the majority of the participants in both groups (94.7%-control group, 85.3%-study group) had a tertiary level of education. This may reflect the fact that more highly educated people patronized the hospital being a teaching hospital than non-educated individuals.

The menstrual cycles of all the subjects and controls were within the normal range and likewise, the duration of bleeding for subjects and controls was within the normal range (100%). This might be because both the subjects and control group were apparently normal without any morbidity as stated in the inclusion criteria.

Approximately 38% and 35% of the study group and control respectively had a history of miscarriages.

The plasma level of protein C, protein S, and antithrombin III for both groups were within the normal reference range but slightly higher among the pregnant women than the control subjects. However, these differences were not statistically significant. This study revealed that there is just a slight increase of the NCI in normal pregnancy to combat the hypercoagulable state of pregnancy thereby preventing the occurrence of thromboembolic events associated with pregnancy. There was also a downward trend of the NCI as the gestation period increased which corroborated that pregnancy is a hypercoagulable state with the likelihood of consumption of NCI as pregnancy advanced. A study done in Turkey by Cengiz et al had a similar report.¹² Also, Nwagha et al reported a decrease in protein C from the second to the third trimester.⁷

However, Imoru et al have a contradictory report with protein C increasing significantly across the trimesters while antithrombin III fluctuated across the trimesters when compared with control.³ This might be because of the small sample size that was studied (n=10). It was also noted in a study done by Buseri et al that protein C was significantly lower when compared with the control while antithrombin III showed no significant difference however the study group was not divided into trimesters, which might account for the contrary opinion.¹⁹

The mean antithrombin III and protein C of the pregnant women decreased across the gestation period from the first trimester to the third trimester with statistical significance only in antithrombin III (p=0.001). This is in keeping with the hypercoagulable state of pregnancy and increased consumption of the NCI as the pregnancy progressed to the third trimester.^2,5,6 Nwagha et al reported the same pattern with protein C.⁷ Other reports by Imoru et al and Ajayi et al showed different patterns and fluctuated values for protein C and antithrombin III at different gestation periods with no statistical significance.^3,19

The mean protein S of the normal pregnant women decreased significantly from the first trimester to the second trimester and maintained at that level throughout the pregnancy. Domenico et al and Feroza et al reported a progressive fall of total protein S with increasing gestation age.^11,20 Variation of protein S by various authors during pregnancy could be associated with different types of reagents used, techniques employed and sample sizes considered.

CONCLUSION

Pregnancy is a hypercoagulable state regulated by natural coagulation inhibitors (NCI) which were higher among subjects in this study and were also noted to decrease as pregnancy advanced. It is therefore recommended that all pregnant women with high-risk factors of thromboembolic events should have a plasma level of prothrombin time, and activated partial thromboplastin time done as baseline investigations to identify the disorders early enough. Such pregnant women with deranged coagulation profiles could benefit from anticoagulation either prophylaxis or therapeutic and further clinical evaluations.

Funding: No funding sources

Conflict of interest: None declared

Ethical approval: The study was approved by the Institutional Ethics Committee of the LAUTECH Teaching Hospital (LTH) Ogbomosho, Oyo State, Nigeria

REFERENCES

1. 1. Hoffbrand AV PAHM. Essential Heamatology. 6th Ed. Wiley-blackwell; 2011:345-380,413-423.
2. 2. Battinelli EM, Marshall A, Connors JM. The role of thrombophilia in pregnancy. Thrombosis. 2013;2013:1-9.
3. 3. Olutayo IM, Ajayi I. Haemostatic changes during pregnancy and puerperium in kano, north-western Nigeria. J Hematol Thromb Dis. 2015;03(05).
4. 4. Victor Hoffbrand A, Catovsky D, Edward GD. Tuddenham ARG. Postgraduate Haematology. 6th ed. Wiley-blackwell;2011:746-928.
5. 5. Sibai BM, How HY, Stella CL. Thrombophilia in pregnancy: Whom to screen, when to treat. Obg Manag. 2007;19(1):50-64.
6. 6. Kaushanky K, Lichtman MA, Beutler E, Thomas J. Kipps, Uri Seligsohn JTP. Williams Haematology. 8th ed. McGraw-Hill; 2010:1721-2219.
7. 7. Nwagha UT, Nwagha UI, Ibegbulam OG, Ocheni S, Okpala I. Increased prevalence of activated protein c resistance during pregnancy may implicate venous thrombo embolic disorders as a common cause of maternal mortality in Nigeria. J Bas Clin Reproduct Sci. 2012;1(1):19-24.
8. 8. Walker ID, Infirmary R. Thrombophilia in pregnancy. J Clin Pathol. 2000;53(8):573-80.
9. 9. Dawood F. Pregnancy and Thrombophilia. J Blood Disord Transfus. 2013;4(5):1-11.
10. 10. Avwioro GO. Prothrombin time and activated partial thromboplastin time in pregnant women in southern Nigeria. J Appl Pharm Sci. 2013;3(April):179-81.
11. 11. Babker AM, Gameel FE. Molecular characterization of prothrombin G20210A gene mutations In pregnant Sudanese women with spontaneous recurrent abortions. Rawal Med J. 2015;40(2):207-9.
12. 12. Demir C, Dilek I. Natural coagulation inhibitors and active protein c resistance in preeclampsia. Clinics. 2010;65(11):1119-22.
13. 13. Markoff A, Bogdanova N, Samama MM. Hereditary thrombophilic risk factors for recurrent pregnancy loss. Hereditary Genetics. 2012;1:2161-1041.
14. 14. Katz D, Beilin Y. Disorders of coagulation in pregnancy. Br J Anaesth. 2015;115(suppl 2):75-88.
15. 15. Okwesili A, Ibrahim K, Nnadi DC, Barnabas B. Fibrinogen levels among pregnant women of african descent in sokoto north Western Nigeria. Front Biomed Sci. 2016;1(September):7–11.
16. 16. Charan J, Biswas T. How to calculate sample size for different study designs in medical research?. Ind J Psychol Medi. 2013;35(2):121-6.
17. 17. Oladosu-olayiwola O, Olawumi H, Babatunde A, Ijaiya M. Fibrinolytic proteins of normal pregnancy and pre-eclamptic patients in North West Nigeria. Afr Health Sci. 2011;18(3):576-83.
18. 18. Ronke O. Socio economic Characteristics of Residents of Multihabited Houses in Ogbomoso, South West, Nigeria. Glob J Human-Soci Sci Res. 2016;16(6):1-9.
19. 19. Imoru Momodu FIB. Protein c and Antithrombin III Activities in Healthy Nigerian Women. Inter J Haematol Res. 2015;1(1):20-3.
20. 20. Domenico Prisco, Gabriele Cuiti MF. Hemostatic changes in normal pregnancy. Haematol Repor. 2005;1(10):1-5.