Correlation of Biochemical Parameters in Paired Maternal and Umbilical Cord Sera in Pregnancies Complicated with Severe Anaemia

Objective: To evaluate the biochemical profile of patients with severe anemia in pregnancy and to study relation between maternal and umbilical cord serum levels of iron, ferritin, cobalamin and folate. Methods: It is a prospective case control study with 200 pregnant women with severe anemia (Hb< 7 gm/dl) and pregnant women with no anemia (Hb>11gm/dl were recruited. Serum ferritin, folate and cobalamin were measured by electro-chemiluminescence method. Cord blood samples were collected for estimating the levels of iron, ferritin, folate and cobalamin. Results: Cases were divided into three groups, microcytic anemia (47%), macrocytic anemia (37.5%) and dimorphic anemia (15.5%). Mothers with iron deficiency were 37.5%, whereas cobalamin and folate deficiency were seen in 47% and 11% respectively. In women with microcytic anemia, iron and ferritin levels in umbilical cord blood were 5.54 and 19.41 times higher, respectively, than maternal serum values. However, in macrocytic anemia, cobalamin levels were only 1.22 times more in cord blood as compared to maternal serum values. Conclusion: Cord blood values show that fetus is able to compensate for low levels of maternal iron and folate but not for cobalamin, indicating the absence of preferential transfer of cobalamin to the fetus. INTRODUCTION Anemia in pregnancy has been almost synonymous with irondeficiency anemia but there appears to be an emerging evidence of macrocytic anemia. Most of the previous studies have reported a lower incidence of macrocytic anemia and hence its importance in pregnancy has been neglected [1,2]. A study evaluating the clinical profile of patients having severe anemia in pregnancy was published earlier where macrocytic picture was seen in 40% of cases of severe anaemia [3]. It also showed that the maternal and fetal morbidity and mortality appears to be higher in macrocytic anemia as compared to iron-deficiency anemia. The prior study mainly dealt with clinical profile and therefore this study was planned to focus on biochemical characteristics and relationship between maternal and umbilical cord hematological parameters in women with severe anemia with reference to type of anemia [3]. The reason to conduct such a study was to assess the differential transfer of nutrients in iron deficiency and megaloblastic anemia; so as to know the impact of type of anemia on fetal development. There is paucity of literature addressing this issue. It was also done to evaluate the effect of various types of anemia on the fetus as this may subsequently have long term consequences. MATERIAL AND METHODS This was a prospective longitudinalstudyinvolving400 pregnant woman (Test=200 and Control = 200) from August, 2010 to February, 2012 in the Department of Obstetrics and Gynaecology of a tertiary care hospital in Northern India. Ethical clearance was taken from Institutional Ethical Committee. The study included 200 pregnant women with severe anemia (Hb< 7gm/dl) who were admitted to the maternity ward as test group together with 200 pregnant women without anemia (Hb>11gm/ dl) attending antenatal clinic who served as controls after written informed consent. Only women with singleton pregnancies presenting after 34 weeks of gestation were recruited. Women with twin pregnancies or who were admitted and treated before 34 weeks were not recruited in the study. Central Bringing Excellence in Open Access   Singh et al. (2018) Email: J Hematol Transfus 6(2): 1081 (2018) 2/11 All women underwent a detailed history followed by thorough general physical, systemic and obstetric examination. Blood samples of all subjects were collected before commencement of any treatment and analyzed for routine antenatal investigations (blood grouping and Rh typing, hemoglobin estimation, antibodies against human immunodeficiency virus, Australia antigen, standard test for syphilis, glucose challenge test, urine routine and microscopy, urine culture and sensitivity and Naked Eye Single Tube Red Cell Osmotic Fragility test (NESTROFT) [4]. At enrolment, approximately 4 ml venous blood was taken from each patient and divided into 2 parts. 1 ml of blood was transferred to a vacutainer containing EDTA solution for the estimation of hemoglobin, mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC) and for peripheral blood smear evaluation. Hemoglobin, hematocrit, MCV, MCH and MCHC were estimated by automated counters. (Biologics ACCUCOUNT 1000 Automated Counter) A peripheral blood smear was stained with Leishman’s stain to determine the morphology of the blood cells and to diagnose type of anemia. The other part of the sample (3 mL) was used for the estimation of serum values of ferritin, iron, folate and cobalamin. It was centrifuged at 2000 rpm for 10 minutes. The separated serum was then transferred to aliquots and the aliquots were stored at -80°C for later measurement of above mentioned parameters. Serum ferritin, folate and cobalamin were measured by electro-chemiluminescence method using automated clinical immune-analyzer in a closed system (Elecsys 2010, Roche Diagnostics, Switzerland). Serum iron estimation was done using fully automated clinical chemistry analyzer (Olympus, Japan) by spectrophotometric method using ready to use Randox reagent kits (Randox laboratories, United Kingdom). Treatment of anemia was done. Reticulocyte count was evaluated after six days of commencing treatment by staining peripheral blood smear with Brilliant Cresyl blue stain to evaluate response to treatment. Monitoring of women in anemia group was done by measuring hemoglobin, hematocrit and red blood cell indices. Routine antenatal care with special importance for treatment of anemia was provided to the anemic women. Nonanemic women were followed up as routine on outpatient basis. All patients were followed till delivery. Delivery was conducted in hospital for all the patients included in the study. Cord blood samples were collected at the time of delivery for all patients for estimation of hemoglobin, hematocrit, red cell indices and serum values of iron, ferritin, folate and cobalamin. The study plan is represented in Figure 1. RESULTS A total of 200 patients with severe anemia (Hb< 7 gm %) as cases and 200 controls (Hb>11 gm %) were studied. On the basis of haematological indices and peripheral smear evaluation the cases were divided into three groups. Though microcytic hypochromic anemia was most common (94 patients, 47%), a significant number of patients were found to have macrocytic anemia (75 patients, 37.5%). Dimorphic anemia was present in a smaller fraction of women with severe anemia (31 patients, 15.5%). In Table 1 the demographic profile of the study population is reported. Majority of study population (148 patients, 74%) belonged to lower socioeconomic status according to modified Kuppuswamy scale and were multiparous (172 patients, 86%). One hundred and thirty five patients (67.5%) consumed vegetarian diet which is deficient in iron and vitamin B12. Further history revealed that only 90 patients (45%) of total study population had taken iron supplementation; this number was significantly less in patients with microcytic anemia (27 of 94 patients, 28.7%). Incidence of multivitamin (28 patients, 14%) and folic acid intake (11 patients, 5.5%) was also very minimal. Out of total anemic population, 110 patients (55%) received blood transfusion. Table 2 shows the biochemical parameters in different study groups. Deranged liver function test was seen more commonly in macrocytic anemia. Out of 200 women, who were anemic, 94 (47%) patients were iron deficient whereas cobalamin and folate deficiency were seen in 75 (37.5%) and 22 (11%) patients respectively. Table 3 shows comparison of maternal and cord blood biochemical parameters in different types of anemia. Tables 4 clearly shows that there is preferential transfer of all iron, ferritin and folate from mother to fetus but in case of cobalamin, fetal levels mirror maternal levels very closely. The reason for this lack of preferential transfer is not clearly understood. Figures 2-5 show comparison of iron, ferritin, cobalamin and folate levels respectively in maternal and cord serum in different types of anemia. Serum levels of iron, ferritin, cobalamin and folate were low in maternal blood. Cord blood levels showed correspondingly higher values of iron, ferritin and folate (in the lower normal range) but were almost similar to maternal levels in the case of cobalamin indicating the absence of preferential transfer to the fetus. DISCUSSION Anaemia is the most common nutritional deficiency disorder in the world. Anemia is of great concern during pregnancy because of its reported association with a number of adverse outcomes on both maternal and neonatal health. Lower socioeconomic status, poor nutritional status, closely spaced multiple pregnancies without supplementation of iron and multivitamins were found to be associated risk factors for anemia [1,2]. In the present study, macrocytic anemia was diagnosed in substantial number of patients (37.5%). This is in accordance with results of a study conducted in the same department three years back which showed 40% of overall patients of severe anemia had macrocytic picture [3]. However, in another Indian study, it was observed that amongst 130 pregnant women with severe anemia (Hb<5 g/dL), prevalence of microcytic anemia was 49%, followed by dimorphic 35% and macrocytic anemia 10% [2]. In western countries, prevalence of macrocytic anemia is comparatively less [5]. Prevalence of vitamin B12 deficiency is much higher in countries like India because majority of individuals consume purely vegetarian diet which is likely to be vitamin B12 deficient [6-8]. The increasing prevalence of vitamin B12 deficiency in different groups of population is highlighted in various studies [9-11], but there is paucity of literature regarding prevalence