Association of Age, Parity and Body mass Index with Hemoglobin and Serum Ferritin Levels in Pregnant Women in Baghdad City

Hemogloin (Hb) and serum ferritin levels are used to assess anemia in pregnancy. Some studies referred to the influence of maternal age, body mass index (BMI) and parity on Hb and serum ferritin levels. The study aimed to examine the possible association of maternal Hb and serum ferritin with maternal age, parity, and BMI in a sample of pregnant women in Baghdad. Ninety healthy pregnant women, grouped in three equal groups according to the pregnancy trimester, and thirty apparently healthy non-pregnant women from Baghdad were enrolled in this observational study. Blood and serum samples were obtained for the estimation of Hb and serum ferritin levels. The pooled data of participants showed a negative correlation between parity and each of blood Hb concentrations (r= -0.147, P=0.046) and plasma ferritin levels (r= -0.186, P= 0.038). The negative correlation of parity with blood Hb concentration was reported in participants in the third trimester of pregnancy (r= -0.270, P=0.048); and between parity and plasma ferritin levels in the second (r= -0.088, P= 0.046) and third (r= -0.398, P=0.029) trimester pregnant. The study did not report a significant correlation between age and BMI with blood Hb concentrations or serum ferritin levels in pregnant women at any trimester of pregnancy. There is a negative correlation between parity and each of blood Hb concentration and serum ferritin levels in pregnant women in Baghdad. While, there is no such correlation with maternal age and BMI at any trimester of pregnancy.


Introduction
Globally, anemia during pregnancy represents one of the most prevalent public health problems, causing negative maternal and fetal health effects (1) .The World Health Organization (WHO) defines anemia in pregnancy as the condition where hemoglobin (Hb) concentration is less than 11g/dL, at any trimester of pregnancy (2) .
While, according to the Centers for Disease Control and Prevention guidance, anemia in pregnancy is when Hb concentration less than 11 g/dL at the first trimester and less than 10.5 g/dL in the second or third trimesters (3) . It is estimated that 56% of pregnant women in low and middle-income countries and more than 10% in high-income countries have anemia. (1) Iron deficiency is the primary cause of anemia in about 75% of all anemia cases during pregnancy. (4) Other causes that are prevalent in developing countries involve micronutrient insufficiencies such as folic acid, vitamin A and vitamin B12; some parasitic infections including malaria and hookworm; and chronic infections like tuberculosis and acquired immunodeficiency syndrome (5-7) . In pregnancy, without supplementary iron, Hb concentration declines due to physiological hemodilution. Serum ferritin is sensitive indicator of body iron stores, and it is considered as a reliable indicator of iron status in pregnant women. (8) At the beginning of pregnancy, serum ferritin concentrations vary more widely than corresponding Hb values (9) . Some studies referred to the influence of maternal age, body mass index (BMI) and parity on Hb and ferritin levels (10)(11)(12) . The aim of the present work is to study the possible association of maternal Hb and serum ferritin with maternal age, parity, and BMI in a sample of Iraqi pregnant women.

Subjects and Methods
A cross-sectional study conducted at the Antenatal clinic in Baghdad Teaching Hospital in Baghdad city, the capital of Iraq, during the period extending from May to July 2019. Ninety pregnant women, aged 18-45 years, were divided into three equal groups according to the trimester of pregnancy; and 30 apparently healthy non-pregnant married women to serve as control group, were enrolled in the study. Participants with history of diseases or treatments that interfere with blood Hb concentrations or serum ferritin levels were excluded from the study. Data regarding participants' age, BMI (calculated as weight in kilograms (kg) divided by height in meters squared (m 2 )) and parity, are collected and recorded using a data collection sheet designed for the purpose of the study. Five ml blood samples were collected, 2 ml in EDTA tube for estimation of Hb and 3 ml in plain tube for serum separation. The collected anticoagulated blood samples were used for Hb concentrations measurement using an automated hematology analyzer. While, serum samples were stored at -20°C until the time of estimation of ferritin levels by enzyme-linked immuno-sorbent assay (ELISA) test.

Ethical considerations
The ethics committee of the College of Pharmacy, University of Baghdad approved the study. Informed consent was obtained from each participant before participation in the study.

Statistical analysis
Statistical analysis was performed using SPSS ® software version 22 for windows. Data are presented mainly as mean values and standard deviation (SD) or frequencies and percentages. Shapiro Wilk test was used to check the distribution normality for the dependent variables, blood Hb concentrations and serum ferritin levels. Mann-Whitney U test was conducted to examine differences between means of two groups, while Kruskal-Wallis H test was used to compare means among three or more groups. The Pearson's correlation coefficient (r) was used to evaluate the association between variables. A P value < 0.05 was considered statistically significant.

Results
The present study showed a significant differences between the means of blood Hb concentration and plasma ferritin levels of the pregnant and non-pregnant participants (P= 0.001 and P= 0.033; respectively). Meanwhile there is no significant difference between the means of age, BMI and parity of the pregnant and non-pregnant participants (P≥0.05) ( Table 1). Comparison of participants' characteristics by the trimester of pregnancy showed that there is significant differences among means of BMI (P< 0.001), blood Hb concentrations (P< 0.001) and serum ferritin levels (P=0.029) of the studied groups (Table 2). Pairwise comparisons revealed that BMI mean of the participants in the third trimester of pregnancy is significantly higher than that of the non-pregnant or the first trimester pregnant participants. While, the mean of blood Hb concentrations of the participants in the third trimester of pregnancy is significantly lower than that of the non-pregnant or the first trimester pregnant participants. Moreover the mean of blood Hb concentrations of the participants in the second trimester of pregnancy is significantly lower than that of the non-pregnant participants. Finally, mean serum ferritin levels of the participants in the third trimester of pregnancy is significantly lower than that of the non-pregnant participants. Correlation tests of the pooled data of pregnant participants of age, parity and BMI with blood Hb concentrations and serum ferritin levels, showed that there is a negative correlation between parity and each of blood Hb concentrations (r= -0.147, P=0.046) and plasma ferritin levels (r= -0.186, P= 0.038). Other correlation pairs testing did not show a significant correlation (P>0.05) ( Table  3).
Correlation tests of age, parity and BMI with blood Hb concentrations and serum ferritin levels of the study groups showed that there is a negative correlation between parity and blood Hb concentrations in the third trimester pregnant (r= -0.270, P=0.048); and between parity and plasma ferritin levels in the second (r= -0.088, P= 0.046) and third (r= -0.398, P=0.029) trimester pregnant. Other correlation pairs testing among the study groups did not show a significant correlation (P>0.05) ( Table 4).

Discussion
The study showed that 52.2% (47 of 90) of the pregnant participants are anemic with mean Hb concentration less than 11 gm/dL. The prevalence of anemia in pregnant women in the present study is higher than that reported by a recent study conducted in Baghdad (33.8%) which was conducted on larger cohort (400 participants). (13) Yet, the prevalence of anemia in pregnant women in low and middle-income countries was reported to be 56%. (1) The present study also showed that serum ferritin levels in pregnant women showed a significantly lower value than that of the nonpregnant. Physiologic anemia occurring during a healthy pregnancy is attributed mainly to increase in the plasma volume without sufficient increase in red cells count. (14) In young adult women, the stored iron is roughly about one tenth of the iron incorporated in hemoglobin. (15,16) During pregnancy, the red cell volume expands to about 24-31% of the non-pregnant levels; (17,18) this expansion of red cell volume results in depletion of the iron stores in most of pregnant women and hence serum ferritin levels. (19) On the other hand, the present study showed that 13.3% (4 of 30) of the nonpregnant participants have mean Hb concentration less than 11 gm/dL. Using the cut-off point of Hb for diagnosis of anemia in non-pregnant women is 11.9 will escalate the percent of anemic non-pregnant participants to 23.3% (7 of 30). The prevalence of anemia in the non-pregnant group is high, but it is still slightly lower than that reported in Iraq as of 2016 (29.1%). (20) Hypervolemia and the increase in fetal weight may represent the main causes of the increase in BMI with the advancement in gestational age. The results showed a negative correlation between parity and each of blood Hb concentration and serum ferritin levels, which suggests a reduction of iron store with increasing parity. The results of this study in this regard occur in accordance with many reports, (21,22) and disagree with others that reported no association between high parity and anemia or low iron stores. (23)(24)(25) Moreover, some studies reported a reduction in risk of anemia with high parity. (26,27) Increased risk of bleeding during pregnancy as well as during and after delivery may represent the link between anemia and low iron store and increasing parity. Several mechanisms were proposed to explain the increased risk of bleeding with increased parity; yet, none of these mechanisms have been proven. (28) The study did not show any significant correlation between age and BMI with blood Hb concentrations or serum ferritin levels in pregnant women at any trimester of pregnancy. This result do not agree with the results of other studies. For example, Mocking et al. has reported a positive correlation between BMI in first trimester and blood Hb concentrations in Indonesian and Ghanaian women; (29) while, Ramussen et al. has reported a significant association of blood Hb concentrations during pregnancy with maternal age, and with body mass index; with no correlation with serum ferritin levels (11) .

Conclusion
Anemia is very prevalent among pregnant women in Baghdad. There is a negative correlation between parity and each of blood Hb concentration and serum ferritin levels. While, there is no such correlation with maternal age and BMI at any trimester of pregnancy. The present study has some limitations. First, the causality between the correlated variables cannot be determined because of the nature of cross-sectional study. Second, being a single center study on relatively small sample size, prevent the finding to be generalized on pregnant women in Baghdad. Multi-center and larger cohort study is recommended.