A comparative study on ABO blood group and fertility hormones in infertile women in Calabar, Southern Nigeria

Received 09 May 2018 Revised 24 May 2018 Accepted 28 May 2018 Online 30 June 2018 Print 30 June 2018 Infertility, a disorder of the reproductive system, is commonly linked to hormonal, pituitary, cervical, uterine, immunological or psychological factors. Besides these factors, it can also be idiopathic or unexplained. Hence, there is a need for more research to unravel the causes of the unexplained infertility. This work aimed at finding out whether there is any relationship between ABO blood group system and female infertility. The study design was cross-sectional. Three hundred women between 18 and 40 years attending fertility clinic at the University of Calabar Teaching Hospital, Calabar between 2011 and 2012 were recruited for this study. Serum progesterone, prolactin, follicle stimulating hormone (FSH), luteinizing hormone (LH) and estradiol were determined using Enzyme linked immunosorbent assay (ELISA) while ABO blood grouping was determined using the tube method. Statistical analysis was done using SPSS version 18. The confidence level was set at 95% where p-value less than 0.05 was considered statistically significant. The mean age of the women was 30.65 ± 5.47 years and the percentage distributions of the blood groups among the women were as follows: 24% A, 12% B, 4% AB and 60% O. The mean FSH levels of blood groups A and O individuals were significantly higher (p<0.05) than that of groups B. However, there was no significant difference (p>0.05) in the mean levels of progesterone, prolactin, LH and estradiol in the respective groups. From this study, 38% of the population had increased levels of progesterone, 58% and 18.7% had elevated prolactin and FSH levels respectively while 11.33% and 43.3% had reduced levels of LH and estradiol levels respectively. Though, there was high prevalence of hyperprolactinemia observed in this study, there was no strong association between ABO blood group and female infertility but, the increased FSH levels observed in blood groups A and O may be a potential link between blood group and infertility and therefore may be beneficial for further study. Keywords

Infertility, a disorder of the reproductive system, is commonly linked to hormonal, pituitary, cervical, uterine, immunological or psychological factors. Besides these factors, it can also be idiopathic or unexplained. Hence, there is a need for more research to unravel the causes of the unexplained infertility. This work aimed at finding out whether there is any relationship between ABO blood group system and female infertility. The study design was cross-sectional. Three hundred women between 18 and 40 years attending fertility clinic at the University of Calabar Teaching Hospital, Calabar between 2011 and 2012 were recruited for this study. Serum progesterone, prolactin, follicle stimulating hormone (FSH), luteinizing hormone (LH) and estradiol were determined using Enzyme linked immunosorbent assay (ELISA) while ABO blood grouping was determined using the tube method. Statistical analysis was done using SPSS version 18. The confidence level was set at 95% where p-value less than 0.05 was considered statistically significant. The mean age of the women was 30.65 ± 5.47 years and the percentage distributions of the blood groups among the women were as follows: 24% A, 12% B, 4% AB and 60% O. The mean FSH levels of blood groups A and O individuals were significantly higher (p<0.05) than that of groups B. However, there was no significant difference (p>0.05) in the mean levels of progesterone, prolactin, LH and estradiol in the respective groups. From this study, 38% of the population had increased levels of progesterone, 58% and 18.7% had elevated prolactin and FSH levels respectively while 11.33% and 43.3% had reduced levels of LH and estradiol levels respectively. Though, there was high prevalence of hyperprolactinemia observed in this study, there was no strong association between ABO blood group and female infertility but, the increased FSH levels observed in blood groups A and O may be a potential link between blood group and infertility and therefore may be beneficial for further study.

Introduction
Fertility can be referred to as childbearing performance while fecundity means childbearing ability although both terms are used interchangeably [1]. Ideally in most societies once a marital relationship is established between a man and a woman their expectation and that of their 20 Ann Med Physiol. 2018; 2(2) relatives as well is reproduction which follows in the sequence of conception, pregnancy and a healthy live birth. No deep thought is put into the physiology and processes of reproduction if maternity is hitch-free [2]. However, sometimes apparently healthy couples experience a condition of inability to conceive after 12 months or more of regular unprotected sexual intercourse. This condition is often referred to as infertility [3]. Infertility as a spectrum of reproductive failure is a social, economic, psychological as well as medical problem [4]. The incidence of infertility in a population has important demographic and health implications as high infertility has a dampening effect on population growth rate [3]. As a result of the social and psychological consequences of infertility, women have been known to manifest various psycho-sexual disorders while some may have decreased sexual drive and some may seek multiple sexual partners in an attempt to achieve a desired conception [5]. The World Health Organization (WHO) estimates that 8-12% of couples globally experience problems in conception [6].
Psychologically, infertility investigation probes into the most intimate and private life of an individual as the history involves questions about the patient's sex life, menstrual cycles of females and examination commonly incorporates genital examinations [7]. The common factors described to be the causes of female infertility include ovarian or hormonal factors, pituitary, cervical, uterine, immunological, psychological and iatrogenic factors [8]. In addition to the factors above, infertility can be idiopathic; in such cases it is loosely termed unexplained infertility [9]. That is why effort is demanded on research to unravel the latent cause of the unexplained infertility as a step in tailoring the solutions to patients looking for assistance [10]. There are a lot of contradicting reports on the relationship of ABO blood groups with female fertility. A study conducted in China suggested that women with blood types A and AB were at increased risk of Diminished Ovarian Reserve (DOR) while those with type O were at low risk. Although, this is in contrast to the findings of Nejat et al who conducted a similar study in the United States of America (USA) and suggested that A blood group antigen was protective against DOR in sub-fertile women than the O blood group antigens [11]. While another study by Timberlake et al in the USA and Yilmaz et al in Turkey reported that they found no clinical implications for particular blood type as a risk factor for DOR [12,13]. However, the underlying mechanisms behind these findings are still not clear. These studies were on Caucasian and Asian populations. Therefore, this work investigated the relationship if any between ABO blood group and infertility in African women.

Study design and subject selection
The design of the study was cross-sectional. A total of three hundred women participated in this study. The participants were consecutively recruited from women of reproductive age (18-40 years) attending the fertility clinic at University of Calabar Teaching Hospital (UCTH), Calabar in Southern Nigeria. All the subjects were infertile and any subject above forty years of age was excluded. Ethical approval was obtained from the Ethics committee of the University of Calabar Teaching Hospital, Calabar (RP/REC/2015/311). The purpose and nature of the research was explained to the participants and informed consent was obtained from them.

Sample size calculation
The number of samples used in this research was determined using the Leslie Kish formula [14] where N = desired sample size Zα = the α level of the coefficient interval at 95% (1.96) p = proportion of occurrence q = (1-p) proportion of non-occurrence d = precision Substituting the expected occurrence of p=12% i.e. 0.12 from WHO [6], we have

Sample collection
On the 21 st day of their menstrual cycle, four milliliters (4 ml) of blood were aseptically collected from each subject into a plain container and allowed to clot at room temperature. The samples were spun at 3000 rpm for 10 minutes. The supernatant sera were then harvested into clean specimen containers and labeled accordingly. The sera were stored frozen at 4⁰C until needed for assay of the hormonal parameters. Some of the serum and cells were used immediately for serum and cell grouping respectively.

Estimation of hormones and ABO blood group determination
Serum progesterone, prolactin, follicle stimulating hormone (FSH), luteinizing hormone (LH) and estradiol were analyzed using Enzyme Linked Immunosorbent Assay (ELISA) kits obtained from Syntron Bioresearch Incorporated Carlsbad, California. The analyses were carried out using the

Statistical analysis
This was done using the PAWS 18, a statistical package from SPSS Inc, Chicago, USA. The results were expressed as Mean ± SEM. The data was analyzed using one way analysis of variance (ANOVA) and the significance level was set at 95% confidence interval where p-value less than 0.05 (p<0.05) was considered as statistically significant.  Table 1 and Table 2) while there was no significant difference (p>0.05) in the mean serum levels of progesterone, prolactin, LH and estradiol of the different blood groups. Table 3 shows the percentage distribution of the different hormones of the subjects with different blood groups within normal, above normal and below normal reference limits. More than 22% of blood group O subjects had elevated FSH levels while 50% had reduced estradiol levels. Hyperprolactinemia was observed across all the groups with group B having the highest distribution (69%) and group A with the lowest (55%). A reduction in the LH levels was also observed in more than 45% of each group. Table 4 shows the percentage distribution of hormones in the sample population. Thirty eight percent (38%) of the total population had increased progesterone levels though more than 50% had normal progesterone levels. Fifty eight percent (58%) had elevated prolactin levels in the study. Low levels of estradiol were observed in 43.3% of the study population while 18.7% and 53.3% had increased levels of FSH and LH respectively.    . Although the exact mechanisms that tie blood type to ovarian reserve and infertility are unclear, the possibilities that glycosyltransferases may play an important role in ovarian function or that the presence of a gene that co-segregates with DOR, are both enticing and constitute an area for further investigations [11]. In summary, from this study, 38% of the sample population had reduced levels of progesterone, suggestive of anovulation and amenorrhea. 58% had elevated prolactin level while 53.3% had reduced levels of LH which is seen in the cases of hyperprolactinemia and 45.3% had depressed estradiol levels.

Conclusion
Though, a high prevalence of hyperprolactinemia among the infertile women was observed, a strong evidence of relationship between female infertility and ABO blood group system was not established from this study but, the increased FSH levels observed in blood groups A and O may be a potential link between blood group and infertility and therefore may require further study.

Limitation of the study
In our study, day 21 of the participants' menstrual cycle was considered as all of them came to the clinic for luteal function assessment. Those for ovarian reserve function that came to the clinic on day 2 or 3 were few in number and were excluded from the study in order to maintain uniformity and obtain a larger sample size. We recommend that further studies be carried out, with hormonal profile on day 2 or day 3 of the participants' menstrual cycle taken into consideration.