Pa ge 1 Pa ge 14 1 American Journal of Medical Science and Innovation (AJMSI) Serum Levels of Thyroid Hormones in Infertile and Fertile Women Attending a Tertiary Care Hospital in North-Central Nigeria: A Comparative Reproductive Medicine Study Halima Inya Aliu-Ayo1, Kikelomo Temilola Adesina2, Abiodun Akanbi Gafar Jimoh2, Aloysius Obinna Ikwuka3* Francis Chigozie Udeh3, Sikiru Abayomi Biliaminu4, Olayinka Waheed Ayo5 Volume 2 Issue 2, Year 2023 ISSN: 2836-8509 (Online) DOI: https://doi.org/10.54536/ajmsi.v2i2.2044 https://journals.e-palli.com/home/index.php/ajmsi Article Information ABSTRACT Received: August 28, 2023 Accepted: September 30, 2023 Published: October 14, 2023 Thyroid hormones have profound effects on reproduction. Normal thyroid function is necessary to maintain reproductive physiology, but its functional abnormalities may cause infertility manifesting as menstrual cycle irregularities. Aim of this research was to determine the serum levels of thyroid hormones in infertile and fertile women attending a tertiary care hospital in North-Central Nigeria. This comparative, reproductive medicine, cross-sectional study was carried out among one hundred and six (106) women attending the Gynecology Clinic of the Department of Obstetrics and Gynecology and the Family Planning Clinic of a tertiary care hospital in North-Central Nigeria. Women with a history suggestive of either primary or secondary infertility and the control women with no history of infertility were evenly grouped. Venous blood samples were collected and analyzed using Enzyme- Linked Immunosorbent Assay (ELISA) technique to determine fT3, fT4 and TSH levels. Data obtained were entered and analyzed using Statistical Package for Social Sciences (SPSS) version 23, and the level of statistical significance was set at p<0.05 and at a confidence interval of 95% for all inferential statistics. Majority (90.6%) of the 106 participants were euthyroid. However, most of the thyroid disorders (80%) were noticed in the infertile women, as 15.1% of them had one form of thyroid disorder. The mean TSH, fT3 and fT4 levels were higher in the infertile patients than in the control group, but the values were still within normal reference range. Mean values of TSH, fT3 and fT4 for the patients and controls were 1.35±1.65 versus 0.85±1.08 mIU/mL, 2.79±1.51 versus 2.19±1.15 pg/ mL, and 1.15±0.33 versus 0.99 ± 0.29 ng/dL respectively. Median for TSH, fT3 and fT4 in the patients and controls were as follows: 0.80 (0.40–1.55) versus 0.60 (0.40-0.85) mIU/ mL, 2.40 (1.45-4.20) versus 1.70 (1.35-2.95) pg/mL, and 1.10 (1.00-1.30) versus 1.00 (0.90- 1.10) ng/dL respectively. The differences in the median between the patients and controls were statistically significant for fT3 and fT4 with p-values of 0.031 and 0.002 respectively. Subclinical hypothyroidism was the most prevalent (7.5%) thyroid disorder identified in the infertile women. Serum levels of thyroid hormones and prevalence of thyroid disorders were significantly higher in the infertile patients compared to the fertile women, although the overall prevalence of thyroid disorders was quite low. Keywords Thyroid Hormones, Thyroid Disorders, Female Infertility, Female Fertility, Reproductive Medicine, Comparative Study, Reproductive Endocrinology, Reproductive Gynecology 1 Department of Obstetrics and Gynaecology, R-Jolad Hospital, Lagos, Nigeria 2 Department of Obstetrics and Gynaecology, University of Ilorin & University of Ilorin Teaching Hospital, Ilorin, Nigeria 3 College of Medicine and Health Sciences, American International University West Africa, Banjul, The Gambia 4 Department of Chemical Pathology and Immunology, University of Ilorin & University of Ilorin Teaching Hospital, Ilorin, Nigeria 5 The Bridge Clinic, Lagos, Nigeria * Corresponding author’s e-mail: aloysiussweet@yahoo.com INTRODUCTION Gametes are ova and sperm cells that are haploid and have one copy of each type of chromosome i.e. 1–22 X or 1–22 Y (Ikwuka, 2023a). The sperm cell must fertilize an ovum in vivo or in vitro for conception (pregnancy) to occur. Infertility is a global health problem, a social demoralizing condition for couples and it is an important cause of marital disharmony (Panti, 2014). It is defined as failure to achieve clinical pregnancy after 12 months (1 year) of regular unprotected sexual intercourse – regular sexual intercourse is 2-3 times sexual intercourse per week (World Health Organization, 2010). Infertility can be further classified into primary infertility, in which no previous pregnancies have occurred and secondary infertility in which a prior pregnancy, although not necessarily a live birth, has occurred (Yao, 2002). The incidence of infertility suggests that 10-15% couples experience infertility (Bhattacharya, 20007), constituting the major cause of gynecological consultations in Nigeria (Jimoh, 2011; Obuna, 2012; Okonofua, 2005). One of the greatest desires of couples is successful reproduction, especially in Africa, where a high premium is placed on childbearing. Infertility therefore causes severe emotional and social distress for the couple especially the social stigma attached to it in Africa (Gerias, 1992). The endocrine system is the second key regulator of organ system function after the nervous system. The endocrine system uses hormones as chemical messengers in signaling, once stimulated. Upon stimulation, the thyroid gland secretes thyroid hormones (THs) which include tri-iodothyronine (T3) and thyroxine (T4). These hormones have a role in controlling basal metabolic rate (BMR), growth, and the development and differentiation of many cells in the body (Habbu, 2016). Normal thyroid gland function is described as euthyroidism, below normal as hypothyroidism, and above normal as hyperthyroidism. Pa ge 14 2 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. Innov. 2(2) 141-152, 2023 Hyperthyroidism has been linked with oxidative stress and various systemic immune inflammatory processes. In inducing oxidative stress, major free radicals that are of physiological significance are superoxide anion, hydroxyl radical, and hydroperoxyl radical, while non- radical is hydrogen peroxide (Ikwuka, 2023b). In addition, the interplay, role and effects of metabolic syndrome diseases on female fertility are still being investigated by different researchers. Metabolic syndrome diseases, MSD (Hypertension, etc) are interrelated diseases with very high morbidity and mortality rates (Ikwuka, 2015; Ikwuka, 2017a; Ikwuka, 2017c; Ikwuka, 2023c; Virstyuk, 2016). Results from different studies have shown that high levels of blood pressure, glucose and lipid metabolic disorders, asymptomatic hyperuricemia, activation of systemic immune inflammation and fibrogenesis, contribute to kidney damage (Ikwuka, 2017d; Ikwuka, 2017e; Ikwuka, 2018a; Ikwuka, 2018c; Ikwuka, 2018d; Ikwuka, 2019a; Ikwuka, 2019c; Ikwuka, 2022; Ikwuka, 2023d; Virstyuk, 2017a; Virstyuk, 2018a; Virstyuk, 2019; Virstyuk, 2021a; Virstyuk, 2021b). Adiposity, diabetes mellitus and dyslipidemia have also been linked with erectile dysfunction (Baysah, 2023). Other than hormonal imbalance, several factors including hazards linked to certain occupations (long distance driving, military, etc); previous history of genital infections (e.g. gonorrhea, orchitis, etc); previous history of surgery in the genital tract or inguinal region; lifestyle choices (sedentary lifestyle, lack of exercise, sleep deprivation, etc); environmental factors (e.g. exposure to ionizing radiation, pesticides, heat from tight underwears and hot baths) can adversely affect semen parameters and lead to male infertility (Baysah, 2023). In addition, lifestyles such as alcohol and tobacco consumption have been reported to cause infertility in men (Baysah, 2023). The effects of alcohol intake on semen quality have been reported to be caused by oxidative stress caused by an imbalance between Reactive Oxygen Species (ROS) produced by the alcohol consumed in the form of free radicals that contain one or more unpaired electrons and antioxidants (Ekechi, 2023). Nicotiana tabacum (local snuff) has been reported to have adverse effects on fertility and pregnancy in female wistar rats (Udeh, 2023a; Udeh, 2023b). An established association between thyroid dysfunctions and infertility exists. These dysfunctions in the form of hyper- or hypothyroidism can adversely affect fertility by the presence of anovulatory cycles, luteal phase defects, high prolactin (PRL) levels, sex hormone imbalances, delayed onset of puberty, menstrual abnormalities, and miscarriages (Doufas, 2000; Poppe, 2003; Poppe, 2007; Rijal, 2011). However, the relative frequency and the chronology of the onset of reproductive dysfunction concerning the onset and type of thyroid disorder have not been well defined (Deshmukh, 2015). Increasing evidence derived from experimental and clinical studies suggests that the hypothalamic-pituitary- thyroid axis and hypothalamic-pituitary-ovarian axis are physiologically related. Thyroid hormones (THs) receptors and their mRNA have been detected in human granulosa cells and direct effects of iodine and THs on ovarian function have been proposed recently (Poppe, 2003). TH scan directly affects granulosa cells, corpus luteum and oocytes (Biswas, 1993), and they regulate follicle-stimulating hormone (FSH) stimulation in follicles and prevent their apoptosis (Kabodmehri, 2021). Thyroid-stimulating hormone (TSH) working in synergy with FSH can proliferate the granulosa cells (Kabodmehri, 2021). Therefore, an increase in TSH and thyrotropin-releasing hormone (TRH) can cause ovulatory dysfunction or corpus luteum dysfunction. In addition, the underperformance of the thyroid gland can affect ovarian function indirectly by decreasing the binding activity of sex hormone binding globulin (SHBG), increasing prolactin (PRL) levels and delaying luteinizing hormone (LH) response to gonadotropin- releasing hormone (GnRH) (Lee, 2014). The prevalence of thyroid dysfunction in infertile women is quite high. In a study done at Aminu Kano Teaching Hospital, Kano, Nigeria; thyroid disorders were observed in 23.4% of the subjects (Emokpae, 2011). The prevalence of thyroid dysfunction in infertile women was found to be 33.3% in a study by (Rahman, 2008) in India and 23% by (Sharma, 2012) while (Shivaleela, 2012) found a prevalence of 42%. However, a study done in Port-Harcourt, Nigeria by (Orazulike, 2018) showed a much lower prevalence of thyroid disorders in infertile women, which was 4.6%. Subclinical thyroid dysfunctions (i.e. subclinical hyperthyroidism or subclinical hypothyroidism) may go unnoticed by unwary clinicians because these patients do not exhibit clinically overt physical symptoms and signs. This may lead to avoidable surgical interferences and related complications. With the advent of modern techniques, the estimation of various hormones can be done rapidly and reliably. The diagnosis of thyroid disorders can easily be made and appropriate treatment instituted with the results usually very gratifying. Metabolic Syndrome Diseases require new and effective treatment regimens. Dapagliflozin which is a Sodium- Glucose Linked Transporter 2 (SGLT-2) inhibitor and Liraglutide which is a Glucagon-like Peptide 1 Receptor Agonist (GLP-1 RA) have been found to increase the effectiveness of treatment and improve the clinical course of type 2 diabetes mellitus and hypertension in patients with such comorbidities (Ikwuka, 2017b; Ikwuka, 2018b; Ikwuka, 2019b; Ikwuka, 2021; Virstyuk, 2017b; Virstyuk, 2018b; Virstyuk, 2018c). Rauwolfia vomitoria has a neuroprotective ability at it elevates antioxidants and suppresses lipid peroxidation (Ekechi, 2023). Proper management of thyroid dysfunction results in improvement in health status, normalization of menstrual abnormalities and restoration of normal fertility (Micińsk, 2006). Therefore, it is very important to screen thyroid abnormalities in women with infertility, particularly in countries considered as areas with endemic goitre since female infertility associated with thyroid dysfunction in these areas is common (Zimmermann, 2008). Pa ge 14 3 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. Innov. 2(2) 141-152, 2023 MATERIALS AND METHODS Study Setting This study was conducted at the Gynecology Clinics of the Department of Obstetrics and Gynecology of the University of Ilorin Teaching Hospital (UITH), Ilorin, Kwara State, North-Central Nigeria. UITH is located at Oke-Oyi, Old Jebba Road in Ilorin. The hospital serves as a major referral centre for Kwara State and parts of the nearby states of Oyo, Osun, Ekiti, Kogi and Niger. UITH is a tertiary care hospital, although it also offers primary and secondary healthcare services, and it is approved to undertake undergraduate and postgraduate medical training. The Gynecology Clinic is open to all, and an average of 144 women access healthcare at the clinics weekly, with 25% of them being for infertility cases (follow-up and new cases). Study Population The study population consisted of reproductive age women (18-45 years) attending the Gynecology Clinic of UITH, and with a history of inability to conceive of more than one year duration. This included those with previous pregnancies irrespective of outcome (secondary infertility) and those with no previous pregnancy (primary infertility). The control group consisted of consenting fertile women, who had carried pregnancy to term within two years prior to this study, and who were new patients at the Family Planning Clinic of UITH. Sample Size and Sampling Technique The formula for sample size calculation for a comparative, cross-sectional study was used (Charan, 2013). The parameter, proportion of thyroid disorders in women, was extracted from a previous study by (Fatima, 2014). N= (2(Z(α/2)+ Zβ ) 2 ˟ P(1-P))/(P1-P2 ) 2 N= Sample size Z(α/2)= 1.96 at type 1 error of 5% Zβ= 1.28 at 90% power P1= Proportion of thyroid disorder in infertile women (47%) P2= Proportion of thyroid disorder in fertile women (16%). P1 and P2 were extracted from a previous study by (Fatima, 2014). P= Pooled prevalence = [prevalence in one group (P1) + prevalence in another group (P2)]/ 2 P= [(0.47 + 0.16) / 2] = 0.32 N= (2(1.96+ 1.28)2 ˟ 0.32(1-0.32))/(0.47-0.16)2 N= 2(10.497)(0.2176)/(0.31)2 N= 2(10.497)(0.2176)/0.0961 N= 47.5 N~ 48 patients To make provision for attrition, 10% of the sample size was added. Thus 53 women in each arm of the study, and a total sample size of one hundred and six (106) women were recruited for the study. Respondents (patients and controls) were recruited consecutively till the sample size was complete. Inclusion Criteria Study participants were consenting women of reproductive age group (18-45 years) with primary or secondary infertility. Control group consisted of consenting fertile women matched for age. The control group members were new clients at the Family Planning Clinic, apparently healthy, with no history of infertility, have carried a pregnancy to term, and had no record of contraceptive use two years prior to this study. Exclusion Criteria Female patients outside the age group of 18-45 years; with previous or present thyroid disorders; women on steroids, hormonal contraceptives, or intrauterine contraceptive devices; women with a history of bleeding disorders; women with co-existing uterine fibroids (leiomyoma uteri). Data Collection Method The study spanned 6 months. Patients who presented with infertility were selected based on the inclusion criteria. Eligible women were informed and counseled about the study in a simple language that they understood. The study proforma was designed to obtain respondents’ socio-demographic status, history of infertility, history of anterior neck swelling and other symptoms suggestive of thyroid disorder, medical conditions and past surgical procedures. Sample Collection Four milliliters (4ml) of whole blood sample was collected from a peripheral vein from each participant after an overnight fast of 8-10 hours into a labeled plain sample bottle. Food affects TSH levels, TSH decreases postprandial and a possible explanation for this is food-induced elevation of circulating somatostatin and consequent suppression of TSH (Takano, 1995). Patients who had coincidentally fasted for 8-10 hours at the first contact with the researcher were recruited while those who had already eaten were recruited at their next visit after they had been informed. Blood samples were allowed to stand for about 1-2 hours to allow clotting and retraction. The sera was drained from the clot and later centrifuged at 1,000 revolutions per minute (rpm) for 10 minutes. The whole sera obtained was transferred into labeled plain containers and kept frozen at -20ºC before analysis. Laboratory Procedures Serum concentrations of TSH, fT3 and fT4 were determined by using Enzyme-linked immunosorbent assay (ELISA) kits manufactured by Monobind Ltd using Rayto microplate well reader. All reagents, calibrators/ standards and patients’ specimens were brought to room temperature (20-27ºC). The microplate wells for each calibrator/ standard and patient specimen were formatted, and the following steps described by (Winter, 2012) were taken: 1. 50μl of the calibrator and the patient specimen were dropped into the assigned well with a pipette. Pa ge 14 4 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. Innov. 2(2) 141-152, 2023 2. 100μl of TSH, fT3, and fT4 enzyme reagent was added to each well. 3. The microplate was swirled gently for 20-30 seconds and then incubated for 60 minutes at room temperature. 4. The contents of the microplate were discarded by decantation, and the plate was tapped and blotted dry with absorbent paper. 5. 350μl of wash buffer was added followed by decantation, a process that was repeated 2 additional times. 6. 100μl of working substrate solution was added to all wells without shaking the plates. This was incubated for 15 minutes at room temperature. 7. 50μl of stop solution was added to each well and mixed for 15-20 seconds. 8. The absorbance/optical density was read in each well at 450nm. The results were read within 30 minutes of adding the stop solution. 9. Calibration curves were plotted for each analyte to determine the corresponding concentration of these analytes in the patients’ samples. Criteria for Diagnosis of Thyroid Dysfunction The reference range of values for THs that was used for this study is the range of normal values on the kit used (TSH: 0.39-6.16 mIU/mL; fT3: 1.4-4.2 pg/mL; and fT4: 0.7-2.0 ng/dL) (Monobindinc, 2012a; Monobindinc, 2012b; Monobindinc, 2012c). Abnormal thyroid function can be categorized as hyperthyroidism (elevated fT3, fT4, and decreased TSH), hypothyroidism (decreased fT3, fT4, and elevated TSH), subclinical hyperthyroidism (normal fT3, fT4, but low TSH), and subclinical hypothyroidism (normal fT3, fT4, but elevated TSH) (Habbu, 2016). Data Analysis Data was analyzed using Statistical Package for Social Sciences (SPSS) version 23. Categorical variables (thyroid disorders) were presented as proportions and analyzed using the Chi-square test with Fisher’s exact correction used in cases where more than 20% of expected counts were less than 5. The continuous variable that is normally distributed (like age) was presented as mean and analyzed using the T-test while those that were not normally distributed (the THs levels) were presented as median and interquartile range and analyzed using the Mann- Whitney U test. The test of normality was done using the Shapiro-Wilk test. The level of statistical significance was set at p<0.05 at a confidence interval of 95% for all inferential statistics. Ethical Considerations Ethical approval for this study was obtained from the Ethical Review Committee of UITH. Informed written consent was obtained from each participant after adequate counseling and all data from the study were treated with confidentiality and used solely for the study. Patients with thyroid abnormalities were referred for further evaluation, treatment, and were co-managed with endocrinologists. RESULTS AND DISCUSSION The findings in the one hundred and six (106) study participants are as shown below. Socio-Demographic Variables of Participants Table 1 shows the socio-demographic variables of the infertile women and control group. Women in the age group 25-28 years had the highest frequency of 30.2%, while women below 25 years of age had the lowest frequency of 13.2%. The mean age for the infertile and control group was 32.18±6.47 years and 31.4±5.74 years respectively. The majority of the patients and control (79.2% and 81.1%) respectively were employed. The majority of the study participants had a tertiary level of education (69.8%) and were Yorubas (83%). There were no significant differences in the age, occupation, educational status and ethnicity of the study participants in the two groups. Table 1: Socio-demographic variables of the infertile women (patients) and fertile (control) group Socio-demographic variable Patient n (%) Control n (%) Total n (%) χ2/ t p-value Age (years) < 25 9 (17.0) 5 (9.4) 14 (13.2) 2.932 0.569 25 – 29 14 (26.4) 18 (34.0) 32 (30.2) 30 – 34 7 (13.2) 11 (20.8) 18 (17.0) 35 – 39 15 (28.3) 12 (22.6) 27 (25.5) 40 – 45 8 (15.1) 7 (13.2) 15 (14.2) Total (n) 53 53 106 Age (Mean ± SD)* 32.18±6.47 31.40±5.74 0.651t 0.516 Occupational status Employed 42 (79.2) 43 (81.1) 85 (44.3) 0.509 0.808 Unemployed 11 (20.8) 10 (18.9) 21 (19.8) Total (n) 53 53 106 Educational status None 1 (1.9) 1 (1.9) 2 (1.9) 1.784f 0.759 Pa ge 14 5 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. Innov. 2(2) 141-152, 2023 Gynecological Variables and Symptoms Found among the Infertile and the Fertile Groups Figure 1 illustrates the types of infertility found among the infertile patients. Secondary infertility was identified in 75.5% of the infertile women while 24.5% had primary infertility. Table 2 illustrates the symptoms present in the infertile patients and the control (fertile) group. None of the participants presented with anterior neck swelling. Two (3.8%) of the infertile patients experienced palpitations compared to three (5.7%) in the control group. Only one (1.9%) patient lost weight as against none in the control. Four (7.5%) patients had a history of abnormal weight gain as against two (3.8%) in the control group. Previous history of miscarriages was more prominent in infertile women (30.2%) than in fertile women (7.5%). Heat or cold intolerance was only evident among the infertile women (3.8%), and this group had more members (7.5%) with decreased libido than 1.9% seen in the control group. Of all these symptoms, only previous miscarriages presented a significant difference (p-value = 0.003) between the two groups. Primary 1 (1.9) 2 (3.8) 3 (2.8) Secondary 16 (30.2) 11 (20.8) 27 (25.5) Tertiary 35 (66.0) 39 (73.6) 74 (69.8) Total (n) 53 53 106 Ethnicity Yoruba 45 (84.9) 43 (81.1) 88 (83.0) 1.137f 0.822 Hausa 2 (3.8) 1 (1.9) 3 (2.8) Igbo 2 (3.8) 3 (5.7) 5 (4.8) Others 4 (7.5) 6 (11.3) 10 (9.4) Total (n) 53 53 106 χ2: Chi square; t: Independent Samples T test; f: Fisher’s exact test Figure 1: Types of infertility among the participants Table 2: Symptoms found in women with infertility (patients) and the control (fertile) group Variables Patient n (%) Control n (%) Total n (%) χ2 p-value Anterior neck swelling No 53 (100.0) 53 (100.0) 106(100.0) Palpitations Yes 2 (3.8) 3 (5.7) 5 (4.7) 0.210f 1.000 No 51 (96.2) 50 (94.3) 101 (95.3) Weight loss Yes 1 (1.9) 0 (0.0) 1 (0.9) 1.010f 1.000 No 52 (98.1) 53 (100.0) 105 (99.1) History of abnormal weight gain Yes 4 (7.5) 2 (3.8) 6 (5.7) 0.707f No 49 (92.5) 51 (96.2) 100 (94.3) 0.678 Previous miscarriage Yes 16 (30.2) 4 (7.5) 20 (18.9) 8.874 No 37 (69.8) 49 (92.5) 86 (81.1) 0.003* Heat or cold intolerance Yes 2 (3.8) 0 (0.0) 2 (1.9) 2.038f No 51 (96.2) 53 (100.0) 104 (98.1) 0.495 Decreased libido Yes 4 (7.5) 1 (1.9) 5 (4.7) 1.889f No 49 (92.5) 52 (98.1) 101 (95.3) 0.363 χ2: Chi square test; f: Fisher’s exact test; *: p-value <0.05 Pa ge 14 6 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. Innov. 2(2) 141-152, 2023 Comparison of the Serum Levels of Thyroid Hormones and Thyroid Dysfunctions in the Infertile Patients and the Control (Fertile) Group Table 3 compares the serum levels of thyroid hormones of infertile and fertile women. The mean TSH, fT3 and fT4 levels were higher in the infertile women than in the fertile women with mean values of 1.35±1.65 versus 0.85±1.08 mIU/mL for TSH, 2.79±1.51 versus 2.19±1.15 pg/mL for fT3, and 1.15±0.33 versus 0.99±0.29 ng/dL for fT4. A significant difference was only in fT3 (p-value = 0.023) and fT4 (p-value = 0.009). The median for TSH, fT3 and fT4 was higher in the infertile group than in the fertile group, although only significant in fT3 (p-value = 0.031) and fT4 (p-value = 0.002). The lower and upper limits of the three parameters were within the normal reference range. Table 3: Comparison of thyroid hormone levels of patients with infertility and the control (fertile) group Variables Patients Control U p-value TSH (mIU/mL) Mean ± SD 1.35±1.65 0.85±1.08 1149.000 0.064 Median (IQR) 0.80 (0.40-1.55) 0.60 (0.40-0.85) 0.105 fT3 (pg/mL) Mean ± SD 2.79±1.51 2.19±1.15 1062.500 0.023* Median (IQR) 2.40 (1.45-4.20) 1.70 (1.35-2.95) 0.031* fT4 (ng/dL) Mean ± SD 1.15±0.33 0.99±0.29 924.000 0.009* Median (IQR) 1.10 (1.00-1.30) 1.00 (0.90-1.10) 0.002* U: Mann-Whitney U test; *: p-value <0.05 Table 4: Prevalence of thyroid disorders among the infertile women (patients) and the control (fertile) group Variable Patient n (%) Control n (%) Total n (%) χ2 p-value OR (95% CI) Thyroid disorders Present 8 (15.1) 2 (3.8) 10 (9.4) 3.975f 0.046* 4.533 (0.915 – 22.465) Absent 45 (84.9) 51 (96.2) 96 (90.6) χ2: Chi square; OR: Odds ratio; 95% CI: 95% Confidence interval; *: p- value <0.05 Table 4 demonstrates the prevalence of thyroid disorders among the infertile women (patients) and the control (fertile) group. Eight (15.1%) patients had thyroid disorders compared to two (3.8%) in the control group, a difference statistically significant (p-value = 0.046). Forty- five (84.9%) infertile and 51 (96.2%) fertile women were euthyroid. Table 5 compares the pattern of thyroid disorders among infertile and fertile women. Four (7.5%) of the infertile women had subclinical hypothyroidism while one (1.9%) of the fertile women had subclinical hypothyroidism, and the difference was not significant (p-value = 0.37). Overt i.e. clinical hypothyroidism was present in two of the infertile patients but was absent in the fertile women. Overt i.e. clinical hyperthyroidism was present in two of the infertile women while only one of the fertile women had it, and the difference was not significant (p-value = 1.000). Majority of the study participants were euthyroid – as observed in 45 (84.9%) of the infertile women as against 51 (96.2%) in the fertile women. Table 6 shows the association between thyroid dysfunction and menstrual cycle abnormality. Five (62.5%) out of eight infertile women with thyroid dysfunction had menstrual irregularities, and others (37.5%) had normal menses. Thirteen (28.9%) out of forty-five euthyroid infertile women had menstrual irregularities, and others (71.1%) had normal menses. Of the two fertile women with a thyroid disorder, one had a menstrual anomaly and the others did not. Six (11.8%) of the fifty-one euthyroid fertile women had menstrual irregularities, while others (45) did not. These differences were not statistically significant (p-value >0.05). Table 5: Pattern of thyroid dysfunction among infertile women and control (fertile) women Thyroid disorder Patients n (%) Control n (%) Total n (%) χ2/ t p-value Subclinical hypothyroidism 4 (7.5) 1 (1.9) 5 (4.7) 0.800 0.371 Overt (clinical) hypothyroidism 2 (3.8) 0 (0.0) 2 (1.9) NA Overt (clinical) hyperthyroidism 2 (3.8) 1 (1.9) 3 (2.8) 0.000 1.000 None 45 (84.9) 51 (96.2) 96 (90.6) 0.260 0.610 Total 53 53 106 χ2: Chi-square; NA: Chi-square not available because of zero ‘0’ value Pa ge 14 7 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. Innov. 2(2) 141-152, 2023 Table 6: Association between thyroid dysfunction and menstrual cycle abnormality Thyroid disorder Menstrual abnormality Present Absent Total χ2 p-value OR (95% CI) n (%) n (%) n (%) Patient Present 5 (62.5) 3 (37.5) 8 3.421f 0.104 4.103 (0.854 – 19.716) Absent 13 (28.9) 32 (71.1) 45 Total 18 35 53 Control Present 1 (50.0) 1 (50.0) 2 2.454 0.249f 7.500 (0.413 - 136.271) Absent 6 (11.8) 45 (88.2) 51 Total 7 46 53 χ2: Chi square; OR: Odds ratio; 95% CI: 95% Confidence interval Undiagnosed and untreated thyroid disease can be a cause of infertility as well as sub-fertility which have important medical, economic, and psychological implications in our society (Verma, 2012). Aside from infertility, thyroid dysfunction could cause other reproductive disorders such as abnormal sexual development or menstrual irregularities. Even in fertility, pregnancy can be complicated by anemia due to SCD and despite the significant need for effective treatment options for SCD patients, current treatments both traditional and newly developed, only ameliorate acute and chronic SCD manifestations without addressing the underlying cause (Musa, 2023). Of the infertile women, 75.5% had secondary infertility and 24.5% primary infertility. This is consistent with the findings of (Panti, 2014) which reported that 67.2% infertile female patients had secondary infertility and 32.8% had primary infertility. This dominance of secondary infertility in this study agrees with previous studies in Africa (Bala, 2003; Ekanem, 2006; Obuna, 2012; Okonofua, 2003), but contrasts the finding in most Western societies where primary infertility dominate with 63.1-80.0% (Habbu, 2016; Nasir, 2016). In this study, 30.2% of infertile patients had previous miscarriages, a value close to the 37% reported by (Orazulike, 2018). Previous miscarriage was the only pregnancy outcome reported in their study by its design. It was also reported by the patient with some possible recall bias. Notwithstanding, this suggests that subfertility and pregnancy wastage are associated with thyroid disorders in women. Other symptoms presented by the subjects were palpitation (3.8%), weight loss (1.9%), abnormal weight gain (7.5%), heat/cold intolerance (3.8%), and decreased libido (7.5%). None of the 106 participants had anterior neck swelling. The mean serum levels of thyroid hormones (TSH, fT3 and fT4) in this study were within the normal reference range although higher in the infertile women. The overall prevalence of thyroid disorders was 15.1% and 3.8% in the infertile and fertile women respectively. A similar study done in Port-Harcourt, Nigeria by (Orazulike, 2018) found a much lower prevalence of thyroid disorders of 4.6% in infertile women. Similar studies done in India by (Rahman, 2008) and another by (Rijal, 2011) reported a prevalence of 25.6% and 33.0% respectively. Similarly, (Habbu, 2016), (Sharma, 2012), and (Shivaleela, 2012) reported higher prevalence rates (between 23.0- 42.3%) of thyroid disorders in infertile women. Thyroid hormone synthesis is influenced by iodine and it varies from clime to clime depending on water, soil, diet and fortification (Utiger, 2006). With this knowledge, the difference in the prevalence of thyroid disorders in these studies can therefore be related to the variation in the study particpants’ consumption of iodine-containing meals (Elahi, 2007). TSH was higher, but fT3 and fT4 levels were significantly higher when serum levels of thyroid hormones of the patients are compared with that of the control group. (Habbu, 2016) and (Shivaleela, 2012) also reported findings of elevated mean serum fT3 and fT4 but low TSH in the infertile patients. (Orazulike, 2018) found no significant difference in the mean serum fT3, fT4, TSH, and TPOAb between patients and controls. These findings were different from the findings of (Fatima, 2014) where serum fT3 and fT4 were significantly decreased and serum TSH was significantly increased in infertile females when compared with fertile females. Serum fT3 and fT4 may be higher where a large proportion of the patients are hyperthyroid and the reverse may be the case when a large proportion of the patients are hypothyroid. On types of thyroid dysfunction in this study, 9.4% had one type of thyroid dysfunction and 90.6% were euthyroid. In agreement with this, (Elahi, 2007) reported that 89.3% infertile patients and 93.4% fertile control were euthyroid. This finding is consistent with that of (Goswami, 2009), (Orazulike, 2018), and (Rijal, 2011). With infertile women being euthyroid, this suggests that other cause(s) of infertility aside from thyroid disorders should be evaluated during infertility examination as there can be significant single or multiple causes of infertility. Subclinical hypothyroidism was the most predominant thyroid disorder in this study. A study done in Kano Pa ge 14 8 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. Innov. 2(2) 141-152, 2023 Nigeria by (Emokpae, 2011) got a higher prevalence (14.9%) for subclinical hypothyroidism as against the 9.4% in this present study. This may be because Emokpae’s study assessed only TSH in hyperprolactinemic women focusing only on subclinical thyroid diseases while overt (clinical) thyroid diseases were not considered at all. Hypothyroidism is associated with increased production of TRH, which stimulates the anterior pituitary to secrete TSH and prolactin. Hyperprolactinemia adversely affects fertility potential by impairing GnRH pulsatility and thereby affecting ovarian function (Poppe, 2003). Therefore, it is advisable to check TSH and PRL levels in every infertile female, regardless of their menstrual rhythm (Abdul, 2015). Moreover, concerning subclinical hypothyroidism (Poppe, 2003) reported a prevalence of 0.9% and (Bohnet, 1981) reported a prevalence of 11%. The prevalence of subclinical hypothyroidism in subfertile women has been reported to vary from 0.7% to 43% (Poppe, 2007). This wide range is due to the differences in the sensitivity of serum TSH measurement. The revised clinical practice guidelines of the Endocrine Society recommend the measurement of serum TSH in women over the age of 30 years with infertility or a prior history of miscarriage, to screen for thyroid dysfunction (De Groot, 2012). In clinical practice guidelines for hypothyroidism in adults, the American Association of Clinical Endocrinologists (AACE) and the American Thyroid Association (ATA) have recommended that treatment with L-thyroxine should be considered in women of childbearing age with subclinical hypothyroidism when they are planning a pregnancy (Garber, 2012). If infertile women planning pregnancy are diagnosed with subclinical hypothyroidism as well as overt (clinical) hypothyroidism, they are recommended to be treated to reduce risks of miscarriage and fetal developmental impairment or to improve the in vitro fertilization outcome (Lee, 2014). Evidence suggests that treating thyroid disorders and keeping TSH levels below 2.5 mIU/L may improve conception rates in infertile women and reduce early pregnancy loss (Garber, 2012). In the study by (Bohnet, 1981), subclinical hypothyroidism was considered an infertility factor by itself because treatment with L-thyroxine 50 mg/day normalized the mid-progesterone secretion and two out of the eleven treated women became pregnant. Thyroid disorders have a known association with menstrual irregularities which may lead to infertility (Krassas, 1990). In this study, menstrual irregularities were significantly more predominant in the infertile patients than in the fertile control and 62.5% of the infertile patients with thyroid disorders had menstrual irregularities. (Orazulike, 2018) reported menstrual irregularities in 50% of the patients. (Nasir, 2016) also reported menstrual irregularities in 19.6% of the patients. (Goswami, 2009) similarly reported menstrual irregularities in 61.2% of the infertile patients and that 50% infertile patients with hypothyroidism had menstrual irregularities. The impact of hypothyroidism on ovulation and menstrual function is related to numerous interactions of thyroid hormones with the female reproductive system, in turn causing infertility. In hypothyroidism, increased TRH production leads to hyperprolactinemia and altered gonadotropin-releasing hormone (GnRH) pulsatile secretion. This leads to a delay in luteinizing hormone (LH) response and inadequate corpus luteum leading to abnormal follicular development and ovulation. At the cellular level, thyroid hormone receptors are expressed in human oocytes and granulosa cells, and the hormones synergize with the follicle-stimulating hormone-mediated luteinizing hormone/human chorionic gonadotropin (hCG) receptor to exert direct stimulatory effects on granulosa cell function of progesterone production (Poppe, 2004). Altering the peripheral metabolism of estrogen and decreasing sex hormone-binding globulin (SHBG) production is another pathway by which hypothyroidism may impact on fertility. These pathways may result in abnormal feedback at the pituitary level and consequently infertility (Bassey, 2015). Several aspects of the reproductive axis influenced by hyperthyroidism are comparable with the situation in hypothyroid women. In hyperthyroidism, the SHBG production, the conversion of androgens to estrogens, and the gonadotrophin response to GnRH are increased. The decrease in menstrual flow may also be related to effects on hemostatic factors, including the synthesis of factor VIII (Krassas, 2000). Women with hyperthyroidism and fertility problems should be treated with antithyroid drugs and/or surgery according to the cause of hyperthyroidism (Krassas, 2000). CONCLUSION Serum levels of thyroid hormones and the prevalence of thyroid disorders were significantly higher in the infertile patients when compared with the fertile women. Although the overall prevalence was quite low, thyroid abnormalities could still be implicated as a cause of female infertility. It is recommended that all infertile women should have thyroid function tests as part of their infertility work-up as many patients with thyroid diseases can easily be missed if they are not screened. Direction for future research on this topic would be to evaluate the menstrual pattern and irregularities and relate these to thyroid disorder patterns among women within this study setting. This will give a clearer picture of the relationship between symptoms of menstrual irregularities and thyroid dysfunction which varies from region to region. Acknowledgments Special thanks to all the infertile and fertile women who voluntarily participated in this study after making an informed decision. Authors’ Contribution All authors contributed in different aspects of the research. Pa ge 14 9 https://journals.e-palli.com/home/index.php/ajmsi Am. J. Med. Sci. 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