Vitamin D Deficiency Among Subfertile Women: Case Study

Vitamin D Deficiency Among Subfertile Women: Case Study Abstract Objective: To compare the dietary vitamin D and calcium intake among subfertile women (cases) versus pregnant women (controls) and to determine the vitamin D levels in the subfertile and pregnant women Study design was an observational case-control study where a total of 181 (83 previously diagnosed subfertile cases from various causes and 98 pregnant controls) women of reproductive age (20-40 years old) attending the reproductive endocrinology and infertility clinics were recruited. Ethical approval was granted from the Institutional Review Board at KFMC and written informed consent was obtained from each participant prior to inclusion. A validated questionnaire was used where it focused on key indicators evaluating vitamin D related factors. Blood was withdrawn for the measurement of serum calcium, albumin and phosphate to exclude secondary causes that might affect Vitamin D level. Data Analysis Data was analyzed using SPSS version 16.0 (SPSS, Chicago, IL, USA). Frequencies were presented as percentages (%) and continuous variables were presented as mean  ± standard deviation. Chi-square was done to elicit differences in frequencies between cases and controls. Independent T-test was done to compare differences among continuous variables. Significance was set at p < 0.05. Results: The prevalence of vitamin D deficiency was significantly higher in the subfertile group than controls (59.0% versus 40.4%; p < 0.01). Calcium supplements intake was significantly higher in controls than the subfertile group (64.6% versus 10.0%; p-value < 0.001). Total dietary vitamin D intake (> 400IU/day) was significantly higher in the controls than the subfertile group. Conclusions: Total dietary vitamin D intake (> 400IU/day) was found to be significantly higher in the controls than the subfertile group. Thus, dietary vitamin D intake greater than 400IU/day and calcium supplementation should be encouraged amongst subfertile women since it may improve their fertility, but this needs to be proven prospectively. Optimization of serum calcium and vitamin D levels are encouraged. Key Words: Vitamin D, infertility, calcium, supplementation, Middle Eastern Condensation Dietary vitamin D intake and calcium supplementation may improve fertility. Optimization of serum calcium and vitamin D levels are encouraged. Introduction Vitamin D deficiency phenomenon over the recent years has earned the notorious reputation of being significantly associated to a plethora of many diseases, making it a major global public health burden [1]. Evidence that vitamin D is also associated with the reproductive physiology is just recently accumulating yet limited to animal models and very few human studies [2]. Animal and human studies have suggested vitamin D role in the reproductive system where vitamin D has recently been described to modulate reproductive processes in women and men [3]. Studies have found the presence of vitamin D receptor (VDR) and vitamin D metabolizing enzymes in reproductive tissues of women [3]. Additionally, low 25-hydroxyvitamin D (25(OH)D) levels have been associated with obesity, metabolic, and endocrine disturbances in PCOS women and vitamin D supplementation has been suggested to improve menstrual frequency and metabolic disturbances in those women [3]. Moreover, vitamin D might influence ste roidogenesis of both estradiol and progesterone in healthy women where low levels of 25(OH)D levels might be associated with infertility and high levels might be associated with endometriosis [4]. The most up to date vitamin D studies outside its conventional role of calcium homeostasis in the Middle East so far have covered most of the obesity-related diseases including diabetes mellitus, hypothyroidism, and the full metabolic syndrome [5-9]. Despite the abundance and overflow of recent literature with respect to vitamin D role in health and disease, there are an equal overwhelming number of questions left unanswered and this probably explains the rationale to conduct more research on vitamin D, namely the reported association between vitamin D deficiency and infertility. In brief, the target effects of vitamin D on certain reproductive parameters including pregnancy, spermatogenesis, implantation and progeny have been studied in both animal and human studies [10,11,12,13], whereas folliculogenesis and steroidogenesis were only proven in animal models [11]. Studies involving humans have so far pointed the vitamin D role in the endometrial regulation [11, 12] rather than folliculogenesis [13, 14]. In the Middle East where most women of reproductive age are vitamin D deficient, follicular fluid has been observed to be reflective of vitamin D status, but does not have a role in assisted reproductive technology [15]. A recent large, prospective, cross-sectional study has shown that low serum levels of vitamin D may impair a woman's chances of conceiving through in vitro fertilization (IVF) [14]. The authors concluded that a serum vitamin D level of 20 ng/mL or more was associated with a significantly greater chance of obtaining 3 or more high-quality embryos and with successful embryo implantation and clinical pregnancy compared with lower levels [14,16]. Given the novel and emerging role of vitamin D in the field of reproductive biology, the present observational study aims to determine on a nutritional level, whether differences exist in the dietary intake of vitamin D among sub-fertile Middle-Eastern women attending in-vitro fertilization (IVF) clinics versus pregnant women which was assessed by 25 (OH) D serum level. Material and Methods A total of 181 (83 previously diagnosedsubfertile cases from various causes and 98 pregnant controls) Saudi women of reproductive age (20-40 years old) attendingthe reproductive endocrinology and infertility clinics at King Fahad Medical City (KFMC), Riyadh, Saudi Arabia from August 2012 to January 2013 were recruited for this observational case-control study. Subjects requiring immediate medical attention and those with co-morbidities such as malignancies, chronic infection, chronic and cardiovascular-related diseases were excluded. Ethical approval was granted from the Institutional Review Board at KFMC and written informed consent was obtained from each participant prior to inclusion. Data Collection Variables include demographics (age and weight), subfertility types and causes. In addition, dichotomous variables (yes or no) on what type of vitamin D-rich food sources were also collected as well as data on other vitamin D related factors such as sunlight exposure for at least 10 minutes a day, use of Vitamin D supplement, and intake of Vitamin D rich food. A validated questionnaire was used to capture from recruited patients. It focused on key indicators evaluating vitamin D related factors. These indicators were adopted from pre-tested patient questionnaires used in various accredited hospitals. Our questionnaire was validated bybenchmarking against similar ones in the literature targeting diverse populations and using different approaches. Initially, the questionnaire was tested on 20 patients from KFMC to determinewhether the questions were clear, understandable, and in a logical order (face validity). Besides, 2 health professionalswho had experience in clinical research were asked to criticize the content of the questionnaire (content validity). Blood was withdrawn for the measurement of serum calcium, albumin and phosphate to exclude secondary causes that might affect Vitamin D level using routine laboratory methods. Serum 25(OH)vitamin D was also assessed using Cobas e411 (Roche Diagnostics, Mannheim, Germany). For the purpose of this study, vitamin D deficiency was defined as serum 25(OH)vitamin D level less than 20ng/ml [2]. Data Analysis Data was analyzed using SPSS version 16.0 (SPSS, Chicago, IL, USA). Frequencies were presented as percentages (%) and continuous variables were presented as mean  ± standard deviation. Chi-square was done to elicit differences in frequencies between cases and controls. Variables exhibiting non-Gaussian distribution were logarithmically transformed prior to analysis. Independent T-test was done to compare differences among continuous variables. Multinomial logistic regression analysis was done using the grouping variable (presence or absence of subfertility) as the dependent variable and dietary vitamin D intake > 400IU/day as independent variables with obesity, sunlight exposure and skin color as co-variates. Significance was set at p < 0.05. Results One hundred and eighty one patients were analyzed, 83 cases and 98 controls. There was no statistical difference between the mean age of cases ,29.5  ± 5.5 years, verses controls, 29.9  ± 5.2 (P=0.64).Vitamin D deficiency (< 20ng/ml) was observed in 48.9% of the entire cohort. Comparatively, the prevalence of vitamin D deficiency was significantly higher in the subfertile group than controls (59.0% versus 40.4%; p < 0.01). On the other hand, calcium supplements intake was significantly higher in controls than the subfertile group (64.6% versus 10.0%; p-value < 0.001). The controls significantly took more vitamin D supplements than the subfertile group (p < 0.05). The rest of the comparisons were unremarkable (Table 1). Table 2 shows the percentage differences in the vitamin D dietary intake of both controls and the subfertile group. The controls had significantly higher percentage consumption of food than the subfertile group in 9 out of 14 vitamin D dietary sources included in the study (wild salmon, farmed salmon, mushrooms, egg yolk, fortified milk, fo rtified orange juice, yogurts, butter and breakfast cereals; all p-values < 0.001). Consequently, total dietary vitamin D intake > 400IU/day was significantly higher in the controls than the subfertile group (p < 0.001) (Table 2). Table 3 shows the metabolic characteristics of subjects. There was a modest, borderline significance in BMI and vitamin D status between groups, with controls being higher than the subfertile group (p-values 0.051 and 0.08, respectively). The control group also had significantly higher levels of corrected calcium (p < 0.001) while the infertile group had significantly higher levels of circulating albumin (p < 0.001) (Table 3). Comments This study addresses the possible protective effects of increased dietary vitamin D intake and calcium supplementation among Middle Eastern women that harbor infertility conditions. Previous studies in Saudi Arabia considered obesity as a major risk factor for infertility [17]. Consequently, obesity is a major risk factor for vitamin D deficiency and both abnormal metabolic states can be manipulated through dietary interventions [18]. The issue of vitamin D status correction to improve fertility status among vitamin D deficient women of reproductive age may still require rigorous investigation since randomized trials are scarce. Nevertheless, several studies can already attest that micronutrient supplementation including vitamin D improves fertility outcomes [19,20]. Furthermore, available evidence confirms present findings in the beneficial effects of vitamin D in female infertility, with favorable outcomes among women with endometriosis and polycystic ovarian syndrome [4,22]. Calcium supplementation to improve female fertility is not a surprise since it is directly regulated by vitamin D, receptors of which are found in various reproduction tissues, and correction of levels confer positive effects in terms of follicle maturation, menstrual regularity and improvement of hyperandrogenism especially among women with polycystic ovarian syndrome (PCOS) [21,22]. The other equally important but less novel finding in the present study is the high vitamin D deficiency prevalent in both pregnant and sub fertile women. Vitamin D deficiency is relatively common in the Middle East in general, but more so for women because of cultural traditions and manner of clothing. Prevalence of hypovitaminosis D among this group ranges from 80-100% from the most recent local studies [23,24]. The modest but still insignificant difference in mean vitamin D status levels in both groups in our study could be attributed to a more vigilant behavior among pregnant women to ingest more food and vitamin supplements although comparison by prevalence clearly shows the high significant difference between the subfertile group and the control group. The authors acknowledge several limitations. Findings of the present study may only be true for women and as such a separate study should be done to reinforce if not negate the present results in non-pregnant women in comparison to subfertile women. Furthermore the case control nature of the study maybe subject to selection bias. Prospective studies are essential to determine whether vitamin D correction through dietary intervention and lifestyle modification can improve fertility among vitamin D deficient women. In conclusion, Middle-Eastern (Saudi) subfertile women have a higher prevalence of vitamin D deficiency as compared to apparently healthy controls, and this is aggravated by their lower intake of foodrich in vitamin D and lower calcium supplementation. Dietary vitamin D intake greater than 400IU/day and calcium supplementation may improve fertility in this population, but this needs to be proven prospectively. Optimization of vitamin D and calcium status among women of reproductive age are encouraged.