European Journal of Obstetrics & Gynecology and Reproductive Biology 159 (2011) 132–137

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European Journal of Obstetrics & Gynecology and

Reproductive Biology
journal homepage: www.elsevier.com/locate/ejogrb

Predictive value of the level of vitamin D in follicular fluid on the outcome

of assisted reproductive technology
Ashraf Aleyasin a, Marzieh Agha Hosseini a, Atossa Mahdavi a,*, Leila Safdarian a, Parvin Fallahi a,
Mohammad Reza Mohajeri b, Mohammad Abbasi b, Fatemeh Esfahani c
a
Department of Infertility, Shariati Hospital, Tehran University of Medical Sciences, North Karegar Street, Tehran 1411713135, Iran
b
Endocrine and Metabolism Research Centre, Shariati Hospital, Tehran University of Medical Sciences, Tehran, Iran
c
Shariati Hospital, Tehran University of Medical Sciences, Tehran, Iran

A R T I C L E I N F O A B S T R A C T

Article history: Objective: To assess the correlation between the levels of vitamin D in follicular fluid and serum, and to
Received 19 December 2010 determine whether the level of 25-hydroxyvitamin D (25OH-D) in follicular fluid of infertile women
Received in revised form 9 May 2011 undergoing assisted reproductive technology (ART) is associated with the outcome.
Accepted 11 July 2011
Study design: Eighty-two infertile women undergoing ART at an academic tertiary care centre were
recruited for a prospective cohort study. Levels of 25OH-D in follicular fluid and serum were measured.
Keywords: Standardized regimens for pituitary downregulation and controlled ovarian hyperstimulation were
Vitamin D
employed. Patient and cycle parameters, and clinical pregnancy (defined as evidence of intra-uterine
Infertility
Pregnancy
gestation sac plus heart rate on ultrasound) were determined.
Follicular fluid Results: A significant correlation was found between the levels of vitamin D in follicular fluid and serum
Intracytoplasmic sperm injection (r = 0.767, p = 0.001). The overall rates of chemical, clinical and ongoing pregnancy were 35.5% (n = 29),
29.3% (n = 24) and 23.2% (n = 19), respectively. No significant difference was found in these pregnancy
rates between the tertiles of 25OH-D level in follicular fluid (p = 0.959, 0.995 and 0.604, respectively).
The median serum level of vitamin D was 8.13 (range 5.37–13.62) ng/ml in the clinically pregnant group
and 8.29 (range 5.93–21.23) ng/ml in the non-pregnant group (p = 0.235). Interestingly, the median level
of vitamin D in follicular fluid was 9.19 (range 5.25–19.51) ng/ml in the clinically pregnant group and
10.34 (range 5.89–29.69) ng/ml in the non-pregnant group (p = 0.433). The fertilization rate decreased
significantly and the implantation rate increased (not significantly) with increasing tertiles of 25OH-D
level in follicular fluid.
Conclusions: The level of 25OH-D in follicular fluid is reflective of body stores of vitamin D. Most subjects
in this study were deficient in vitamin D, but this study found that vitamin D deficiency does not play a
pivotal role in the outcome of ART.
ß 2011 Elsevier Ireland Ltd. All rights reserved.

1. Introduction and fertility capacity, diminished mating success, impaired

The well-established function of the steroid hormone vitamin D
is to maintain calcium and phosphorus homeostasis, and to
promote bone mineralization. Since this vitamin can have a
significant influence on the growth and differentiation of a variety
of tissues, and decreases the risk of many chronic illnesses
including common cancers, autoimmune diseases, infectious
diseases and cardiovascular disease [1–7], it has become the focus
of many studies in recent years. The discovery that most tissues
and cells in the body have a vitamin D receptor has provided new
insights into the function of this vitamin [1]. Reduced fertility rates
* Corresponding author. Tel.: +98 21 88008810; fax: +98 21 88220050.
E-mail address: atossa_mahdavi@yahoo.com (A. Mahdavi).
neonatal growth, increased pregnancy complications, gonadal
insufficiency, reduced aromatase gene expression, low aromatase
activity, hypogonadism, bone malformations, uterine hypoplasia,
impaired folliculogenesis and infertility have been reported in
vitamin-D-deficient animal models [8–10]. Also, it has been
reported that vitamin D affects placental steroidogenesis, placental
calcium transport, expression of placental lactogen and decid-
ualization of the endometrium [2,11,12].
Although the biological actions of vitamin D are mediated
through a member of the steroid nuclear hormone receptor
superfamily but there is a debate about the mechanism of
reproductive function of this vitamin [10,13–15]. While some
attributed the direct effect of vitamin D but others realize that
hypocalcemia associated with vitamin D deficiency is responsible
for reduced fertility in vitamin D–deficient animals [16].

0301-2115/$ – see front matter ß 2011 Elsevier Ireland Ltd. All rights reserved.
doi:10.1016/j.ejogrb.2011.07.006
 A. Aleyasin et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology 159 (2011) 132–137 133

Due to limited specific human data in this regard and the
epidemic of vitamin D deficiency in Iran [17], this study was
undertaken to evaluate the predictive value of the level of vitamin
D in follicular fluid on assisted reproductive technology (ART)
outcome in infertile patients.
2. Materials and methods
A prospective cohort study was undertaken at Shariati Hospital,
Tehran, Iran. In total, 82 infertile women undergoing ART were
enrolled between August 2009 and March 2010. The study protocol
was in accordance with the guidelines of the Declaration of
Helsinki, and was approved by the Institutional Review Board of
the Infertility Department and Deputy of Research affiliated to
Tehran University of Medical Sciences.
Women >38 years of age, with systemic illness, consuming
drugs interfering with vitamin D metabolism, with hypothalamic
amenorrhoea or galactorrhoea were excluded from the study.
Couples requiring testicular sperm extraction and aspiration for
sperm recovery were also excluded.
After complete desensitization (long-term protocol) using the
gonadotrophin-releasing hormone agonist, buserelin (Daroo Sazi
Samen Co., Iran), ovarian stimulation with Gonal F (Serono,
Switzerland) was commenced on Day 3 of the next cycle. The
starting dose was selected on the basis of age, weight, early
follicular phase levels of follicle-stimulating hormone and the
number of antral follicles. Transvaginal ultrasound (Sonoline G20,
Siemens Medical Solutions USA, Inc., Mountain View, CA, USA) was
performed every 3–5 days to study follicular development. Final
oocyte maturation was triggered when at least two follicles with
diameter 17 mm were observed, with 10,000 IU human chorionic
The level of parathyroid hormone in peripheral plasma was
estimated using enzyme-linked immunosorbent assay (ELISA;
Immunodiagnostic Ltd). The interassay coefficient of variation was
4.23% (<6.7%). Intra-assay coefficients of variation were 1.5% for
the first control and 2% for the second control, respectively (<4.4%).
The oestradiol level was measured using ELISA (Germany).
Serum samples for calcium, phosphorus, alkaline phosphatase,
vitamin D and parathyroid hormone were assayed using a
photometric authoanalyser (Hitachi 902, Japan).
2.2. Statistical analysis
The sample size was calculated to show if there was a linear
correlation of at least 0.85 between serum and follicular fluid levels
of vitamin D, with a level of significance of 0.05. Considering a
clinical pregnancy rate of 30%, this sample size would also be
sufficient to detect a difference of 20 mmol/l in the mean level of
vitamin D in follicular fluid between pregnant and non-pregnant
women, assuming a standard deviation (SD) of 30 mmol/l, with
power of 80% and a significance level of 5%.
Statistical Package for the Social Sciences Version 12 (SPSS,
Chicago, IL, USA) was used for data analysis. The normality of
distribution was checked using the Kolmogorov–Smirnov test.
According to the distribution of continuous data, they are
presented as mean  SD or median (range). Correlation analysis
was performed using Spearman’s rho test. Pregnant and non-
Table 1
Basal and clinical characteristics of all the patients.
n = 82

gonadotrophin (hCG; Ferring Co., Germany) administered as a Female age (years), mean  standard deviation 29.81  4.25
single intramuscular injection. Serum samples (calcium, phospho- Male age (years) 33 (24–51)
Duration of infertility (years) 7 (1–20)
rus, alkaline phosphatase, vitamin D and parathormone) were
Type of infertility, n (%)
collected on the day of ovum pick up and stored at 20 8C prior to Primary 60 (73%)
assay. Oocytes were collected 36–38 h later using transvaginal- Secondary 22 (27%)
guided follicle aspiration under general anaesthesia. Follicular Gravidity 0 (0–3)
fluid was collected from follicles with diameter 14 mm following Number of previous ART cycles 0 (0–5)
Polycystic ovarian syndrome, n (%) 17 (20.7%)
oocyte isolation, after which the fluid should be clear and not Female body mass index (kg/m2) 26.76 (17.80–35.80)
contaminated with blood. Follicle-stimulating hormone (mIU/ml) 6.15 (1.4–14)
The follicular fluid for each patient was pooled and centrifuged Luteinizing hormone (mIU/ml) 4.8 (4–21)
at 3000  g for 15 min, and the supernatant was stored at 80 8C Oestradiol (pg/ml) 44.50 (5–300)
Antral follicle count 11 (5–20)
prior to assay. After fertilization through intracytoplasmic sperm
Total good sperm (million) 4 (0.1–60)
injection, up to three good-quality embryos were transferred Stimulation days 0 (8–17)
transcervically 3 days later. Luteal phase support was started the Gonadotropin injections 35 (16–104)
day after ovum pick up by daily administration of the progesterone Oestradiol on hCG day (pg/ml) 1822.5 (606–6100)
suppository Cyclogest (Actavis, UK) 800 mg. Pregnancy was Total oocytes retrieved, n (%) 11 (3–24)
Mature oocytes 7 (1–20)
detected by serum beta-hCG analysis 14 days after embryo Pronuclear number 6 (1–15)
transfer, and a transvaginal ultrasound scan was scheduled for 2 Embryos transferred 4 (1–5)
weeks later to detect the intra-uterine gestation sac. Each pregnant Having freezed embryos, n (%) 32 (39%)
woman was followed by ultrasound scan until the fetal heart was Ovarian hyperstimulation syndrome 4 (4.8%)
(mild–moderate), n (%)
documented (clinical pregnancy), and was followed until 20 weeks
Calcium (mg/dl) 9.5 (8.2–10.2)
of gestation (ongoing pregnancy). Phosphorus (mg/dl) 3.2 (2–4.9)
Alkaline phosphatase (U/l) 144 (81–350)
2.1. Hormone assay Parathormone (pmol/l) 2.1 (0.5–4.8)
Vitamin D in serum
nmol/l 21 (13.40–53)
Levels of 25OH-D in peripheral plasma and follicular fluid were ng/ml 8.41 (5.37–21.33)
estimated via enzyme immunoassay (Immunodiagnostic Ltd, UK, Vitamin D in follicular fluid
USA, Germany, France, Scandinavia). The interassay coefficient of nmol/l 24.6 (13.1–74.1)
variation was 7.9% (<8.7%). Intra-assay coefficients of variation ng/ml 9.86 (5.25–29.67)
were 3.4% for the first control and 2.1% for the second control, ART, assisted reproductive technique; hCG, human chorionic gonadotropin.
respectively (<6.7%). Follicle-stimulating hormone, luteinizing hormone and oestradiol were measured
Based on previously defined serum criteria [1], 25OH- on Day 3 of the cycle.
Calcium, phosphorus, alkaline phosphatase, parathormone, vitamin D in serum and
D > 30 ng/ml in follicular fluid was taken to reflect ‘sufficient’ vitamin D in follicular fluid were measured on the day of ovum pick up.
vitamin D status. Levels of 20–30 ng/ml and <20 ng/ml were taken Unless otherwise indicated, values are median (range).
to reflect vitamin D insufficiency and deficiency, respectively. p < 0.05.
134 A. Aleyasin et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology 159 (2011) 132–137

pregnant women were compared using independent Student’s t-test The mean levels of 25OH-D in follicular fluid in the lowest to
or Mann–Whitney’s U-test. Chi-squared test and Fisher’s exact test highest tertiles were 7.9 (SD 0.8, median 8.1, range 5.2–8.8), 10.0
were used as indicated to compare qualitative variables. The (SD 0.9, median 9.9, range 8.9–11.5) and 16.1 (SD 5.3, median 13.3,
fertilization and implantation rates were compared between tertiles range 11.6–29.7), respectively. The median levels of 25OH-D in
of 25OH-D level in follicular fluid using Chi-squared test. p < 0.05 was serum in the lowest to highest tertiles were 6.85 (range 5.37–7.41),
taken to indicate statistical significance. 8.41 (range 7.49–9.58) and 11.3 (range 9.62–21.23) ng/ml,
respectively. There was no significant difference between the
3. Results tertiles of 25OH-D level in follicular fluid according to maternal
age, BMI, follicle-stimulating hormone, luteinizing hormone,
This study included 82 infertile women with a mean age of 29.8 oestradiol, stimulation days, total ampoules including total human
(SD 4.2) years. The serum level of 25OH-D was <20 ng/ml in all menopausal gonadotrophin, oocytes retrieved and metaphase II
except one woman. The mean levels of 25OH-D in serum and oocytes.
follicular fluid were 9.1 (SD 2.1, median 8.4) and 11.3 (SD 4.7, The overall frequencies of chemical, clinical and ongoing
median 9.8) ng/ml, respectively (Table 1). pregnancy were 35.5% (n = 29), 29.3% (n = 24) and 23.2%
There was a significant linear correlation between the levels of (n = 19), respectively. There was no significant difference in these
25OH-D in serum and follicular fluid (r = 0.77, p < 0.001) (Fig. 1). pregnancy rates between the tertiles of 25OH-D level in follicular
Also, the serum level of 25OH-D showed a significant linear fluid (p = 0.959, 0.995 and 0.604, respectively) (Fig. 2).
correlation with age (r = 0.28, p = 0.014), but not with body mass Basal characteristics, stimulation parameters and serum
index (BMI) (r = 0.12, p = 0.299). The correlation between the level biochemistry were similar between the comparison groups (Table
of 25OH-D in follicular fluid and age and BMI was not significant 2). The logistic regression model including the variables related to
(r = 0.20, p = 0.068; r = 0.09, p = 0.428, respectively). There was no clinical pregnancy in univariate analysis with PV  0.2 and level of
linear correlation between metaphase II (and/or embryo quality) 25OH-D in follicular fluid did not show the level of 25OH-D in
and levels of 25OH-D in follicular fluid and serum (r = 0.073, follicular fluid to be an independent predictor of clinical
p = 0.517; r = 0.018, p = 0.784, respectively). pregnancy. Women with ongoing pregnancies had median levels

30
Vitamin D in follicular fluid (ng/ml)

25

20

15

10
r=0.77
p=0.001
5
5 10 15 20 25 30
Serum vitamin D (ng/ml)

80

70 70.00
Follicle vitamin D = -3.66 + 1.41 * servitd
2
R =0.77
60 60.00
Follicle vitamin D

Follicle vitamin D

50 50.00

40 40.00

30 30.00

Linear regression with 95%

20 20.00 mean prediction interval

10
10 20 30 40 50 60 20.00 30.00 40.00 50.00
Serum Vitamin D Serum vitamin D

Fig. 1. Level of 25OH-D in follicular fluid provides a reliable reflection of the serum level.
 A. Aleyasin et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology 159 (2011) 132–137 135

Fig. 2. Pregnancy rates for women in each tertile of 25OH-D level in follicular fluid (p = 0.959, 0.995 and 0.604, respectively).

of 25OH-D in serum and follicular fluid of 7.77 (range 5.37–
10.26) ng/ml and 9.07 (range 5.25–12.02) ng/ml, respectively.
Although vitamin D deficiency was detected in both groups, the
fertilization rate was 71.5% among all participitants. The fertiliza-
tion rates were 79.0%, 67.8% and 68.3% in the lowest to highest
tertiles of 25OH-D level in follicular fluid, respectively (p = 0.018).
The fertilization rates across tertiles of 25OH-D level in follicular
fluid were examined using trend analysis in SPSS one-way analysis
of variance, and there was no significant linear trend (p = 0.274) or
quadratic components of trend (p = 0.574) in the data.
The implantation rate was 12.6% in all women, and 9.6%, 13%
and 15.9% in the lowest to highest tertiles of 25OH-D level in

Table 2
Participant and assisted reproductive technology cycle characteristics by cycle outcome.

Clinical pregnancy (n = 24) Not pregnant (n = 53) p-Value

Female age (years), mean  standard deviation 28.67  3.84 30.28  4.35 0.119
Male age (years) 32 (27–50) 34 (24–51) 0.312
Duration of infertility (years) 5.5 (1–20) 7 (1–19) 0.396
Type of infertility, n (%) 0.161
Primary 15 (62.5%) 45 (77.6%)
Secondary 9 (37.5%) 13 (22.4%)
Gravidity 0 (0–2) 0 (0–3) 0.290
Number of previous ART cycles 0 (0–2) 0 (0–5) 0.378
Polycystic ovarian syndrome, n (%) 4 (16.7%) 13 (22.4%) 0.766
Female body mass index (kg/m2) 27.18 (18.29–35.70) 26.70 (17.80–34.29) 0.654
Follicle-stimulating hormone (mIU/ml) 5.35 (1.4–11) 6.8 (2–14) 0.129
Luteinizing hormone (mIU/ml) 4 (1.5–20) 4.8 (4–21) 0.650
Oestradiol (pg/ml) 44 (14–300) 45.5 (5–245) 0.956
Antral follicle count 11 (5–20) 11 (5–20) 0.512
Total good sperm (million) 4 (0.1–30) 4 (4–60) 0.473
Stimulation days 10 (9–17) 11 (8–14) 0.667
Gonadotropin injections 34 (18–104) 36 (16–80) 0.428
Oestradiol on hCG day (pg/ml) 2011 (657–4300) 1760 (606–6100) 0.631
Total oocytes retrieved, n (%) 11 (5–24) 11 (3–23) 0.396
Mature oocytes 8 (3–20) 7 (1–16) 0.070
Pronuclear number 7 (2–15) 6 (1–13) 0.092
Embryos transferred 4 (2–5) 4 (1–5) 0.182
Having freezed embryos, n (%) 12 (50%) 20 (34.5%) 0.190
Ovarian hyperstimulation syndrome (mild–moderate), n (%) 2 (8.3%) 2 (3.4%) 0.289
Calcium (mg/dl) 9.5 (8.2–10) 9.40 (8.60–10.20) 0.931
Phosphorus (mg/dl) 3.2 (2.10–4.80) 3.2 (2–4.90) 0.901
Alkaline phosphatase (U/l) 156 (85–236) 139 (81–350) 0.131
Parathormone (pmol/l) 2.3 (0.7–4.8) 2 (0.5–4.30) 0.159
Vitamin D in serum 0.235
nmol/l 20.3 (13.4–34) 21.45 (14.80–53)
ng/ml 8.13 (5.37–13.62) 8.29 (5.93–21.23)
Vitamin D in follicular fluid 0.433
nmol/l 22.95 (13.10–48.70) 25.80 (14.70–74.10)
ng/ml 9.19 (5.25–19.51) 10.34 (5.89–29.69)

ART, assisted reproductive technique; hCG, human chorionic gonadotropin.

Follicle-stimulating hormone, luteinizing hormone and oestradiol were measured on Day 3 of the cycle.
Calcium, phosphorus, alkaline phosphatase, parathormone, vitamin D in serum and vitamin D in follicular fluid were measured on the day of ovum pick up.
Unless otherwise indicated, values are median (range).
p < 0.05.
136 A. Aleyasin et al. / European Journal of Obstetrics & Gynecology and Reproductive Biology 159 (2011) 132–137
follicular fluid, respectively (p = 0.791). In addition, the implanta-
tion rate across the tertiles of 25OH-D level in follicular fluid was
examined, but no significant linear trend (p = 0.857) or quadratic
components of trend (p = 0.628) were found.
Levels of 25OH-D in serum and follicular fluid were not
significantly different between patients with and without polycystic
ovarian syndrome (PCOS) (p = 0.938 and 0.158, respectively).
4. Discussion
This study found that the level of 25OH-D in follicular fluid is
reflective of body stores of vitamin D. However, most of the
subjects in this study were deficient in vitamin D, and it was not
possible to calculate the effect of vitamin D deficiency on ART
outcome. However, according to the ART results (36% chemical
pregnancy and 30% clinical pregnancy), it is considered that
vitamin D deficiency does not play a pivotal role for the outcome of
ART. The prevalence of vitamin D deficiency found in this study is
in accordance with previous research in Iran [17], but is in contrast
to a study by Ozkan et al. which found significantly higher levels of
vitamin D in follicular fluid in women achieving clinical pregnancy
following in vitro fertilization (IVF) [16].
A more recent study by Anifandis et al. [18] found that higher
levels of vitamin D in combination with decreased levels of glucose
in follicular fluid was associated with a lower likelihood of
pregnancy due to a detrimental impact on IVF outcome. The present
observation may be in accordance with their results, because no
differences were detected between the levels of vitamin D in
pregnant women and non-pregnant women considering the high
prevalence of vitamin deficiency in the sample. Also, the present
study found that the fertilization rate was highest in the first tertile
of 25OH-D level in follicular fluid; this difference was significant.
In accordance with the findings of Ozkan et al., the most recent
study by Rudick et al. [19] showed that vitamin D deficiency and
insufficiency was associated with lower pregnancy rates in
recipients of egg donation, suggesting that the effects of vitamin
D may be mediated through the endometrium. Furthermore, in
these two recent studies, the quality of embryos transferred in
relation to the level of vitamin D was not investigated. The present
study did not find any significant linear correlation between
metaphase II (and/or embryo quality) and 25OH-D levels in
follicular fluid and serum (r = 0.073, p = 0.517; r = 0.018, p = 0.784,
respectively). This may be explained by the vitamin D deficiency
observed in the study subjects.
Strengths of this study included the use of clinical pregnancy
(detection of gestational sac plus fetal heart rate) as an outcome
and following the patients to 20 weeks of gestation. Comparison of
the ongoing pregnancies with non-pregnancies did not reveal any
difference in vitamin D stores, but the fertilization rate decreased
significantly and the implantation rate increased (not significantly)
with increasing tertiles of 25OH-D level in follicular fluid. In
addition to considering factors affecting reproduction, this study
may indicate that the higher reproduction rates observed in
underdeveloped countries, such as Middle Eastern countries,
which have a high prevalence of vitamin D deficiency [17,20,21],
may not be affected by male or female vitamin D reservoirs.
Although vitamin D deficiency was prevalent among the pregnant
women in this study, it can be concluded cautiously that
conception is not correlated with vitamin D stores [22].
On the other hand, serum and follicular levels of 25OH-D did
not differ with the cause of infertility (e.g. PCOS). However, a
significant direct association has been reported between serum
levels of 25OH-D and PCOS [23], and it is likely that 25OH-D levels
increase to compensate for insulin resistance [24]. This issue was
not investigated in the present study, but deserves attention in
future research.
It appears that skin complexion, poor sun exposure, dietary
habits (e.g. vegetarian), air pollution, clothing habits (especially
among women), socio-economic factors such as lifestyle, older age,
institutionalized or hospitalized persons, and lack of vitamin D
food fortification programme can explain the high prevalence of
vitamin D deficiency in Iran and other countries in the Middle East
despite their sunny climate. In addition, this research was
conducted between August and March, so seasonal variation is
another explanation for the vitamin D deficiency of most of the
subjects, as nearly one-third of healthy adults are vitamin D
deficient at the end of winter [25].
While this study is of interest in evaluating the level of vitamin
D in follicular fluid, the observed prevalence of vitamin D
insufficiency in an otherwise healthy young female population
is of concern.
Prevalent vitamin D deficiency was the main limitation of this
research. This study was not able to demonstrate a lack of
correlation between vitamin D and the ART success rate, and did
not find that vitamin D deficiency cannot be a valid predictive
value in ART. However, the logistic regression model including the
variables related to clinical pregnancy did not show the 25OH-D
level in follicular fluid to be an independent predictor of clinical
pregnancy. This study did not prove that higher levels of 25OH-D in
follicular fluid have implications for ovarian steroidogenesis and
improved ovarian response to controlled ovarian hyperstimulation
or higher endometrial receptivity. As such, given the outcomes of
ART without considering the level of vitamin D, the assessment of
vitamin D status is not considered to be a part of routine infertility
work-up.
Due to limited human data about the role of vitamin D in
reproductive physiology, and small sample sizes in existing
research [16,18,19], including the present study, more powerful
studies with much larger sample sizes are needed to compare
pregnancy and livebirth rates with more realistic results.
Acknowledgements
This study was the Infertility Fellowship Thesis of Dr. Atossa
Mahdavi and was supported by Grant No.: 9462-30-03-88 from the
Deputy of Research, Tehran University of Medical Sciences. The
authors wish to thank the staff of the Infertility Department of
Shariati Hospital for all their help. In addition, the authors wish to
thank the Endocrine and Metabolism Research Centre of Shariati
Hospital for conducting the laboratory tests.
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