Vitamin D plays a crucial role in calcium and phosphate homeostasis, by increasing intestinal calcium absorption and renal calcium reabsorption. For vitamin D, accumulating evidence from observational human studies suggests a key role for both male and female fertility.
Vitamin D plays a crucial role in calcium and phosphate homeostasis, by increasing intestinal calcium absorption and renal calcium reabsorption 
. It is found in two major forms, D2
(ergocalciferol) and D3
(cholecalciferol). The former is produced by ergosterol upon irradiation in plants and fungi. The latter is produced by 7-dehydrocholesterol upon irradiation in the epidermis 
. After hydroxylation at carbon 25 [producing 25-hydroxyvitamin D, 25(OH)D], it is transported to the kidney, where it is hydroxylated by 1α-hydroxylase (CYP27B1) at the carbon 1 of the A ring, producing 1,25-dihydroxy-vitamin D [1,25(OH)2
D], the active form of vitamin D 
. CYP27B1 is also present in extrarenal sites, such as macrophages, osteoblasts, epithelial, endocrine, placental and cancer cells 
. The mechanism of 1,25(OH)2
D action involves its binding to vitamin D receptor (VDR), a transcription factor, member of the steroid hormone nuclear receptor family 
. VDR and CYP27B1 are expressed in various cells, indicating that vitamin D is characterized by a plethora of extra-skeletal actions, such as those on the immune and cardiovascular system 
Vitamin D deficiency is defined as 25(OH)D concentrations <20 ng/mL (50 nmol/L), whereas vitamin D insufficiency as 25(OH)D concentrations 20–30 ng/mL (50–75 nmol/L) 
. The prevalence of vitamin D deficiency ranges from 8 to 90% in Europe (reaching >50% in Western European populations) and from 14 to 89% in North America 
2. Vitamin D and Infertility
2.1. Vitamin D and Male Infertility (Observational Studies)
An accumulative body of evidence from observational studies suggests a potentially key role for vitamin D in male reproductive function, including semen quality and androgen status. This is indicated by a positive correlation between vitamin D concentrations and sperm motility 
and vitamin D concentrations and normal sperm morphology in infertile men 
. Particularly, a cross-sectional study including 300 men showed that men with severe vitamin D deficiency [25(OH)D <10 ng/mL] had a lower proportion of motile spermatozoa (62% vs. 70%; p
= 0.027), progressive motile spermatozoa (56% vs. 64%; p
= 0.035) and % of morphologically normal spermatozoa (6% vs. 8%; p
= 0.044) compared with those with vitamin D sufficiency 
. Similar results were obtained from a subsequent prospective study including 1427 infertile men, which demonstrated higher sperm motility in men with 25(OH)D >30 ng/mL compared with those with 25(OH)D <10 ng/mL [45% (31–63%) vs. 34% (22–54%), respectively; p
= 0.030]. However, no differences were observed regarding total sperm count, sperm concentration, sperm volume or sperm morphology 
. On the other hand, a cross-sectional study, including 170 men, showed a U-shaped correlation of vitamin D concentrations with semen parameters, supporting that not only low, but also high vitamin D concentrations are associated with impaired sperm quality 
. In detail, men with high vitamin D concentrations (≥50 ng had lower sperm concentration [46.7 (95% confidence interval (CI) 27.2 to 73.9) vs. 84.0 (95% CI 70.3 to 99.3) million/mL; p
< 0.05), progressive motile sperm [38.4% (95% CI 29.3 to 49.2) vs. 52.6% (95% CI 47.6 to 58.0); p
< 0.05] and normal sperm morphology [18% (95% CI 12.1 to 25.6) vs. 27.4% (95% CI 23.8 to 31.3); p
< 0.05) compared with those with 25(OH)D concentrations between 20 and 50 ng/mL 
Furthermore, serum 25(OH)D concentrations are associated not only with semen quality, but also with androgen status. Data from cross-sectional studies have shown a positive association between 25(OH)D and testosterone concentrations 
. In particular, a large cross-sectional study of 2299 men demonstrated higher total testosterone concentrations, free androgen index, and lower SHBG concentrations in vitamin D-sufficient compared with vitamin D-insufficient or -deficient men (p
< 0.05 for all) 
. Likewise, results from another cross-sectional survey of 3369 men in eight European centers (the European Male Ageing Study) supported a linkage between vitamin D deficiency and secondary or compensated hypogonadism [relative risk ratio (RRR) = 1.16, p
= 0.05], as 25(OH)D concentrations were positively associated with total and free testosterone and negatively with estradiol and LH concentrations 
2.2. Vitamin D and Male Infertility (Interventional Studies)
Few interventional studies have assessed the effect of vitamin D supplementation on semen quality, male fertility and testosterone concentrations. In a recent triple-blinded, randomized clinical trial, 330 men with infertility and vitamin D insufficiency received either a single dose of 300,000 IU cholecalciferol (followed by 1400 IU/day, combined with calcium 500 mg/day for 150 days) or placebo. Although there was no difference in sperm concentration, the number of spontaneous pregnancies was higher in the vitamin D compared with the placebo group (7.3% vs. 2.4%; 95% CI −0.6% to +10.5%) 
. In another study, 86 infertile men with idiopathic oligoasthenospermia were randomized to oral cholecalciferol 200 IU/day with calcium 600 mg/day, or a combination of vitamin E 100 mg plus vitamin C 100 mg, t.i.d. After three months, semen quality, especially the progressively motile sperm count per ejaculate and the proportion of progressively motile sperm were increased only in the vitamin D group. In particular, the mean count of progressively motile sperm per ejaculate was increased from 9.8 ± 3.7 × 106
to 21.5 ± 6.5 × 106
< 0.05) in the vitamin D group, while it was increased from 9.5 ± 6.3 × 106
to 12.4 ± 4.4 × 106
> 0.05) in the control group. The proportion of progressively motile sperm was also increased, from 18.4 ± 9.8% to 28.3 ± 4.5% (p
< 0.05) in the vitamin D group, while it did not increase in the control group (17.8 ± 5.3% to 21.4 ± 2.4%; p
> 0.05). In addition, pregnancy rates were higher in the vitamin D group (16.3%) compared with the control group (2.3%) (p
< 0.05) 
Additional interventional studies assessed the association between vitamin D and androgen status. A randomized controlled trial (RCT) in vitamin D-deficient men evaluated the effect of vitamin D supplementation (cholecalciferol 3330 IU/day, n
= 31) on testosterone concentrations, compared with placebo (n
= 23) 
. An increase in total (10.7 ± 3.9 to 13.4 ± 4.7 nmol/L; p
< 0.001), bioactive (from 5.2 ± 1.9 to 6.3 ± 2.0 nmol/L; p
< 0.001) and free testosterone concentrations (from 0.22 ± 0.08 nmol/L to 0.27 ± 0.09 nmol/L; p
< 0.001) was observed in the vitamin D supplemented group, while there was no change in the placebo group 
. Similar results were demonstrated by a prospective study, including 102 men who received a single dose of ergocalciferol (600,000 IU). Significant increase in serum total testosterone concentrations (from 12.46 ± 3.30 to 15.99 ± 1.84 nmol/L, p
< 0.01) and erectile function scores (from 13.88 ± 3.96 to 20.25 ± 3.24, p
< 0.01) were observed after 12 months 
The aforementioned data suggest a potentially adverse effect of low vitamin D status on male fertility, although a U-shape is more representative of its association with infertility. Vitamin D supplementation may improve sperm quality and increase spontaneous pregnancy rates and testosterone concentrations. Thus, while there is no level 1 evidence, vitamin D supplementation, achieving sufficient but not high (i.e., 30–50 ng/mL) 25(OH)D concentrations, may have a beneficial effect on male infertility 
2.3. Vitamin D and Female Infertility (Observational Studies)
Recently, research has focused on the role of vitamin D concentrations in women undergoing assisted reproductive technologies (ART). Based on data reported in a systematic review and meta-analysis of 11 cohort studies, including 2700 women, investigating the association between vitamin D status and ART outcome, higher live birth rates have been reported in vitamin D-sufficient compared with vitamin D-deficient and -insufficient women [odds ratio (OR): 1.33 (95% CI 1.08 to 1.65), seven studies] 
. Similar results were shown in another recently published meta-analysis of nine cohort studies, which supported the decreased live birth rates after in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) in women with vitamin D deficiency compared with those of sufficient vitamin D status [relative risk (RR): 0.74 (95% CI 0.58 to 0.90), three studies] 
Vitamin D has also been involved in the development of specific gynecological conditions affecting fertility, such as endometriosis and polycystic ovarian syndrome (PCOS). In particular, a cohort study of 49 women showed a significant linear correlation between 25(OH)D concentrations and the diameter of ovarian endometriomas (r = −0.3, p
= 0.03) 
. Moreover, a prospective comparative study, evaluating 25(OH)D concentrations in 135 women with endometriosis and 90 controls, showed that the incidence of women with vitamin D deficiency/insufficiency was significantly higher in women with endometriosis compared with the control group (80% vs. 33.3%; p
< 0.001) 
. Moreover, the impact of vitamin D concentrations on reproductive outcomes, in women with PCOS undergoing ovulation induction, has been investigated in a retrospective cohort study (n
= 540). This study showed that vitamin D-deficient women were less likely to achieve ovulation compared with those with 25(OH)D >20 ng/mL (OR 0.43, 95% CI 0.25 to 0.76, p
= 0.006). Furthermore, live birth rates after ovulation induction were increased by 2% for each 1 ng/mL increase in 25(OH)D concentrations (OR 1.02, 95% CI 1.00 to 1.04, p
= 0.040) 
. However, data are still insufficient in order to establish a possible causality between vitamin D and endometriosis or vitamin D and PCOS. Further studies are needed to confirm these associations.
2.4. Vitamin D and Female Infertility (Interventional Studies)
Few RCTs currently exist evaluating the effect of vitamin D supplementation on ART outcome, yielding inconclusive results. In a recent RCT 
, infertile women undergoing ICSI, using both fresh and frozen embryo transfers, were randomized to either cholecalciferol (50,000 IU/week) supplementation for six weeks (n
= 42) or placebo (n
= 43). Higher clinical pregnancy rates were shown in the vitamin D group compared with the placebo group (38.1% vs. 20.1%, p
= 0.019) 
. On the other hand, another RCT including 128 infertile women with vitamin D insufficiency, who underwent frozen-thawed embryo transfer cycles after IVF/ICSI and were treated either with cholecalciferol (50,000 IU/week) for 6-8 weeks (n
= 57) or with no intervention (n
= 57) 
, did not show any difference in clinical pregnancy rates between the two groups (25.5% vs. 21.8%, respectively; p
= 0.810) 
. Although safe conclusions cannot be drawn by these two studies [of note, both were conducted in Iran, with baseline 25(OH)D concentrations of 12.7–15.8 ng/mL], one should underline some differences between them, which may have had an impact on this discrepancy. First, participants of the former study 
were of normal body weight in contrast to those of the latter (mean BMI >26 kg/m2
. Second, the type of fertilization and the method of embryo transfer (i.e., fresh or frozen) also differed between studies. Third, 25(OH)D concentrations achieved after vitamin D supplementation also differed between studies (37 ng 
vs. 47 ng/mL 
With respect to PCOS patients, there is evidence for a possible beneficial effect of vitamin D supplementation on fertility outcomes. Particularly, a recent meta-analysis of nine RCTs (502 PCOS women) showed that vitamin D supplementation (in different doses) resulted in increased number of dominant follicles (>14 mm) compared with placebo or metformin (1000–1500 mg/day) (OR: 2.34, 95% CI 1.39 to 3.92, four RCTs) 
. Moreover, the same meta-analysis showed that vitamin D supplementation especially combined with metformin in women with PCOS seems to regulate menstrual cycles compared with women treated with metformin only (OR 1.85, 95% CI 1.01 to 3.39, three RCTs) 
. In addition, a recent double-blinded RCT was conducted aiming to evaluate the role of vitamin D supplementation on ICSI outcomes in 105 PCOS infertile women. Patients were randomized either to treatment group (vitamin E, 400 mg/day and vitamin D3, 50,000 IU every two weeks, n
= 52) or placebo group (n
= 53) for eight weeks. A higher clinical pregnancy rate was observed in the treatment group compared with the placebo group (62.1% vs. 22.6%; p
= 0.002), suggesting thus a beneficial effect of combined supplementation of vitamin D and E on ICSI outcomes in PCOS patients 
Thus, accumulating evidence supports the notion that vitamin D may play an important role in female fertility. Observational studies associate vitamin D status with IVF outcome, endometriosis and reproductive success after ovulation induction in women with PCOS (Table 1). However, RCTs have not yet established the effect of vitamin D supplementation on the success of ART. Hence, while there is no level 1 evidence, vitamin D sufficiency may be required for improving female fertility outcome.
Table 1. Vitamin D and infertility.
Linear or U-shaped correlation between vitamin D concentrations and sperm motility/morphology .
Sufficient vitamin D concentrations associated with high testosterone concentrations .
Supplementation of vitamin D improved semen quality and pregnancy rates .
Supplementation of vitamin D increased testosterone concentrations .
Contradictory data on whether supplementation of vitamin D is associated with pregnancy rates .
Higher live birth rates in vitamin D-sufficient women .
Linear correlation between vitamin D concentrations and diameter of ovarian endometriomas .
Higher incidence of vitamin D deficiency/insufficiency in women with endometriosis .
Linear correlation between vitamin D levels and reproductive success rates after ovulation induction in women with PCOS .
Abbreviations: ART: assisted reproductive techniques; PCOS: polycystic ovary syndrome.