Vitamin D Supplementation and Male Infertility: a Double Blinded Randomized Clinical Trial
Background In 2008, approximately 8% of all newborn children were conceived by assisted
reproduction. Intracytoplasmatic sperm injection (ICSI) accounts for ~50% of all treatments
and is mainly used in cases of severely reduced semen quality. This is in many cases most
likely caused by prenatal factors adversely affecting the developing testicles, and it is
therefore unlikely that any treatment of adult men will be able to completely normalize
their semen quality. However, less would also be clinically relevant. If reduced semen
quality could be improved prior to any assisted reproduction a less invasive treatment would
be needed. Eg. classical in vitro fertilization (IVF) rather than ICSI, simple intrauterine
insemination (IUI) rather than IVF or natural conception rather than IUI.
Several endocrine factors have been implicated in sperm production and maturation, but
little is known about the potential role of VD. VD is a key regulator of calcium homeostasis
and bone mineralization, although expression of the vitamin D receptor (VDR) in various
tissues has been related to several diverse actions. VD affects reproduction in several
animal species, convincingly shown in rodents, where VD deficiency in male rats resulted in
reduced sperm counts, and female rats inseminated with semen from VD deficient male rats had
lower fertility rates. The impaired reproductive performance is reversible and can be
corrected either by supplying VD or by normalizing calcium levels. Supported by VDR knockout
mice, which showed decreased sperm counts, reduced sperm motility and histological
abnormalities of the testis, which unlike the VD deficient male rats only partly can be
restored by calcium supplements.
The investigators have recently shown expression of VDR and the VD metabolizing enzymes in
the human testis, ejaculatory tract and in mature spermatozoa. The investigators
subsequently showed that VD in physiological concentrations increased intracellular calcium
in spermatozoa. VD acts through a rapid non-genomic response and the VD induced increase in
calcium may be crucial for the spermatozoa, because VD induced sperm motility and the
acrosome reaction. Expression analysis of spermatozoa from fertile and infertile men showed
that men with impaired semen quality have fewer that can metabolize VD than normal men(p <
0.0005) making presence of one of the proteins a potential marker of semen quality.
Furthermore, the investigators have just completed a cross sectional study of 300 young men
from the general population and found that low serum VD was associated with reduced sperm
motility and morphology.
SETTING, SCIENTIFIC PLAN AND RECRUITMENT Participants will be included among men referred to
the Department of Growth and Reproduction (dept. of GR), Rigshospitalet (RH) for evaluation
of male infertility
DESIGN This is a prospective, double blinded, two-arm randomized controlled trial
Group of intervention: Each man will receive 300,000 IU (7500 ug) cholecalciferol (D3)
orally once after blood- and semen sampling and performed DXA scan. Thereafter they will
receive VD tablets of 1,400 IU (35 ug) + 500 mg calcium daily for 3 months. Telephone
control after 4 weeks and at 3 months a clinical control and blood sampling will be
performed, followed by continued daily intake of 1400 IU VD + 500 mg calcium for 2
additional months. At end of treatment at five months after inclusion the men deliver two
semen samples, have a blood sample drawn and a DXA scan performed.
Group receiving placebo: Will follow the same chain of events, although VD will be replaced
PARTICIPANTS Screening of ~700-800 infertile men will be performed. The investigators assume
that ~340 men will be excluded and ~100 men do not wish to participate. Thus, 300 men will
be included in the study and half will be randomized to active VD treatment. The
investigators expect a small drop out (< 20) because of high motivation and no adverse
ANALYSIS AND INTERVENTION Reproductive hormones and growth factors will be analyzed at dept.
of GR, Rigshospitalet. An aliquot of the first blood sample will be analyzed before
inclusion to the study and another kept to be analyzed at the same time as the second blood
sample at the end of the trial to overcome the interassay variation. Other serum analyses
will be analyzed at department of clinical biochemistry, Rigshospitalet. Semen analyses and
DXA scans will be performed at dept. of GR.
SAMPLE SIZE CALCULATION AND STATISTICS In our association study with 300 participants the
investigators are able to detect significant dose response relationship between VD status
and sperm motility and morphology, and found significant differences between men with low
and high VD levels. Based on these results, the investigators assume to be able to increase
serum levels of VD with 50 nM with our setup. Including 150 men in each group will enable
us to detect changes in sperm motility and morphology of 15% and a 25% change in Inhibin B
levels. Intention to treat principles will be used and co-variate analyzes will be used to
analyze the potential effect of VD.
SCREENING AND TIME COURSE Men, who are investigated at dept. of GR due to infertility will
be screened for eligibility to the study, and those who meet the criteria for participation
will be informed, and if they consent allocated to active treatment with VD or placebo.
Allocation will be done by minimization using minim to avoid unbalanced grouping due to
randomization failure. Following variables will be balanced: Sperm concentration, BMI, serum
inhibin-B, and VD level.
ETHICS AND SIDE EFFECTS All the patients will have fulfilled their investigation, before
they are invited to the study. They will be informed of potential adverse effects, signs of
intoxication and they can leave the trial at any point without any consequences. The trial
will run in accordance with "good clinical practice". VD treatment gives virtually no side
effects and the risk of intoxication is almost nonexisting in the suggested setting, with
relatively low doses of VD and close monitoring of VD and calcium status. The participants
will be exposed to oral VC/placebo, to 2 DXA scans, have 3 extra blood samples drawn and
deliver 2 extra semen samples. All participants will be informed and counseling according to
their VD status.
PUBLICATION OF RESULTS All results, positive or negative will be submitted to peer reviewed
scientific journals. Data will successively be obtained and transferred to a statistical
PRACTICAL ISSUES Dept. of GR investigates annually more than 500 infertile men. Participants
will be included from that group. The department have the clinical expertise, experience and
capacity to perform all the investigations, except for the measurements of some of the blood
samples, which will be analyzed by department of clinical biochemistry. VD and placebo oral
suspensions are purchased from Glostrup apotek, while VD and placebo tablets are purchased
from Ferrosan, MD Martin. The investigators are collaborating with associate professor in
biostatistics J.H. Petersen, University of Copenhagen.
Allocation: Randomized, Endpoint Classification: Efficacy Study, Intervention Model: Parallel Assignment, Masking: Double Blind (Subject, Caregiver, Investigator, Outcomes Assessor), Primary Purpose: Treatment
difference in semen quality (semen variables total sperm count, sperm concentration, sperm motility, sperm morphology and semen volume) between VD and placebo treated men after 150 days of treatment.
Martin Blomberg Jensen, MD
Department of growth and reproduction, Rigshospitalet
Denmark: Danish Dataprotection Agency