Vitamin D and Vascular Function in Obese Children
Phase: NA
Conditions: Obesity, Vitamin D Deficiency
Sponsor: University of Pittsburgh

 
 
 
 
 
 
 
 
ClinicalTrials.gov Identifier: NCT01797302 
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Vitamin D and Vascular Function in Obese Children 
Clinical Trial Protocol 
 
ClinicalTrials.gov Identifier: NCT01797302 
 
Document Date: 12/2/2019 
 
Principal Investigator: 
Dr. Kumaravel Rajakumar, MD, MS 
Professor of Pediatrics 
University of Pittsburgh School of Medicine 
 
Supported by: 
(1) National Heart, Lung, and Blood Institute (NHLBI) of the National Institutes of Health 
R01HL112985 
(2) Office of Dietary Supplements (ODS) of the Office of Director of the National Institutes of 
Health R01HL112985 
 
 
 
 
 
 
 

 
 
 
 
 
 
 
 
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Study Title 
Vitamin D and Vascular Function in Obese Children 
Objectives 
The primary objective of this randomized clinical trial is to determine the efficacy of enhanced 
vitamin D3 supplementation in improving vascular endothelial function, arterial stiffness, insulin 
sensitivity, and metabolic syndrome risk status in 10- to 18-year-old vitamin D-deficient (serum 
25-hydroxyvitamin D <20 ng/mL) obese or overweight (BMI ≥85th percentile) children; and to 
assess whether these effects are dose-dependent. As a secondary objective, we will examine 
the vitamin D supplementation-induced effect on adipokines and inflammatory markers 
relevant to CVD risk. 
Design and Outcomes 
In a double-blinded, controlled trial, we propose to randomize 229 eligible children to receive 
either 600 IU (conventional supplementation, current recommended dietary allowance), or 
1000 IU or 2000 IU (enhanced supplementation) of vitamin D3 daily for six months. 
The primary outcome measure will be change in brachial flow-mediated dilation percentage at 
6 months (FMD%); FMD% is a measure of vascular endothelial function. The secondary 
outcome measures will include (1) pulse-wave velocity (PWV) – a measure of arterial stiffness, 
(2) central and systemic blood pressure, (3) fasting blood glucose concentration, (4) 1/fasting 
insulin concentration as a surrogate for insulin resistivity, (5) lipid profile indices, (6) 
adipocytokines (plasma leptin, and adiponectin) and systemic markers of inflammation (plasma 
high-sensitivity CRP, IL-6, TNF-α). 
Eligible participants will be scheduled for enrollment within eight weeks of the initial screening 
visit. Follow-up visits will occur approximately at three months and six months after enrollment. 
FMD%, PWV, biochemical measurements, and inflammatory markers will be obtained at each 
study visit; DXA scan will be done at enrollment and at 6 months. 
Interventions and Duration  
Enrolled children will receive daily oral supplementation of either 600 IU or 1000 IU or 2000 IU 
of vitamin D3 for six months. 
Sample Size and Population  
We aim to enroll 229 otherwise healthy 10- to 18-year-old, overweight (BMI percentile, ≥85th to 
<95th) or obese (BMI percentile, ≥95th), vitamin D-deficient (serum 25(OH)D concentration <20 
ng/mL) children. 
 
 

 
 
 
 
 
 
 
 
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1. STUDY OBJECTIVES 
1.1 Primary Objective 
To determine whether enhanced vitamin D3 supplementation in 10- to 18-year-old obese and 
overweight children with serum 25(OH)D <20 ng/mL results in: (a) improved vascular health 
(arterial endothelial function and arterial stiffness) and (b) improved insulin sensitivity and 
metabolic syndrome markers (blood pressure, fasting blood glucose, lipid profile). We 
hypothesize that vitamin D3 supplementation with 2000 IU or 1000 IU daily will be more 
effective than 600 IU in improving vascular health and insulin sensitivity and metabolic 
syndrome markers. 
1.2 Secondary Objectives 
To examine whether downregulation of systemic inflammatory markers and adipokines 
underlie the effects of vitamin D on vasculature and metabolism. We hypothesize that vitamin 
D effects on vascular health and metabolism may be driven by the attenuation of obesity-
associated inflammation. 
2. BACKGROUND AND RATIONALE  
2.1 Background on Condition, Disease, or Other Primary Study Focus 
Childhood obesity is a major public health problem in the United States (US). In 2007-2008, 32% 
of 2- to 19-year-old US children were either obese (BMI ≥95th percentile) or overweight (BMI 
85th - 94th percentile).1 Obese and overweight children exhibit cardiovascular disease (CVD) risk 
factors2 and have a high risk of metabolic syndrome3 (i.e. large waist circumference, 
hypertension, fasting hyperglycemia, hypertriglyceridemia, and low levels of high-density 
lipoprotein (HDL) cholesterol). Obesity and its atherogenic risks track from childhood to 
adulthood, increasing the risk of adult CVD and pointing to the need for developing primary 
prevention strategies in affected children. 
2.2 Study Rationale  
Vitamin D deficiency is emerging as an independent risk factor for CVD and metabolic syndrome 
in children4-7 and adults.8,9 Our preliminary work shows that obese children have an increased 
risk of vitamin D deficiency; hypovitaminosis D is also associated with other CVD risk factors, 
such as low HDL cholesterol and lower insulin sensitivity in youth.10,11 Accordingly, optimization 
of vitamin D status in obese children might improve their CVD risk profile and reduce their risk 
of later CVD. As vitamin D is a potent immune modulator, its cardiovascular and metabolic 
benefits might be mediated, in part, through downregulation of inflammatory mediators, 
including adipokines. 
In the proposed study, we will test the central hypothesis that enhancement of vitamin D status 
in otherwise healthy obese and overweight vitamin D-deficient children will improve their 
vascular health and their CVD and metabolic syndrome risk profile. 

 
 
 
 
 
 
 
 
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3. STUDY DESIGN 
We propose a double-masked, randomized, controlled clinical trial of enhanced vitamin D3 
supplementation (600 IU vs 1000 IU vs 2000 IU) for six months in 10- to 18-year-old, overweight 
(BMI percentile, ≥85th to <95th) or obese (BMI percentile, ≥95th), vitamin D-deficient (serum 
25(OH)D concentration <20 ng/mL) children. Children who seek care at the Primary Care 
Centers of CHP (Oakland, Mercy Hospital, and Turtle Creek) or at affiliated PittNet offices 
(Practice Based Research Network, PBRN) will be eligible to participate. Our primary aim is to 
determine whether enhanced supplementation with vitamin D3 (1000 IU or 2000 IU) compared 
to the standard of care vitamin D3 supplementation (600 IU), taken orally once daily, is effective 
in improving brachial FMD% by 25% or more at the end of six months. Secondary outcomes will 
determine effects of vitamin D supplementation in reducing arterial stiffness, improving insulin 
sensitivity, and reducing metabolic syndrome risks. We will also characterize the mechanistic 
effects of vitamin D on systemic inflammatory markers and adipokines relevant to CVD risk. 
4. SELECTION AND ENROLLMENT OF PARTICIPANTS 
Children who seek care at the Primary Care Center of CHP Oakland or at affiliated PittNet 
offices (Practice Based Research Network, PBRN) will be eligible to participate. 
Eligible children will meet the following inclusion and exclusion criteria 
4.1 Inclusion Criteria 
Potentially eligible children will be 
• 10 to 18 years of age 
• otherwise healthy 
• obese or overweight (BMI ≥85th percentile) 
• able to understand all aspects of study procedures 
• able to voluntarily comply with all study procedures across the duration of the study 
• vitamin D-deficient and normocalcemic during screening (serum 25(OH)D concentration <20 
ng/mL; serum calcium concentration: 10-14 years, 8.8-10.8 mg/dL; ≥15 years 8.4–10.2 
mg/dL) 
4.2 Exclusion Criteria 
Children will be excluded from the study if they meet any of the following exclusion criteria: 
• Have hepatic or renal disease, metabolic rickets, malabsorptive disorders, primary 
hyperparathyroidism, hyperthyroidism, previous history of cancer or other chronic 
disorders that could affect vitamin D or glucose metabolism 
• Are receiving treatment with anticonvulsants, systemic glucocorticoids, pharmacologic 
doses of vitamin D (>1000 IU of vitamin D2 or D3 daily), antihypertensives, vasoactive drugs, 
antilipidemics, metformin, antipsychotics, other oral insulin regulators, or cancer 
treatments 
• Have cholelithiasis or urolithiasis 

 
 
 
 
 
 
 
 
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• Have type 1 or type 2 diabetes 
• Have a condition or underlying abnormality that could compromise the safety of the subject 
during study procedures (e.g. sickle cell trait/disease) 
• Found to be hypercalcemic during screening visit (serum calcium >10.8 mg/dL in 10- to 14-
year-old children; >10.2 mg/dL in ≥15-year-old children) 
• Significant fasting hyperglycemia (fasting blood glucose concentration >125 mg/dL) during 
the enrollment visit 
• Post-menarchial girls with a positive pregnancy test at enrollment will be excluded; 
however, contraception is not required for post-menarchial girls 
4.3 Study Enrollment Procedures 
Participants will be recruited from the Primary Care Center of CHP Oakland or at affiliated 
PittNet offices. In addition to the two recruitment locations, participants may also be recruited 
through various community outreach programs. The study will also be advertised through the 
University of Pittsburgh Pitt+Me database to strengthen our recruitment effort. 
Potential participants among children attending the CHP Oakland Primary Care Center will be 
screened for eligibility by reviewing their EPIC electronic medical records. Subjects who are not 
excluded during the preliminary EPIC chart review will be approached for screening. Reasons 
for inclusion and exclusion will be electronically documented on a screening form in the study-
specific electronic database administered by the University of Pittsburgh Center for Research 
on Health Care (CRHC). 
Written informed consent from a biological or legally-adoptive parent and participants’ assent, 
will be obtained prior to enrollment (both screening and enrollment) of 10- to 17-year-old 
children. Participants who are 18 years of age can consent for themselves for both screening 
and enrollment. Legal guardians will have to provide documentation of guardianship from 
relevant agencies and/or courts. 
Participants found to be vitamin D-deficient during screening, and thus eligible for study 
enrollment, will be randomized to one of three vitamin D supplementation doses (600 IU, 1000 
IU, or 2000 IU). The randomization codes will be assigned electronically through the CRHC study 
database using a block randomization scheme set forth by the study statistician prior to the 
onset of the study. Both participants and study investigators will be blinded to the assigned 
supplementation dose throughout the trial. 
5. STUDY INTERVENTIONS 
5.1 Interventions, Administration, and Duration 
Enrolled study participants will be randomized to one of three vitamin D supplementation 
doses: 600 IU (conventional supplementation), 1000 IU, or 2000 IU (enhanced 
supplementation). Assigned study vitamin D supplement bottle will be dispensed to the 
participants at enrollment and 3-month follow-up visit. Supplementation bottles will be given 
with Medical Event Monitoring System caps (MEMS, manufactured by AARDEX Group Ltd., Sion, 
Switzerland) for assessing and enhancing adherence. Participants will be instructed to take only 
one study vitamin D tablet daily for the duration of the study, and to not double their dose if a 

 
 
 
 
 
 
 
 
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previous dose was missed. 
5.2 Handling of Study Interventions 
Study vitamin D supplements will be manufactured by Douglas Laboratories in Pittsburgh, PA. 
The procurement of the required tablets will be coordinated in conjunction with the UPMC 
Investigational Drug Service Research Pharmacist. Study vitamin D supplements will be directly 
shipped to the research pharmacist who will ensure appropriate masking, labeling, bagging, and 
coding of the supplement bottles. The study vitamin D supplements will be identical in color, 
taste, and size. Each dispensed bottle will contain 105 pills. 
The coded study vitamin D supplement bottles will be stored in a locked cabinet in the UPMC 
Osteoporosis Prevention and Treatment Center in the Kauffman Building. Once, the trial is 
completed, all excess vitamin D supplements will be returned to the UPMC Investigational Drug 
Service research pharmacist for discarding. 
If multiple children of the same household enter the study, the dispensed supplement bottles 
will be color-coded to ensure identification of their respective medication. 
5.3 Concomitant Interventions 
Allowed Interventions 
For the duration of the study, participants are permitted to take any medications that are 
not included in the list of prohibited interventions. 
Required Interventions 
Participants will be required to take one study vitamin D3 supplement tablet daily for the 
duration of the study (six months), beginning on the day of their enrollment. 
Prohibited Interventions 
Participants are expected to not take the following medications during their participation in 
the study. If these medications are clinically necessary and prescribed by a provider, the 
participant may be withdrawn from the study based on the PI’s discretion. 
• Multivitamins 
• Anticonvulsants 
• Systemic glucocorticoids 
• Antipsychotics 
• Antihypertensives 
• Vasoactive drugs 
• Antilipidemics 
• Metformin or other oral insulin regulators 
• Chemotherapeutics or other cancer treatments 

 
 
 
 
 
 
 
 
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5.4 Adherence Assessment 
Adherence to the vitamin D supplement will be measured using MEMS cap, pill count, and the 
medication calendar log. Each supplement bottle will be equipped with a MEMS cap, which will 
be activated when the study supplement bottle is dispensed during enrollment. Study staff will 
assess the adherence measures (MEMs cap, pill count, and medication calendar) during follow-
up. To promote valid MEMs cap data, subjects will be advised to take their study vitamin D 
supplement once daily directly from the dispensed bottle—preferably around the same time 
each day. Use of other medication storage devices or dispensers will be discouraged. 
6. STUDY PROCEDURES 
6.1 Schedule of Evaluations 
Assessment 
Screening 
Enrollment 
3-month 
Follow-Up 
6-month 
Follow-Up 
 
(Visit 0) 
(Visit 1) 
(Visit 2) 
(Visit 3) 
Inclusion/Exclusion Criteria 
x 
 
 
 
Informed Consent/Assent Form 
x 
x 
 
 
Demographics 
x 
 
 
 
Medical History 
x 
x 
x 
x 
Anthropometric Measurements 
x 
x 
x 
x 
Skin Color Assessment 
x 
 
 
x 
Short Vitamin D and calcium intake FFQ & 
Sunlight Exposure Questionnaire 
x 
 
 
x 
Blood Chemistries 
x 
x 
x 
x 
Urine Pregnancy Screen* 
 
x 
 
x 
Brachial FMD 
 
x 
x 
x 
Pulse Wave Velocity/Analysis 
 
x 
x 
x 
Dual-Energy X-ray Absorptiometry (DXA) Scan 
 
x 
 
x 
Activity Survey 
 
x 
 
x 
Youth and Adolescent FFQ 
 
x 
 
x 
Pubertal Self-Assessment (Tanner Stage) 
 
x 
 
 
Dispense Study Vitamin D Supplement 
 
x 
x 
 
Medication Adherence 
 
 
x 
x 
*Only post menarchial girls require a urine pregnancy screen as a safety requisite for DXA scan 
6.2 Description of Evaluations 
Potentially eligible patients passing the inclusion and exclusion criteria will be screened. 
Screened participants who are found to be vitamin D-deficient (serum 25(OH)D concentration 
<20 ng/mL) will be eligible to enroll in the study. Eligible participants will be scheduled for 
enrollment visit within eight weeks of their screening evaluation. Study duration is six months. 
Randomized participants will return for follow-up evaluations twice. The first follow-up visit will 

 
 
 
 
 
 
 
 
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occur approximately three months after their enrollment visit, and their final study visit will be 
scheduled approximately three months thereafter. 
Screening Evaluation 
Consenting Procedure 
Written informed consent will be obtained separately for screening and for enrollment, at 
the respective study visits. The parent or medical legal guardian of the child (as defined by 
state law) will be given a copy of the informed consent and will be provided an opportunity 
to read the consent in its entirety. For a legal guardian to consent on behalf of a child to 
participate in research, the guardian must present a copy of the appropriate legal document 
proving guardianship under state law before signing the consent document. A copy of the 
guardianship document will be included in the research subject’s chart. 
The consent form given at the screening visit will describe the procedures that will take 
place only during the screening visit. The consent form given at the enrollment visit will 
detail all procedures to be performed at each study visit. Participants must give their assent 
and only biological parents, or legally-recognized guardians, may provide consent. 
Only trained study coordinators and the principal investigator may obtain informed consent 
from participants. Person obtaining consent will review all aspects of the consent and allow 
the parent or legal guardian the opportunity to ask questions. The following points will be 
explained to the parent(s) or legal guardian(s) at the time of consent: 
• The study involves research 
• The purpose, background and rationale of the study 
• Study involves taking vitamin D supplementation (Vitamin D3 600 IU vs. 1000 IU vs. 2000 
IU once daily for six months) which has been masked and randomly assigned 
• The responsibility of the parent and the participant 
• All reasonably foreseeable risks or inconveniences associated with all the expected 
study procedures (i.e. vascular function tests, DXA scan, blood draw, skin color 
estimation, completion of physical activity, diet and sun exposure surveys) 
• Potential risk of hypervitaminosis D (vitamin D poisoning) and its symptoms 
• The expected benefits of the study 
• The participation in this study is voluntary and that the parent or subject may refuse to 
participate or withdraw from the study at any time without penalty of loss of benefits to 
which he/she may be entitled 
• The accessibility of the records to the regulatory authority(ies), monitor(s), auditor(s), or 
other appropriate reviewers 
• Maintenance of confidentiality of the patient record 
The consent form will include: the parent(s) or legal guardian(s) signature, the printed name 
of the child, and the date, as to document that all-of-the-above was completed. Participants 
who are 18 years of age will sign their own consent. The consenting research staff member 
will also sign the consent form to indicate that he/she has reviewed and explained the 
consent form with the subject and was present at the time of signing. After obtaining signed 

 
 
 
 
 
 
 
 
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consent, a copy of an unsigned consent form will be given to the parent(s) or legal 
guardian(s) for their records. The original signed copy of the consent will be kept within the 
study folder. 
 
Screening 
The following procedures are performed to determine a subject’s eligibility: 
• Age Calculation: To enter the study children should have completed their 10th birthday and 
not have had their 19th birthday. 18-year-old children are eligible to enroll until the day 
before their 19th birthday. 
• BMI Calculation: Height will be measured three times in centimeter (cm) to the nearest 
0.1-cm and averaged to determine the participant’s height in centimeter. Weight will be 
measured three times in kilogram (kg) to the nearest 0.1-kg and averaged to determine 
the participant’s weight in kilogram. BMI will be calculated using the following formula: 
[Weight in kilogram]/[Height in meter]2 
BMI can also be calculated using metric measurements of height (in cm) and weight (in 
kg) through NHLBI online BMI calculator available at: 
http://nhlbisupport.com/bmi/bminojs.htm 
• Medical History Form: Captures anthropometrics, demographic data (age, sex, race, and 
ethnicity), level of education, smoking status, current medications, and type of medical 
insurance. 
• Contact Form: To be completed by parent unless subject is 18 years of age 
• Vitamin D and Sunlight Exposure Questionnaire: To be completed by parents unless 
subject is 18 years of age 
• Waist Circumference Measurement: Waist circumference will be measured three times, 
using a non-stretch tape measure, at the mid-point between the lowest rib and the iliac 
crest at minimal respiration while standing. The respective waist circumference 
measurements will be measured to the nearest 0.1 cm and averaged. 
• Fitzpatrick Sun-reactive Skin Type estimation: Sun-reactive skin type estimate will be 
made by the parent unless the subject is 18 years of age 
• DSM II Colormeter Measurement: Study staff will normalize the 
dermatophotospectrometer on the calibration plate and obtain three readings from the 
forehead, back of the hand, and inner upper arm. The measurements obtained at each 
site will be averaged to estimate the melanin index at the respective sites. 
• Venipuncture, Specimen Collection, and Specimen Processing: 
o Venipuncture: Venipunctures at the Primary Care Center will be done by the clinic 
nurses or phlebotomy-trained medical assistants. For subjects consenting for 
screening at the Pediatric PittNet practices, venipuncture will be performed by the 
pediatric PittNet nurse, the clinic nurse, or the phlebotomy-trained medical 
assistants 

 
 
 
 
 
 
 
 
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o Specimen Collection: At least 5 mL of blood needs to be drawn in one or two red-top 
tubes and 10 mL of blood in one or two green-top tubes 
o Specimen Processing: Label the specimens with screening ID, date of birth, and date 
of visit. Complete lab test order forms (instructions for lab procedures). Enclose the 
collected tubes in a medical-biohazard seal bag with the lab order/procedure 
instruction sheets and drop-off the specimens at the appropriate place in the UPMC 
Central Lab building or in the clinic for transportation of the samples to the Clinical 
Chemistry Lab at UPMC Presbyterian. 
Enrollment 
Enrollment 
Participants that are deemed vitamin D deficient (serum 25(OH)D concentration <20 ng/mL) 
and meet all other inclusionary criteria are eligible to enroll in the study. Prior to entering 
the study (initiation of baseline assessments and randomization), participants will be 
required to complete a second informed consent, which outlines the assessments that will 
take place at the three study visits. 
Baseline Assessments at enrollment 
Patients who have been successfully screened for the study and have enrolled will undergo 
the following baseline assessments: 
• Medical history form 
• Anthropometric measurements 
• Venipuncture, specimen collecting, and specimen processing 
• Urine pregnancy screen: Only post-menarcheal girls require urine pregnancy screen. 
Only subjects with a negative pregnancy screen will be permitted to enroll. 
• Fasting glucose concentration screen: Only participants with fasting serum glucose <125 
mg/dL will be permitted to enroll 
• Activity survey: Participant may complete this survey, with parental help if they are ≤17 
years of age. 
• Nutritional survey: Parent may complete a survey regarding subject’s food frequency 
habits for participants ≤17 years of age. If subject is 18 years of age or older, he or she 
will complete this questionnaire on their own. 
• Pubertal self-assessment: Subjects will self-identify their pubertal stage by looking at 
Tanner Stage drawings and selecting the image that corresponds most closely to their 
stage of development. 
• Vascular function testing (pulse wave velocity, pulse wave analysis, and flow mediated 
dilation, and blood pressure measurements) 
o Blood pressure. Measurements will be obtained by a CTRC vascular technician prior 
to vascular health assessments, and after 10 minutes of participant rest and 
acclimatization while supine, using a model CONTEC08A automated digital 
oscillometric device (cuff 22-32 cm). Measurements will be taken three times, one 
minute apart, and averaged.  

 
 
 
 
 
 
 
 
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o Endothelial function. Brachial artery diameter will be measured using a high-
resolution ultrasound machine (GE, Vivid 7, GE Health care, Milwaukee, WI) 
equipped with a 9-L linear transducer preset to a dedicated vascular scanning 
protocol. We will then occlude brachial arterial flow at the upper forearm using a 5-
cm-wide occlusive cuff (Hokanson SC5) inflated to a pressure of 50 mmHg above the 
systolic blood pressure or to 200 mmHg, whichever is greater, by a rapid release 
sphygmomanometer (Hokanson DS 400) for 5 minutes. Post-cuff release diameter 
measurements during the reactive hyperemic phase, obtained at 60, 120, and 180 
seconds, will be used to calculate FMD%.  
o Arterial Stiffness Indices. Carotid-femoral pulse wave velocity (PWV), Aortic 
Augmentation Index at heart rate of 75/min (AIx-75), and central systolic and 
diastolic blood pressure were measured using arterial tonometry (SphygmoCor 
CVMS V9, CPVH System, Model EM3, AtCor Medical, Sydney, Australia).  
• DXA scan: Measures total body adiposity and percent of body fat  
Participants should enroll within two months of initial screening. If participants do not 
enroll within this time window, they will be required to rescreen to ensure eligibility. 
Subjects that have successfully completed the enrollment process and baseline procedures 
will be randomized to a study intervention dosage, which is blinded to both study team and 
participant.  
Parents will be instructed to record each dose of intervention given to their child in a 
calendar supplied by the investigators. 
3-month Follow-up Visit 
The window for the follow-up visit will be three months after the enrollment date (±15 
days). The following procedures will be performed at this visit: 
• Medical history form 
• Anthropometric measurements 
• Venipuncture, specimen collecting, and specimen processing 
• Vascular function testing (pulse wave velocity, pulse wave analysis, and brachial flow 
mediated dilation, and blood pressure measurements) 
• Medication adherence assessment 
o Collect medication calendar 
o Read MEMs cap and obtain Patient Adherence Report 
• Dispensation of second supply of vitamin D supplements 
Final Visit (6-month Follow-up Visit) 
The window for the final visit will be three months after the 3-month follow-up date (±15 
days). The following study procedures are performed at this visit: 
• Medical history form 
• Anthropometric measurements 

 
 
 
 
 
 
 
 
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• Venipuncture, specimen collecting, and specimen processing 
• Urine pregnancy screen (only post-menarchial girls require urine pregnancy screen). 
Subjects with a positive pregnancy screen will not undergo a DXA scan but can complete 
the remaining study procedures if they are willing.  
• Activity survey 
• Nutritional survey 
• Vitamin D and Sunlight Exposure Questionnaire 
• DSM II Colormeter Measurement: Will be measured at similar body sites as the baseline 
testing 
• Vascular function testing (pulse wave velocity, pulse wave analysis, and brachial flow 
mediated dilation, and blood pressure measurements) 
• DXA scan 
• Medication adherence assessment 
o Collect medication calendar 
o Read MEMs cap and obtain Patient Adherence Report 
7. SAFETY ASSESSMENTS 
7.1 Specification of Safety Parameter 
• Serum calcium levels will be monitored for participant safety. Serum calcium >10.8 
mg/dL in 10- to 14-year-old children and >10.2 mg/dL in ≥15-year-old children will 
prompt further evaluation. An elevated serum calcium level will be repeated and if still 
abnormal, study medication will be discontinued. Children who discontinued study 
medication due to hypercalcemia will have serum 25(OH)D measured to rule out 
vitamin D toxicity (levels ≥100 ng/mL) and will be referred to a pediatric endocrinologist 
for further evaluation and management. Children with hypercalcemia and normal serum 
25(OH)D levels (<100 ng/mL) will be also be referred for further evaluation to their 
primary care provider and a pediatric endocrinologist to rule out other unusual causes 
of hypercalcemia (chronic granulomatous disorders, primary hyperparathyroidism, 
familial hypocalciuric hypercalcemia). 
• Significant fasting hyperglycemia (fasting blood glucose >125 mg/dL) during enrollment 
visit will disqualify subject from enrolling in the study. 
• Post-menarchial girls that test positive during pregnancy screen at the enrollment visit 
will be excluded. Positive screens in the final visit will disqualify subject from getting a 
DXA scan but will not exclude subject from completing the study. 
• At the 3-month follow-up and the final visit, the MEMS cap data will be downloaded to 
assess adherence. 
7.2 Methods and Timing for Assessing, Recording, and Analyzing Safety Parameters 
Serum calcium will be measured at the screening visit and only children with normal calcium 
results for age will be eligible to enter the study. Upon enrollment, serum calcium will be 
monitored for safety at the 3-month and 6-month follow-up visits, and children found to have 
elevated serum calcium (>10.8 mg/dL in 10- to 14-year-old children and >10.2 mg/dL in ≥15-
year-old children) will be further managed as stated above in section 7.1. 

 
 
 
 
 
 
 
 
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7.3 Adverse Events and Serious Adverse Events 
An adverse event is any unintended medical occurrence (symptom, diagnosis, sign, abnormal 
laboratory finding, or missed study procedure) that occurs during the study that was not 
present at baseline. 
• Any medical condition present at the time of enrollment is considered baseline, and not 
reported as an adverse event. 
• If the severity of a pre-existing medical condition worsens at any time during the study 
participation, it will be considered an adverse event. 
• All adverse events will be assessed by the Principal Investigator to determine the 
severity of the event (mild, moderate, severe, life-threatening) and whether the event is 
related to the study product or study intervention 
• All AEs will be recorded in the eCRF in the study database. The event description, date of 
onset and study clinician’s assessment of severity and relationship to study product, 
date of resolution and outcome will be recorded. To elicit any adverse event, the parent 
will be prompted at each encounter to report any change in their child since the last 
contact. 
A serious adverse event (SAE) is any unintended medical occurrence in which the patient 
outcome is death or life-threatening hospitalization, disability, or incapacity. 
• In the event of an SAE, the SAE eCRF must be completed by the research coordinator 
and the details of the relevant SAE should be notified to the PI (Dr. Rajakumar) 
immediately. The event description, date of onset and study clinician’s assessment of 
severity and relationship to study product, and outcome of event will be recorded. 
• SAEs will be logged in the electronic database and will be periodically monitored by the 
study statistician. 
7.4 Reporting Procedures 
• All AEs will be reported to the Data and Safety Monitoring Board annually. 
• Unanticipated protocol-related problems will be notified by the PI (Dr. Rajakumar) to 
the IRB as and when they occur 
• All SAEs must be reported by the research staff to Dr. Rajakumar, the study principal 
investigator, immediately. Principal investigator will report SAEs to the University of 
Pittsburgh Institutional Review Board (IRB) according to the local IRB requirements. 
SAEs related to the research intervention will be notified by the PI (Dr. Rajakumar) to 
the IRB within 24 hrs. AEs of moderate severity related to the research intervention will 
be notified by the PI (Dr. Rajakumar) to the IRB within 5 days.  
7.5 Follow-up for Adverse Events  
• Follow-up of a non-serious adverse event of mild or moderate severity will end when 
the study reporting period ends [i.e., 6-month follow-up visit (final study visit)]. Study 
staff will keep apprised of subject’s adverse event until the parent/subject provides a 
date of resolution for the AE; resolution date will be documented in the CRHC database. 

 
 
 
 
 
 
 
 
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• Follow-up for a non-serious adverse event of severe nature will be the following: 
o At the time of the event, the investigator will provide appropriate treatment or 
follow up, including communicating the event to the child’s primary care provider 
and/or providing a referral to a specialist. 
o The staff will place a follow up phone call to the parent, regardless of the study 
reporting period, to assure that the child is improving or that the family has 
connected with the suggested referral. Study staff will keep apprised of subject’s 
adverse event until the parent/subject provides a date of resolution for the AE; 
resolution date will be documented in the CRHC database. 
7.6 Safety Monitoring 
• Members of the Data and Safety Monitoring Board will evaluate participants’ risks 
(adverse events) vs. the benefits and other factors that can affect study outcome. 
• Real-time reports of SAEs will be sent to the Data and Safety Monitoring Board as 
determined by the principal investigator. 
• We will appoint an Independent Safety Monitor (ISM) who will have expertise in 
pediatrics and whose primary responsibility will be to provide timely safety monitoring, 
to ensure good clinical practice and to quickly identify safety concerns. Safety reports by 
the ISM will be submitted on a regular basis, (e.g., real time, weekly, monthly, quarterly) 
as determined by the DSMB. To minimize bias, the ISM will evaluate SAEs masked to 
treatment assignment, unless the DSMB approves partial or complete unmasking. The 
ISM will participate for the duration of the study. 
• The ISM will be able to readily access participant records. The primary focus of the ISM 
is to independently review SAEs that may be associated with study treatment as well as 
adverse events of special interest. The ISM will investigate any events considered 
serious and unexpected. The ISM will be in communication with Dr. Rajakumar and the 
DSMB chair for any event that needs further evaluation. The ISM will not be directly 
involved in the trial, will not be under the investigator’s supervision, and will have no 
financial, intellectual, proprietary or professional interest in the outcome of the trial. 
8. INTERVENTION DISCONTINUATION 
To ensure children’s safety, we will use a serum calcium cutoff of >10.8 mg/dL in 10- to 14-year-
old children and >10.2 mg/dL in ≥15-year-old children to prompt further evaluation. An 
elevated serum calcium level will be repeated and if it remains abnormal, study medication will 
be discontinued, and participant will be withdrawn from the study protocol. Children who 
discontinued study medication due to hypercalcemia will have serum 25(OH)D measured to 
rule out vitamin D toxicity (levels ≥100 ng/mL) and will be evaluated by a pediatric 
endocrinologist. Children with hypercalcemia and normal serum 25(OH)D levels (<100 ng/mL) 
will be referred for further evaluation to their primary care provider and a pediatric 
endocrinologist to rule out other unusual causes of hypercalcemia (chronic granulomatous 
disorders, primary hyperparathyroidism, familial hypocalciuric hypercalcemia). 

 
 
 
 
 
 
 
 
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9. STATISTICAL CONSIDERATIONS 
9.1 General Design Issues 
In this randomized, controlled, double-blinded clinical trial, eligible children will receive in a 
1:1:1 ratio either 600 IU or 1000 IU or 2000 IU vitamin D3 supplementation orally once daily for 
six months. Our primary outcome measure will be brachial FMD at 6-month follow-up. A 25% or 
more improvement in brachial FMD% in the 1000 IU or the 2000 IU group when compared to 
the 600 IU group, after six months of vitamin D replacement will be considered significant. 
Our secondary outcomes will include: (1) pulse wave velocity (PWV) as a measure of arterial 
stiffness, (2) 1/fasting insulin concentration as a surrogate for insulin sensitivity, and (3) 
metabolic syndrome risk factors (blood pressure, waist circumference, HDL cholesterol, 
triglycerides, and fasting blood glucose) at the 6-month follow-up. 
9.2 Sample Size and Randomization 
Our sample size estimates are based on showing a relative improvement in baseline brachial 
artery flow mediated vasodilation (FMD, %) of at least 25% in children receiving each of the 
higher doses of vitamin D3 (1000 IU and 2000 IU) compared with no improvement in the 600 IU 
group after six months of treatment. We assumed the baseline FMD was ~6.5% based on 
previous studies in overweight children similar to our projected population.12-14 A sample size of 
56 in each treatment group at the end of the trial would have 84% power (two-sided two-
sample comparison of means, α=0.025) to detect this clinically important difference of 25% 
relative improvement (absolute improvement=25%*6.5%≈1.6%) assuming a standard deviation 
of 2.6% (conservative estimates based on published literature).12,14 To provide for a projected 
drop-out rate of 25% by the end of the trial, we propose to recruit 225 children into the trial. 
We have over 90% power to detect a dose-response effect by testing for linear trend assuming 
a 0%, 20%, and 30% improvement in vascular health among children receiving 600 IU, 1000 IU, 
and 2000 IU, respectively. 
Our original sample size of n=252 was based on 90% power and 20% attrition to allow for 
subgroup analyses by race. In December of 2015, the PI and funder decided to re-evaluate the 
targeted sample size due to slower than expected recruitment. Accordingly, with approval from 
NHLBI, our accrual target was revised from 252 participants to 229 participants based on 
projected enrollment and power of at least 80% to detect the original effect size with 25%-30% 
attrition. The lower sample size of n=225 would still allow for detection of clinically significant 
effect of vitamin D supplementation. However, the proportion of African Americans in the 
recruited sample was much larger than anticipated; the small number of Caucasian subjects in 
the cohort precludes doing race-stratified analysis. For the secondary outcomes, we have 89% 
power to detect effect sizes of 0.61 or greater without adjustment for multiplicity. 
 
Treatment Assignment Procedures 
In this randomized, controlled, double-blinded clinical trial, eligible children will be randomly 
allocated, using a permuted block design, in 1:1:1 ratio, to receive either conventional (600 IU) 

 
 
 
 
 
 
 
 
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or enhanced (1000 IU or 2000 IU) vitamin D3 supplementation orally once daily for six months. 
Study supplement bottles were masked and labeled by the research pharmacist with 
appropriate randomization codes. At enrollment, the CRHC database assigned the 
randomization code based on the permuted block scheme. 
Study participants or investigators will not have access to the assigned doses until the end of 
the clinical trial [timing of planned breaking of randomization code]. Unmasking may occur in 
the instance of an SAE. The ISM may divulge the masked dosage to the PI or relevant clinicians 
who are treating the patient. 
9.3 Interim analyses and Data and Safety Monitoring Plan 
No interim efficacy analysis has been planned. Dr. Moore will work with Dr. Rajakumar to 
prepare a DSMB report addressing specific concerns that she anticipates the DSMB may have 
regarding the conduct of the study. This will be distributed to DSMB members along with the 
Open Session report. Likewise, Dr. Moore’s report for the Closed Session will usually contain an 
assessment of the progress of the trial, including recommendations as to whether it should be 
terminated or modified. Interim data reports will generally include the following types of 
information, although only the Closed Session data reports will include comparisons by 
treatment group: 
• Monthly and cumulative accrual compared with targets 
• Baseline characteristics, overall and by treatment group 
• Completeness and quality of data collection forms 
• Status of enrolled participants, overall and by treatment group 
• Participants’ off-protocol treatments 
• Compliance with eligibility criteria and other protocol requirements 
• Subject adherence to the treatment regimen, overall and by treatment group 
• Individual SAEs by subject ID number and a table of event-specific cumulative rates, 
overall and by treatment group 
Dr. Moore will work to ensure that all the information presented at the DSMB meetings 
remains confidential. Each report will be marked as such. At the end of each DSMB meeting, all 
reports will be collected and destroyed. At the end of the meeting, the DSMB will make 
recommendations regarding the continuation of the trial. Dr. Rajakumar will implement the 
DSMB recommendations. Dr. Rajakumar will also submit the DSMB recommendations to the 
IRB. 
9.4 Outcomes 
Primary outcome 
Our primary outcome measure will be change in mean FMD% at 6-month follow-up. A 25% 
or more improvement in FMD% in the 1000 IU or 2000 IU compared to the 600 IU group 
after six months of vitamin D replacement will be considered significant. 
Secondary outcome 

 
 
 
 
 
 
 
 
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Our secondary outcomes will include: (1) pulse wave velocity (PWV) as a measure of arterial 
stiffness, (2) 1/fasting insulin concentration as a surrogate for insulin sensitivity, and (3) 
metabolic syndrome risk factors (blood pressure, waist circumference, HDL cholesterol, 
triglycerides, and fasting blood glucose) at the 6-month follow-up. 
9.5 Data Analyses 
Our primary analysis will use children’s original treatment assignment regardless of adherence 
(intention to treat). We will begin by calculating descriptive statistics for baseline demographics 
and clinical characteristics by vitamin D treatment groups (600 IU vs 1000 IU vs 2000 IU). Our 
primary outcome measure of vascular health is brachial artery FMD expressed as a percentage. 
We will use mixed-effects models with fixed effects of time (baseline, 3 months, and 6 months), 
the treatment*time interaction, and random subject effects to test for differential changes in 
FMD over time and differences in FMD at six months. We are primarily interested in the 
comparisons of each higher dose (1000 IU and 2000 IU) to the lower dose (600 IU) at six months 
(α=0.025 for each comparisons). The mixed-effects model will allow unbiased estimation of the 
treatment effects at six months even if some 6-month data are missing because of the 
incorporated dependence among baseline, 3-month and 6-month assessments (assuming data 
are missing at random). We will test the main effect of vitamin D supplementation (three 
groups) at 3 and 6 months and pairwise comparisons of the groups receiving the larger doses of 
vitamin D with the group receiving 600 IU.  If differences are detected across the 3 groups, we 
will test for a dose response in FMD across doses by using a linear contrast of coefficients test 
(α=0.05). We will use the same methodology to test the effects of higher doses of vitamin D on 
pulse-wave velocity, our primary measure of arterial stiffness. 
The second part of our primary analysis of our trial is to determine whether higher doses of 
vitamin D will result in improvements in insulin sensitivity and metabolic syndrome markers 
over time. We will use mixed-effects models with fixed effects of time (baseline, 3 months, and 
6 months), and the treatment time interaction with random subject effects to test for 
differential changes in secondary outcomes over time. We will test the main effect of treatment 
(three groups) at 3 and 6 months and pairwise comparisons of the groups receiving the larger 
doses of vitamin D with the group receiving 600 IU.  All tests will be two sided with α=0.05. 
We will explore the mediating effects of inflammatory markers (hs-CRP, IL-6, TNF-α) and 
adipokines (leptin and adiponectin) on the temporal relationship between vitamin D and 
vascular health. We hypothesize that vitamin D effects on vascular health may be driven by 
inflammatory processes and adipokines. An analysis that would complement this hypothesis 
would be a mediation analysis where we demonstrate that the effect of vitamin D on vascular 
health is partially mediated by our measures of inflammation and adipokines. We propose to 
use single-mediator and multiple-mediator methods incorporating product of coefficients to 
test for mediation.15 For the simple single-mediator analysis, we first must demonstrate that 
the high dose vitamin D has a significant effect on each mediator (inflammatory markers and 
adipokines) at three and six months using a general linear model. We will then test whether 6-
month vascular health is correlated with each mediator. Both tests would have to be significant 
for there to be evidence of mediation. We will estimate the mediated effect by taking the 
product of the following coefficients: 1) the coefficient for the intervention effect on the 
mediator, and 2) the coefficient for the mediator in a model testing the intervention effect on 

 
 
 
 
 
 
 
 
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vascular health controlling for the potential mediator. This estimate will provide the reduction 
in the effect of vitamin D on vascular health when adjusted for the mediator. The confidence 
interval for the mediated effect will be calculated using bootstrap methods. Since our vitamin D 
intervention is hypothesized to target multiple mechanisms, we will test for a multiple-
mediator model after single mediation is thoroughly investigated. 
Missing Data 
We do not expect substantial missing item-response data due to the electronic nature of data 
collection. Based on our pilot work, we anticipate <20% attrition at the 6-month assessment, 
and we have accounted for this in sample-size analyses. Participants who wish to discontinue 
the study drug will be asked to continue to participate in all follow-up visits; those who no 
longer wish to participate in the study will be considered dropouts, and we will monitor these 
events. We will obtain reasons for dropout in case it appears possible to modify the protocol so 
as to increase retention. At the end of the study, we will compare baseline characteristics and 
treatment arms between participants who have received 3- and 6-month assessments and 
participants who have not, to learn what factors are associated with missing visits. 
For participants with missing 6-month outcome data, we will conduct several sensitivity 
analyses to test the impact on our study conclusions. The methods proposed assume that the 
data are missing at random (MAR), where the missing data mechanism is related to observed 
data, not the unobserved values. Although this assumption is difficult to confirm, our sensitivity 
analyses will allow us to assess the robustness of our findings and threats to the MAR 
assumption. Given that our outcomes are highly mechanistic and less patient-relevant 
symptomatic/functional outcomes, missing data at 6 months would likely be related to baseline 
characteristics or 3-month values. As mentioned previously, we will use linear mixed models for 
analysis, since these models are more robust to missing data than traditional multivariate 
models or single-point-in-time ANOVA. From these models, we can estimate the treatment 
impact at 6 months using the covariance structure of the repeated measurements. This 
approach will be compared to a complete case analysis (missing complete at random) and a 
nonignorable missing data method (pattern mixture modeling). These different methods should 
result in consistent qualitative results if our findings are robust to missing data. 
 
10. DATA COLLECTION AND QUALITY ASSURANCE 
10.1 Data Collection Forms 
Data based on source documents (eligibility, anthropometrics, demographics, etc.) will be 
assessed and entered by the research coordinator. Data collected in paper records (skin-typing, 
sun exposure history, DSM II color-meter readings, pubertal self-assessment, lab results, 
adverse events, etc.) will be entered into the CRHC study database, and data with electronic 
outputs (vascular function results and adherence data from MEMs cap) will be merged into the 
CRHC study database. Tracking reports will be developed to assess recruitment, participant 
status, upcoming visits, and outstanding assessments. Automated reports for data and safety 
monitoring will also be developed. 

 
 
 
 
 
 
 
 
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All paper records containing identifying information will be kept in locked files accessible only 
to study staff and unlocked only while a study staff member is working with the files. 
Information regarding individual subjects will be kept private and shared only with the IRB and 
appropriate government agencies. 
10.2 Data Management 
Data management effort for this clinical trial will be coordinated through University of 
Pittsburgh’s Center for Research on Health Care Data Center and will use a paperless data 
collection systems (eSYSDM), using both the Internet and tablet PCs with backup paper data 
collection. Dr. Rajakumar will work with Dr. Moore to oversee all aspects of data management. 
Along with the lead research coordinator, they will develop an operation manual to standardize 
all procedures and staff training in areas such as participant recruitment; measurements; 
assessments; and data entry, management, and security. 
10.3 Quality Assurance 
Training 
All key personnel and research staff have completed or will complete the required training 
and certification for research integrity, protection of human subjects, HIPPA privacy and 
blood-borne pathogens. 
Quality Control Committee 
Quality Control Committee will include Dr. Rajakumar, Dr. Moore, and Ms. Kearney.  
 Metrics 
• Monthly and cumulative accrual compared to enrollment/follow-up targets 
• Baseline characteristics, overall and by treatment group 
• Completeness and quality of data collection forms 
• Status of enrolled participants, overall and by treatment group 
• Participants’ off-protocol treatments 
• Compliance with eligibility criteria and other protocol requirements 
• Subject adherence to the treatment regimen, overall and by treatment group 
• Individual SAEs by subject ID number and a table of event-specific cumulative rates, 
overall and by treatment group 
Protocol Deviations 
Any protocol deviations recognized by study staff will be communicated to Dr. Rajakumar 
(PI). The PI will determine if the IRB needs to be immediately apprised of the deviation or 
can be informed at the next renewal. This deviation will be documented in the “As Needed” 
forms in the CRHC database. The protocol deviations and the course of actions taken to 
address the deviations will be discussed in the next DSMB meeting. 

 
 
 
 
 
 
 
 
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Monitoring 
Dr. Rajakumar, Dr. Moore, and Ms. Kearney have extensive experience in all areas of clinical 
trial oversight, including developing quality assurance benchmarks, enforcing quality 
control, and monitoring protocol-specific activities through ongoing reporting structures 
and on-site evaluations. Dr. Moore will be responsible for generating reports on 
recruitment and follow up; quality control reports, which include the quality of data 
received; participant follow up adherence data, which include missed visits; and participant 
characteristics. Ms. Kearney will be responsible for monitoring the study site to ensure that 
all regulatory procedures are met. This will include ensuring that all study personnel 
(especially those hired after the trial begins) have completed required training and received 
certification to conduct human subjects’ research. 
Within one month of initiating a protocol, we will conduct an audit to review the study 
procedures and documentation of their use, including standardized procedures for 
obtaining informed consent, proper recording of data, reporting AEs and SAEs, training of 
personnel, and protection of patient identification and confidentiality. This process will be 
repeated every six months to assure that the protocol is being properly implemented, and 
that data are being collected and entered accurately. Enrollment and retention rates, 
protocol deviations, all SAEs, and all informed consent documentation will also be reviewed. 
The following elements will be reviewed under the Data and Safety Monitoring Plan: study 
progress, including an assessment of data quality (monitored monthly); outcomes and 
adverse-events data, including out-of-range laboratory results (ongoing monitoring); any 
pertinent new information (monitored every six months); study procedures designed to 
protect the privacy of the subjects and the confidentiality of the data, i.e., that data and 
charts are stored in locked area and data are recorded to protect the identity of subjects 
(monitored weekly); interim analysis and final conclusions evaluating benefit-to-risk ratio of 
study participation. Data and Safety Monitoring Plan reports will be submitted yearly to the 
Institutional Review Board at the time of renewals. 
11. PARTICIPANT RIGHTS AND CONFIDENTIALITY  
11.1 Institutional Review Board (IRB) Review 
This protocol and the informed consent documents, and any subsequent modifications to these 
documents, will be reviewed and approved by the IRB committee responsible for oversight of 
the study. 
11.2 Informed Consent Forms 
Written informed consent from either a biological or adoptive parent, and participant’s assent, 
are required for enrollment of eligible participants <18 years of age (10- to 17-year-old 
children). Eligible participants who are 18 years of age can provide legal consent for 
themselves. Consent forms will include the purpose of the study, why the subject is eligible to 
participate, the study procedures to be implemented, associated benefits and risks, the 
voluntary nature of the study, and notices of privacy rights. The original, signed consent form 
will be retained by the study and a copy of the consent form will be given to the 

 
 
 
 
 
 
 
 
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parent/guardian. If a non-biological legal guardian consents for the subject, a copy of state 
documentation stating legal guardianship will be retained with the consent form. Consent 
documents will only be in English; non-English speakers/readers will not be eligible to consent 
for the study. 
11.3 Participant Confidentiality 
Subjects will be assigned an anonymizing Participant ID, which will serve as the primary means 
of identifying subject to non-study related personnel (e.g. clinical labs, etc.). All paper records 
containing identifying information will be kept in locked files accessible only to study staff and 
unlocked only while a study staff member is working with the files. All computer and 
networking systems will comply with the University of Pittsburgh standard for digital security 
and password-encryption, as necessary. Information regarding individual subjects will be kept 
private and shared only with the IRB and appropriate government agencies. 
11.4 Study Discontinuation 
The study may be discontinued at any time by the IRB, the NHLBI, the OHRP, the FDA, or other 
government agencies as part of their duties to ensure that research participants are protected. 
12. COMMITTEES 
• Data and Safety Monitoring Board 
• Quality Control Committee will include Dr. Rajakumar, Dr. Moore, and Ms. Kearney.  
13. PUBLICATION OF RESEARCH FINDINGS 
Proposed trial will be registered at ClinicalTrials.gov and the results of the will be submitted to 
ClinicalTrials.gov as per NIH policy. Finding of the trial will also be published in a peer-reviewed 
medical journal. 
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excess adiposity among overweight children and adolescents: the Bogalusa Heart Study. J 
Pediatr 2007;150:12-7 e2. 
3. Ice CL, Murphy E, Minor VE, Neal WA. Metabolic syndrome in fifth grade children with 
acanthosis nigricans: results from the CARDIAC project. World J Pediatr 2009;5:23-30. 
4. Delvin EE, Lambert M, Levy E, et al. Vitamin D status is modestly associated with glycemia 
and indicators of lipid metabolism in French-Canadian children and adolescents. J Nutr 
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5. Kumar J, Muntner P, Kaskel FJ, Hailpern SM, Melamed ML. Prevalence and Associations of 
25-Hydroxyvitamin D Deficiency in US Children: NHANES 2001-2004. Pediatrics 2009. 
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