FOLIC ACID IMPROVES ENDOTHELIAL FUNCTION IN CHILDREN AND

ADOLESCENTS WITH TYPE 1 DIABETES

ALEXIA SOPHIE PEÑA, MD, ESKO WILTSHIRE, MD, FRACP, ROGER GENT, DMU,
CRAIG HIRTE, BSC(HONS), AND JENNIFER COUPER, MD, FRACP

Objective To evaluate the effect of folate supplementation on endothelial function in children and adolescents with type 1
diabetes.
Study design Thirty-six subjects with type 1 diabetes age 13.6 ± 2.6 years completed a randomized, double-blind, placebo-
controlled crossover trial. Each subject received 8 weeks of oral folic acid (5 mg/d) and 8 weeks of placebo, with an 8-week
washout period. Before and after each intervention, we assessed endothelial function by using brachial artery responses to flow
(flow-mediated dilatation [FMD]) and glyceryl trinitrate, von Willebrand factor, glucose, hemoglobin A1c, total plasma
homocyst(e)ine (tHcy), vitamin B12, serum folate, and red cell folate (RCF).
Results Folic acid increased FMD by 2.58 (3.1-5.7) % (95% confidence interval, 1.28-3.88), whereas placebo did not change
FMD (–0.42%; 95% confidence interval, –1.67 to 0.83; P < .001). Folic acid increased serum folate by 14 nmol/L (6.2 ng/mL,
P < .001) and RCF by 467.2 nmol/L (206 ng/mL, P < .001). Change in FMD was related to change in serum folate (r = 0.46,
P = .005) and RCF (r = 0.39, P = .02). Glyceryl trinitrate responses, von Willebrand factor, tHcy, and hemoglobin A1c were not
affected by the intervention.
Conclusions Short-term high-dose folic acid improves endothelial function in children and adolescents with type 1 diabetes
and normal folate status independently of tHcy. (J Pediatr 2004;144:500-4)

he endothelium is a key regulator of vascular function.1 Endothelial dysfunction is

T an early and fundamental event in the development of atherosclerosis.2 Endothelial

dysfunction occurs early in type 1 diabetes3-5 and is critical to the pathogenesis of
microvascular and macrovascular complications of diabetes.6 Markers of endothelial
From the Departments of Endocrinol-
dysfunction precede the onset of microalbuminuria.3,7 Prevention of the vascular ogy and Diabetes, Medical Imaging,
complications of diabetes therefore requires strategies that begin early in the disease, and Public Health Research Unit, and the
endothelial function is a useful surrogate marker to assess the effectiveness of such Department of Pediatrics, University
of Adelaide, Women’s and Children’s
strategies. Hospital, North Adelaide, South Aus-
Hyperglycemia, insulin, and increased total plasma homocyst(e)ine (tHcy) contribute tralia, Australia; and the Department
of Pediatrics and Child Health, Wel-
to endothelial dysfunction in diabetes.8-11 Folic acid improves endothelial function in adults lington School of Medicine and Health
with upper quartile tHcy.12-14 A single dose of folic acid improves endothelial function in Sciences, Wellington South, Welling-
adults with type 2 diabetes, as measured by forearm plethysmography.15 There is a scant ton, New Zealand.
Supported by grants from the Chan-
pediatric literature, but a trial in children with chronic renal failure showed improvement in nel 7 Research Foundation and the
endothelial function with folic acid.16 Women’s and Children’s Research
We have recently shown that children with type 1 diabetes have endothelial Foundation, Australia, and the
Wellington Medical Research Founda-
dysfunction17 despite having lower tHcy levels than age-matched controls.18 Endothelial tion, New Zealand.
function in these children relates to red cell folate (RCF).17 We therefore hypothesized that Submitted for publication July 18, 2003;
folic acid would improve endothelial function in children and adolescents with type 1 last revision received Nov 6, 2003;
accepted Dec 30, 2003.
diabetes. We conducted a randomized, double-blind, placebo-controlled crossover trial to Reprint requests: Alexia S Peña, MD,
evaluate the effect of folic acid on endothelial function as assessed by flow-mediated Women’s and Children’s Hospital, 72
dilatation (FMD) in children and adolescents with type 1 diabetes. King William Rd, North Adelaide,
South Australia 5006, Australia.
E-mail: alexia.pena@adelaide.edu.au.
CI Confidence interval Hb Hemoglobin 0022-3476/$ - see front matter
CV Coefficient of variation RCF Red cell folate Copyright ª 2004 Elsevier Inc. All rights
FMD Flow-mediated dilatation tHcy Total plasma homocyst(e)ine reserved.
GTN Glyceryl trinitrate vWF von Willebrand factor
10.1016/j.jpeds.2003.12.049

500
 METHODS Table I. Baseline subject characteristics
Subjects Group A* Group By
Children and adolescents with type 1 diabetes for (n = 21) (n = 15)
a duration of >1 year were recruited consecutively from the
diabetes outpatient clinics at Women’s and Children’s Age (y) 13.7 ± 2.6z 13.9 ± 2.7
Hospital, Adelaide, Australia, and Wellington Hospital, Sex (M/F) 12/9 6/9
Wellington, New Zealand. No subject had background Diabetes duration (y) 6.5 ± 3.3 6.8 ± 3.2
retinopathy on direct funduscopy or microalbuminuria Insulin dose (U/kg/d) 1.17 1.2
measured by overnight albumin excretion rate. Exclusion Weight (kg) 55.2 ± 16.3 56.4 ± 16.5
criteria were history of smoking, vitamin B12 deficiency, or Height (cm) 157.3 ± 14.5 158.1 ± 14.4
current or recent use of folic acid supplements. Sample size Systolic blood pressure (mm Hg) 116 ± 12 116 ± 12
was calculated with an estimated increment of 1.7% on FMD Diastolic blood pressure (mm Hg) 64 ± 7 63 ± 7
with 80% power at a 5% significance level. The Human vWF (%) 100.9 ± 37.5 87.3 ± 16.7
Research Ethics Committee of each hospital approved the RCF (nmol/L [ng/mL]) 855 ± 294 938 ± 295
study. Written informed consent was obtained from the (377 ± 129) (402 ± 187)
parents or guardians, or from the patient if older than 18 years. Serum folate (nmol/L [ng/mL]) 29.1 ± 7.3 30.4 ± 8.5
(12.8 ± 3.2) (12.4 ± 3.3)
tHcy (lmol/L) 5.1 ± 1.5 4.8 ± 1.1
Study Design Vitamin B12 (pmol/L [pg/mL]) 329 ± 145 343 ± 148
A randomized, double-blind, placebo-controlled cross- (445 ± 197) (465 ± 201)
over trial was performed comparing oral folic acid (5 mg daily) HbAlc (%) 9.3 ± 1.6 9.3 ± 1.1
with placebo (both supplied by Sigma Pharmaceuticals, Fasting glucose (mmol/L [mg/dL]) 13.2 ± 5.1 13 ± 5.2
Melbourne, Australia). Randomization was performed by (237 ± 90) (234 ± 93)
using a Fisher table in blocks of 10 (Pharmacy Department, FMD increment (%) 2.78 ± 3.60 4.31 ± 3.38
Women’s and Children’s Hospital). Subjects were randomized GTN increment (%) 15.46 ± 4.53 18.31 ± 6.07
to receive either folic acid or placebo for 8 weeks. There was an Baseline artery diameter (mm) 0.29 ± 0.04 0.28 ± 0.04
8-week washout period, and the subjects then crossed over to
*Started on folic acid followed by placebo.
placebo or folic acid, respectively, for 8 weeks. The folic acid yStarted on placebo followed by folic acid.
dose was chosen by taking previous studies into account.13,19 zMean ± SD.
The subjects were instructed not to take any vitamins during
the study period. There was no systematic change to insulin
to ensure the measurement occurred at the same place for each
dose or diet during the study period. Compliance was assessed
scan. An electrocardiogram was recorded with the ultrasound
by pill counting.
images. Each study included four scans. The first scan was
Four assessments were performed at 0, 8, 16, and 24
taken at rest. Reactive hyperemia was then induced by
weeks—that is, pretreatment and posttreatment with folic acid
occluding arterial blood flow for 4 minutes by using
or placebo. Baseline clinical data were collected (Table I). At
a sphygmomanometer inflated to 250 mm Hg. Arterial flow
each assessment FMD, glyceryl trinitrate (GTN) responses,
velocity was measured by a pulsed Doppler signal at 60 degrees
von Willebrand Factor (vWF), serum folate, RCF, tHcy,
to the vessel during the resting scan and for the first 15 seconds
vitamin B12, fasting glucose, hemoglobin (Hb) A1c, and blood
after deflation of the cuff. The second scan (reactive hyperemia
pressure were measured. All subjects were well at the time of
or FMD) was recorded 30 to 90 seconds after cuff deflation,
assessment.
with measurements between 45 and 75 seconds after deflation.
Ten to 15 minutes were allowed for vessel recovery, and then
Ultrasound Assessment of Endothelial Function the third scan was taken (resting or recontrol scan). The last
The assessments were performed after fasting overnight scan was taken 4 minutes after the sublingual administration
in a quiet and stable temperature environment. FMD and of the GTN spray (400 lg, Nitrolingual spray, G. Pohl-
GTN-induced dilatation were assessed, as we have previously Boskamp, Hohenlockstedt, Germany).
reported.17 The diameter of the brachial artery (2-15 cm above All images were recorded onto high-quality super VHS
the elbow) was measured in longitudinal section from two- videotape and analyzed by an observer blinded to the stage of
dimensional ultrasound images with a 10.0-MHz linear array experiment and the intervention group (folic acid vs placebo).
transducer (Advanced Technology Laboratories, Bothel, For each scan, measurements were made with ultrasonic
Wash) using an Advanced Technology Laboratories HDI calipers for four cardiac cycles, and the measurements were
3000 ultrasound system. Experienced pediatric vascular averaged. All measurements were made incident with the
ultrasonographers performed all studies. Machine operating electrocardiogram R wave (ie, at end-diastole). The
parameters were not changed during any study. A suitable site measurements were averaged and expressed as percentages of
for imaging the vessel was first selected, with reproducible the first control (resting) scan. There were four final average
ultrasonic markers such as venous valves or vessel bifurcations, measurements in total: resting, FMD, recontrol, and after

Folic Acid Improves Endothelial Function in Children and Adolescents

with Type 1 Diabetes 501

Figure. A, FMD changes over time by treatment group. Mean
change of FMD on folic acid treatment was 2.58%, compared with

0.42% on placebo (P < .001). B, Serum folate changes over time by
treatment group (P < .001). d, Folic acid followed by placebo; ,
placebo followed by folic acid. Values are mean ± SE.
8 years were recruited consecutively into the study; 22 began on
GTN. Reactive hyperemia was calculated as flow in the first 15
seconds after cuff deflation divided by the flow during the
resting scan. Our coefficient of variation (CV) between 20
controls was 3.9% for FMD and 4.0% for GTN-induced
dilatation.17
Laboratory Tests
Overnight fasting venous blood samples were drawn in
all subjects before insulin administration. Samples for the
quantitative determination of vWF were collected into tubes
containing 0.109 M trisodium citrate and separated immedi-
ately, and the plasma was frozen at –708C until sample assay.
Plasma vWF was measured by using a commercially available
immunoturbidimetric assay (Diagnostica Stago, Asnieres,
France). Intra-assay CV was 1.9%, and interassay CV was
2.9%. The reference value was 50% to 160%.
Serum and RCF were measured by using ion capture
technology (Abbott IMx analyzer; Abbott Laboratories,
Abbott Park, Ill) with a normal range of 7.7 to 33.9 nmol/L
(3.4-15.0 ng/mL) and 380 to 878 nmol/L (168-388 ng/mL),
respectively. Interassay CV was 5.5%. Vitamin B12 was
measured with a microparticle enzyme immunoassay (Axsym
System; Abbott Laboratories). tHcy was measured by
a fluorescence polarization immunoassay using the commer-
cial IMx Homocysteine assay (Abbott Diagnostic Division,
Abbott Laboratories). HbA1c was measured by using a latex
immunoagglutination inhibition methodology (DCA 2000
Hemoglobin A1c Reagent Kit, Bayer, Toronto, Canada). The
nondiabetic normal range is 4.0% to 6.0%. The method has
502 Peña et al
been correlated with high-performance liquid chromatogra-
phy in our laboratory (r = 0.97). The high-performance liquid
chromatography method is standardized with Diabetes Con-
trol and Complications Trial control sera. Glucose was mea-
sured by the hexokinase spectrophotometry method (Synchron
cx5ce system; Beckman Instruments, Inc, Fullerton, Calif ).
Statistical Analysis
The data were analyzed by using SPSS software version
10.0.7 (SPSS Inc, Chicago, Ill). A comparison of baseline
characteristics between treatment groups assessed the ade-
quacy of randomization. Statistical significance was inferred
with a P value < .05. Spearman rank correlations were used to
compare the change in FMD against the change in tHcy,
HbA1c, and glucose. The analysis was completed by intention
to treat. The effect of the intervention on FMD, GTN
dilatation, baseline artery diameter, vWF, serum folate, RCF,
tHcy, and HbA1c was assessed by using one-sample t tests for
the period effect and independent-sample t tests for the
carryover, period-treatment interaction, and treatment effect.
RESULTS
Thirty-eight subjects (18 male subjects) age 13.6 ± 2.6
folic acid and 16 on placebo. One subject from each group
withdrew from the trial; one had recurrent ketosis, and one did
not wish to continue in the study. There were no important
differences in the baseline characteristics of the two groups as
defined by first treatment received (Table I). All subjects had
normal folate status and lower normal tHcy at baseline. At
baseline, there was no significant correlation between folate
status and HbA1c (RCF, r = –0.06, P = .73; serum folate,
r = 0.1, P = .4, tHcy, r = 0.05, P = .74). In addition, at
baseline, there was no relation between FMD and HbA1c
(r = 0.1, P = .2). Compliance according to pill counting for
placebo and folic acid tablet ingestion was 92%. There were no
adverse effects.
The mean increment in FMD in the 36 subjects on folic
acid was 2.58 (3.1-5.7)% (95% confidence interval [CI], 1.28-
3.88), compared with no significant change in FMD on
placebo (–0.42%; 95% CI, –1.67 to 0.83; P < .001; Figure, A).
FMD after washout period (week 16) did not differ between
the group that started on folic acid and the group that started
on placebo (P = .78), showing the absence of carryover effect
of folic acid on FMD. FMDs at baselines (week 0 and week
16) in both intervention groups were not significantly different
(P = 0.77), confirming the absence of period effect of the
intervention on FMD (Table II).
Change in FMD related to change in serum folate
(r = 0.46, P < .005) and RCF (r = 0.39, P = .02). Change in
FMD was not related to change in tHcy (r = –0.23, P = .17)
or glucose (r = 0.08, P = 0.69). Folate did not affect GTN
brachial artery responses (P = 0.46), baseline artery diameter
(P = 0.27), vWF levels (P = 0.58), or tHcy (P = 0.31; Table
II).
The Journal of Pediatrics  April 2004
Table II. Effects of intervention on endothelial function, folate status, and HbAlc
Baseline After folic acid Baseline After placebo P

FMD (% group A) 2.7 ± 3.6* 5.7 ± 3.0 2.3 ± 3.3 2.4 ± 3.1 < .001
FMD (% group B) 3.4 ± 3.3 5.5 ± 3.8 4.3 ± 3.3 3.1 ± 3.8
GTN (% group A) 15.4 ± 4.5 17.2 ± 5.1 16.4 ± 5.5 17.5 ± 5.0 .46
GTN (% group B) 17.8 ± 4.8 19.5 ± 5.6 18.3 ± 6.0 18.8 ± 5.9
vWF (% group A) 100.9 ± 37.5 101.3 ± 32.2 108.0 ± 39.5 104.0 ± 41.5 .52
vWF (% group B) 106.3 ± 34.6 104.3 ± 26.9 87.3 ± 16.7 97.2 ± 19.5
Serum folate (nmol/L, 29.0 ± 7.2 (12.8 ± 3.2) 39.6 ± 5.4 (17.5 ± 2.4) 33.0 ± 7.2 (14.6 ± 3.2) 32.3 ± 7.9 (14.3 ± 3.5) < .001
group A)
Serum folate (nmol/L, 28.0 ± 7.4 (12.4 ± 3.3) 41.0 ± 5.2 (18.1 ± 2.3) 30.3 ± 8.3 (13.4 ± 3.7) 26.2 ± 8.3 (11.6 ± 3.7)
group B)
RCF (nmol/L, group A) 855 ± 294 (377 ± 129) 1410 ± 439 (622 ± 194) 1205 ± 357 (532 ± 157) 1063 ± 393 (469 ± 173) < .001
RCF (nmol/L, group B) 911 ± 424 (402 ± 187) 1419 ± 441 (626 ± 194) 938 ± 295 (414 ± 130) 1026 ± 406 (453 ± 179)
tHcy (lmol/L group A) 5.1 ± 1.5 4.5 ± 1.1 4.9 ± 1.1 4.6 ± 1.4 .25
tHcy (lmol/L, group B) 4.9 ± 1.0 4.6 ± 1.3 4.8 ± 1.1 5.2 ± 0.9
HbAlc (% group A) 9.3 ± 1.67 9.0 ± 1.5 8.5 ± 1.2 8.7 ± 1.1 .88
HbAlc (% group B) 8.5 ± 1.2 8.4 ± 0.8 9.3 ± 1.4 9.0 ± 1.3
*All values are listed as mean ± SD, with nanograms per mL in parentheses.

HbA1c improved significantly from 0 to16 weeks during
the trial in both groups (9.35% to 8.52%, P < .001)
irrespective of the intervention (placebo or folic acid;
P = .88). Change in HbA1c between week 0 and week 8
was not related to change in FMD during the same time
(r = –0.05, P = .77).
Folic acid increased serum folate by 14 nmol/L (6.2 ng/
mL; 95% CI, 9.2-20.1; P < .001) and RCF by 467.2 nmol/L
(206 ng/mL; 95% CI, 181.4-753.3; P < .001). There was
a carryover effect for serum folate (3.1 nmol/L; 1.3 ng/mL;
95% CI, 0.79-5.54; P = .006) and RCF (189.1 nmol/L; 83.4
ng/mL; 95% CI, 93.3-284.7; P < .001). RCF had a significant
period effect (P = .001). However, serum folate did not show
a significant period effect (Figure, B, Table II).
DISCUSSION
We have shown that short-term folic acid supplemen-
tation improves endothelial function, as measured by FMD, in
children and adolescents with type 1 diabetes and normal
folate status. Mean FMD after folic acid treatment reached
the lower normal range of pediatric controls,17,20 which is
approximately 60% of the mean of our previous age-matched
controls.17
Folate improves FMD in adults with coronary artery
disease,13,19 but there are no data in subjects with diabetes,
apart from the recent demonstration of a beneficial acute effect
of a single folic acid dose on endothelial function, as measured
by forearm plethysmography, in adults with type 2 diabetes.15
FMD is a noninvasive, accurate, and reproducible method for
assessment of endothelial function21 and correlates with
findings in coronary angiography.22
The effect of folate on FMD was independent of any
change in tHcy. Most studies assessing the effect of folate on
endothelial function have investigated this through lowering
tHcy levels,12-14,16 which occurs with less than 1 mg folate
daily, according to previous meta-analysis.23 However,
children with type 1 diabetes have lower tHcy levels than
age-matched controls.18 Our finding of an effect of folate on
endothelial function, independent of tHcy, and a recent
description of acute effects within hours of a single oral folate
dose provide further evidence of a direct effect of folate on the
endothelium.24 Potential mechanisms include antioxidant
actions19,25 or direct interactions with the endothelial nitric
oxide synthase.26
The marker of endothelial activation, vWF, did not
change with folic acid therapy, consistent with findings in
patients with coronary artery disease13 and renal failure.14 We
measured vWF in addition to FMD because we have
previously found a negative correlation between vWF and
serum folate17 and there is increasing evidence that vWF is an
early marker of microvascular disease in children.27
There was a significant improvement in HbA1c over the
study period in both groups without any systematic change of
insulin regimen or diabetes management. Importantly, this
improvement in HbA1c did not have any effect on FMD, as
has been shown in adults with type 2 diabetes.28 The
improvement in HbA1c in all participants can probably be
explained by the effects of study participation on patient
motivation.
Of interest, in comparison with folate intervention trials
in nondiabetic adults,25 our subjects’ serum folate and RCF
increments were lower after supplementation with the same
folate dose. This is probably explained by the higher basal
folate levels. Compliance by pill counting appeared adequate.
We found a strong carryover effect for RCF and
a small carryover effect for serum folate, without a carryover
effect for FMD, suggesting the need for ongoing treatment

Folic Acid Improves Endothelial Function in Children and Adolescents

with Type 1 Diabetes 503
with high-dose folic acid to maintain an effect on FMD—that
is, long-term supplementation will be required to maintain
improved endothelial function. A similar carryover effect of
RCF was demonstrated in a crossover study design in children
with chronic renal failure.16
A sustained increase in serum folate, above the normal
range, appears necessary to maintain the effect of folic acid on
the endothelium. These results, together with the crucial role
of endothelial dysfunction in the development of vascular
disease, raise the testable hypothesis that long-term folic acid
supplementation, beginning early in the course of diabetes,
may reduce the risk of vascular complications.
We thank Professor David Celermajer, Kaye A. Griffiths, and
Michael Skilton for FMD technical support.
REFERENCES
1. Furchgott RF, Zawadzki JV. The obligatory role of the endothelial cells
in the relaxation of arterial smooth muscle by acetylcholine. Nature
1980;288:373-6.
2. Ross R. The pathogenesis of atherosclerosis: a perspective for the 1990s.
Nature 1993;362:801-9.
3. Meeking DR, Cummings MH, Thornet S, Donald AE, Clarkson P,
Crook JR, et al. Endothelial dysfunction in type 1 diabetic subjects with and
without microalbuminuria. Diabet Med 1999;16:841-7.
4. Elhadd T, Khan F, Kirk G, McLaren M, Newton RW, Greene SA, et al.
Influence of puberty on endothelial dysfunction and oxidative stress in young
patients with type 1 diabetes. Diabetes Care 1998;21:1990-6.
5. McLaren M, Elhadd TA, Greene SA, Belch JJ. Elevated plasma vascular
endothelial cell growth factor and thrombomodulin in juvenile diabetic
patients. Clin Appl Thromb Hemost 1999;5:21-4.
6. Cohen RA. Dysfunction of the vascular endothelium in diabetes
mellitus. Circulation 1993;87:67-76.
7. Stehouwer CD, Fischer A, van Kuijk AW, Polak BC, Donker AJM.
Endothelial dysfunction precedes development of microalbuminuria in
IDDM. Diabetes 1995;44:561-4.
8. Makimattila S, Virkamaki A, Groop PH, Cockcroft J, Utriainen T,
Fagerudd J, et al. Chronic hyperglycemia impairs endothelial function and
insulin sensitivity via different mechanisms in insulin-dependent diabetes
mellitus. Circulation 1996;94:1276-82.
9. Brownlee M. Biochemistry and molecular cell biology of diabetic
complications. Nature 2001;414:813-20.
10. Hoogeveen EK, Kostense PJ, Beks PJ, Mackaay AJ, Jakobs C, Bouter
LM, et al. Hyperhomocysteinemia is associated with an increased risk of
cardiovascular disease, especially in non-insulin-dependent diabetes mellitus:
a population-based study. Arterioscler Thromb Vasc Biol 1999;18:133-8.
11. Vaccaro O, Perna AF, Mancini FP, Iovine C, Cuomo V, Sacco M, et al.
Plasma homocysteine and microvascular complications in type 1 diabetes.
Nutr Metab Cardiovasc Dis 2000;10:297-304.
12. Woo KS, Chook P, Lolin YI, Sanderson JE, Metrewelli C, Celermajer
DS. Folic acid improves arterial endothelial function in adults with
hyperhomocysteinemia. J Am Coll Cardiol 1999;34:2002-6.
504 Peña et al
13. Thambyrajah J, Landray MJ, McGlynn FJ, Jones HJ, Wheeler DC,
Townend JN. A randomized double blind placebo-controlled trial of the effect
of homocysteine-lowering therapy with folic acid on endothelial function in
patients with coronary artery disease. J Am Coll Cardiol 2001;37:1858-63.
14. Thambyrajah J, Landray MJ, McGlynn FJ, Jones HJ, Wheeler DC,
Townend JN. Does folic acid decrease plasma homocysteine and improve
endothelial function in patients with predialysis renal failure? Circulation
2000;102:871-5.
15. van Etten RW, de Koning EJ, Verhaar MC, Gaillard CA, Rabellink TJ.
Impaired NO-dependent vasodilatation in patients with type II diabetes
mellitus is restored by acute administration of folate. Diabetologia
2002;45:1004-10.
16. Bennett-Richards K, Kattenhorn M, Donald AE, Oakley G, Varghese
Z, Rees L, et al. Does oral folic acid lower total homocysteine levels and
improve endothelial function in children with chronic renal failure?
Circulation 2002;105:1810-5.
17. Wiltshire EJ, Gent R, Hirte C, Pena A, Thomas DW, Couper JJ.
Endothelial dysfunction relates to folate status in children and adolescents
with type 1 diabetes. Diabetes 2002;51:2282-6.
18. Wiltshire EJ, Thomas DW, Baghurst P, Couper JJ. Reduced total
plasma homocyst(e)ine in children and adolescents with type 1 diabetes.
J Pediatr 2001;138:888-93.
19. Title LM, Cummings PM, Giddens K, Genest JJ Jr, Nassar BA. Effect
of folic acid and antioxidant vitamins on endothelial dysfunction in patients
with coronary artery disease. J Am Coll Cardiol 2000;36:758-65.
20. Jarvisalo MJ, Ronnemaa T, Volanen I, Kaitosaari T, Kallio K, Hartiala
JJ, et al. Brachial artery dilation responses in healthy children and adolescents.
Am J Physiol Heart Circ Physiol 2002;282:H87-92.
21. Sorensen KE, Celermajer DS, Spiegelhalter DJ, Georgakopoulos D,
Robinson J, Thomas O. Non-invasive measurement of human endothelium
dependent responses: accuracy and reproducibility. Br Heart J 1995;74:247-
53.
22. Anderson TJ, Uehata A, Gerhard MD, Meredith IT, Knab S,
Delagrange D, et al. Close relation of endothelial function in the human
coronary and peripheral circulations. J Am Coll Cardiol 1995;26:1235-41.
23. Homocysteine Lowering Trialists’ Collaboration. Lowering blood
homocysteine with folic acid based supplements: meta-analysis of randomised
trials. BMJ 1998;316:894-8.
24. Doshi SN, McDowell IFW, Moat SJ, Payne N, Durrant HJ, Lewis MJ,
et al. Folic acid improves endothelial function in coronary artery disease via
mechanism largely independent of homocysteine lowering. Circulation
2002;105:22-6.
25. Doshi SN, McDowell IF, Moat SJ, Lang D, Newcombe RG, Kredan
MB, et al. Folate improves endothelial function in coronary artery disease: an
effect mediated by reduction of intracellular superoxide? Arterioscler Thromb
Vasc Biol 2001;21:1196-202.
26. Stroes ES, van Faassen EE, Yo M, Martasek M, Boer P, Govers R, et al.
Folic acid reverts dysfunction of endothelial nitric oxide synthase. Circ Res
2000;86:1129-34.
27. Verotti A, Greco R, Basciani F, Morgese G, Chiarelli FL, von
Willebrand. factor and its propeptide in children with diabetes: relation
between endothelial dysfunction and microalbuminuria. Pediatr Res
2003;53:382-6.
28. Bagg W, Whalley GA, Gamble G, Drury PL, Sharpe N, Braatvedt GD.
Effects of improved glycaemic control on endothelial function in patients with
type 2 diabetes. Intern Med 2001;31:322-8.
The Journal of Pediatrics  April 2004