Urinary glycosaminoglycan excretion in urolithiasis - Archives of ...

Arch Dis Child 1999;80:271–272 271
Department of
Biochemistry,
Cerrahpas¸a Medical
Faculty, Istanbul
University, Turkey
T Akçay
D Konukog˘lu
Y Dínçer
Correspondence to:
Professor T Akçay, Kayıs¸dag˘ı
Caddesi Akçay Apt No
156/4, Kadıköy, Istanbul,
Turkey.
Accepted 20 October 1998
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Urinary glycosaminoglycan excretion in urolithiasis
Tülay Akçay, Díldar Konukog˘lu, Yildiz Dínçer
Abstract
Urinary glycosaminoglycan (GAG) excretion
was measured in children with idiopathic
urolithiasis (15 girls and 10 boys;
mean (SD) age 6.2 (2.4) years) and in
healthy controls (10 girls and 14 boys; mean
(SD) age 6.8 (3.8) years). GAG excretion
was expressed as a GAG/creatinine (mg/g)
ratio and was evaluated using dimethylmethylene
blue. In healthy control children,
the mean (SD) GAG/creatinine ratio
was 31.67 (12.76) and it was similar in girls
and boys. The children with idiopathic urolithiasis
had significantly lower mean (SD)
GAG/creatinine ratios than controls (22.59
(7.35)). Therefore, urinary GAG excretion
may be important in the disease process in
children with urolithiasis, as it is in adults.
(Arch Dis Child 1999;80:271–272)
Keywords: glycosaminoglycans; urolithiasis; renal stones
The urinary inhibitors of crystal nucleation,
growth, and aggregation play an important role
in urolithiasis. 1 Such inhibitors are presumed
to aVord protection against the formation of
stones in normal individuals. Their deficiency
in, or absence from, the urine of patients with
stones is thought to predispose these individu-
als to the disease.
1 2
In vitro studies—except those of Grases and
Costa-Bauza, 3 who reported that pentosan
polysulphate promoted calcium oxalate dihydrate
crystal formation in synthetic urine—
have shown that glycosaminoglycans (GAGs)
are potent inhibitors of crystal growth and
aggregation.
4 5
There have been few studies on urinary
GAG excretion in various forms of childhood
nephrolithiasis. 6–8 therefore, we have attempted
to establish whether the diVerences that we
described previously between controls and
adults with renal stones 9 are also present in
children with urolithiasis.
Subjects and methods
We studied 15 girls and 10 boys, mean (SD)
age 6.2 (2.4) years, with idiopathic renal stone
disease. The calculi were composed of calcium
oxalate (n = 18) or calcium phosphate (n = 7).
The children had taken no medication for at
least two weeks, and were not suVering from
malabsorption, either as a result of tubular acidosis
or malformations of the urinary system.
The control group consisted of 24 healthy children
(10 girls and 14 boys; mean (SD) age 6.8
(3.8) years).
Written consent was obtained from parents.
All subjects had normal renal function, blood
pH, and serum concentrations of parathyroid
hormone, uric acid, sodium, potassium, chlo-
ride, magnesium, calcium, and phosphorus.
Routine urine examinations were normal and
urine was sterile in all subjects.
Patients and controls were placed for three
days on a standard diet containing predetermined
amounts of calories, proteins, and
mineral salts in proportion to age. At the end of
the third day, 24 hour urine specimens were collected
at 4°C without preservatives. On arrival at
the laboratory, the total volume and relative
density of each sample were measured and general
urine analysis tests were carried out.
Calcium and magnesium measurements
were performed by atomic absorption spectrophotometry.
Uric acid, oxalate, and phosphate
concentrations were measured by means of
conventional enzymatic kits.
For GAG measurement, the GAGs were
precipitated with cetylpyridinium chloride and
then reacted with dimethylmethylene blue to
produce a complex with the polyanionic
molecule of sulphated GAGs. 10 The GAG
results were expressed as a GAG/creatinine
(mg/g, respectively) ratio. All chemical materials
were purchased from Sigma Chemicals.
Statistical analysis was performed using the
Student’s t test for unpaired data.
Results
Table 1 lists the concentrations of urinary calcium,
magnesium, phosphate, oxalate, and uric
acid, and pH values. Figure 1 shows the GAG/
creatinine ratios.
There were no significant diVerences in the
excretion of calcium, magnesium, phosphate,
oxalate, and uric acid between the patients and
controls.
Table 1 Urinary calcium, magnesium, oxalate, phosphate,
and uric acid concentrations, and pH values in controls and
patients with urolithiasis
Controls (n = 24) Patients (n = 25)
Calcium 2.70 (1.23) 2.93 (1.32)
Magnesium 2.43 (0.90) 2.41 (1.20)
Oxalate 0.40 (0.13) 0.42 (0.16)
Phosphate 41.5 (8.1) 37.6 (10.5)
Uric acid 4.45 (1.09) 4.70 (1.50)
pH 6.90 (0.50) 7.20 (0.30)
Concentrations are mean (SD) in mmol/l.
35
30
25
20
15
10
5
*
0
Controls Boys Girls Patients
Figure 1 Mean (SD) urinary GAG/creatinine ratio
(mg/g) in controls and in patients with urolithiasis.
*Comparison with controls (p < 0.005).

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272 Akçay, Konukog˘lu, Dínçer
The control subjects had a mean (SD)
GAG/creatinine ratio of 31.67 (12.76). We
found no diVerences between girls and boys in
mean (SD) urinary GAG/creatinine ratio
(33.34 (9.43), 30.39 (14.69), respectively). In
patients, the mean (SD) urinary GAG/
creatinine ratio was significantly lower (22.59
(7.35)) than in control subjects (p < 0.005).
Discussion
In adults with calcium stones a higher urinary
excretion of calcium, oxalate, and urate as well
as a deficiency in inhibiting substances has
been reported repeatedly. There have been
numerous reports showing that urinary GAGs
are low in adult patients with nephrolithiasis.
1 3 11 12 We also found that urinary GAG
concentrations were significantly lower in
adults with urolithiasis. 9 However, although
Michelacci et al suggested that urinary GAG
excretion in children with urolithiasis was
significantly lower, 6 these data have not been
confirmed by others.
7 8
The mean (SD) urinary GAG concentrations
in 15 healthy children were reported as
17.00 (15.60) mg/day by Lama et al. 7 However,
Baggio et al reported that the mean (SD) GAG
concentrations in healthy children were 61.26
(17.94) mg/l and found no significant diVerence
in urinary GAG excretion between
children with idiopathic urolithiasis and
healthy controls. 8 Michelacci et al reported a
mean (SD) GAG/creatinine (mg/g) ratio of
24.33 (1.91) in healthy children 6 ; this result is
close to our findings of 31.67 (12.76).
Discrepancies between data from diVerent
studies might be caused by the diVerent methods
used to measure GAGs. In the literature,
the measurement of urinary GAG concentrations
is commonly performed by the
borate−carbazole method or other procedures
involving the basic metachromatic dye, alcian
blue. The borate−carbazole method measures
the hexuronic acid residues of GAG molecules
and, therefore, cannot detect keratan sulphate
because the hexuronic acid residues are
replaced with galactose in keratan sulphate. 10
In addition, other procedures involving the use
of alcian blue are not specific for urinary GAGs
because negatively charged molecules, other
than sulphated GAGs, interfere with the
assay. 13 Finely dispersed precipitates obtained
with alcian blue are often diYcult to
harvest. 13 14 On the other hand, Hesse et al
indicated that there was a diurnal rhythm in
urinary GAG excretion. 15 As GAG excretion
was highest in the morning and lowest at night,
24 hour urine collection would avoid falsely
high results. Therefore, we performed the
GAG measurements in 24 hour urine specimens
and we also calculated total GAG
concentration by using the GAG/creatinine
ratio in 24 hour urine samples, which gives
more reliable results. 16
A procedure involving the use of the basic
metachromatic dye dimethylene blue, which
we used for the first time to determine GAG
concentrations in the urine of children with
stones, is reported to be more sensitive than the
other methods used to analyse urinary GAG
concentrations in patients with urolithiasis. 12
Our data lead us to propose that urinary
GAG may play an important role in the
prevention and reduction of calculi in children,
as has been found in adults. 9
The authors thank the Department of Paediatrics of Cerrahpas¸a
Medical School for their clinical assistance.
1 Sindhu H, Hemal A K, Thind S, Rath R, Vaidyanathan S.
Comparative study of 24 hours urinary excretion of
glycosaminoglycan by renal stone formers and healthy
adults. Eur Urol 1989;16:45–7.
2 Akçay T, Konukog˘lu D, Çelik Ç. Hypocitraturia in patients
with urulithiasis. Arch Dis Child 1996;74:350–1.
3 Grases F, Costa-Bauza A. Study of factors aVecting calcium
oxalate crystalline aggregation. BrJUrol1990;66:240–4.
4 Fellstrim B, Danielson BG, Karlsson FA, Ljunghall S. Crystal
inhibition: binding of heparin and chondroitin sulphates,
heparin, pentosan polysulphate and Tamm-Hosfall
glycoprotein. Urol Res 1984;12:81–3.
5 Kohri K, Garisde Y, Blacklock NJ. The eVect of glycosaminoglycans
on the crystallisation of calcium oxalate.
BrJUrol1989;63:584–90.
6 Michelacci MY, Glashan BQ, Schor N. Urinary excretion of
glycosaminoglycans in normal and stone forming subjects.
Kidney Int 1989;36:1022–8.
7 Lama G, Gariella Carbone M, Marrone N, Russo P, Spagnuolu
G. Promoters and inhibitors of calcium urolithiasis in
children. Child Nephrol Urol 1994;10:81–4.
8 Baggio B, Gambro G, Favaro S, et al. Juvenile renal stone
disease: a study of urinary promoting and inhibiting
factors. J Urol 1981;130:1133−5.
9 Akçay T, Erbas¸ M, Konukog˘lu D. Urinary excretion of glycosaminoglycans
in normal individuals and patients with
renal stones. Med Sci Res 1994;22:77–8.
10 Pannin G, Naia S, Dallw’Amica R, Chlandetti L, Zachello
F, Catassi C. Simple spectrophotometric quantification of
urinary excretion of glycosaminoglycan sulphates. Clin
Chem 1986;32:2073–6.
11 Nikkila M. Urinary glycosaminoglycans excretion in normal
and stone-forming subjects. Significant disturbance in
recurrent stone formers. Urol Int 1989;44:157–9.
12 Jong JGN, Wevers RA, Laarakkers C, Poortuis BJHM.
Dimethylmethylene blue-based spectrophotometry of glycosaminoglycans
in untreated urine: a rapid screening procedure
for mucopolysaccharidoses. Clin Chem 1989;35:
1472–7.
13 Whitley CB, Ridnour MD, Draper KA, Dutton CM, Neglia
JP. Diagnostic test for mucopolysaccharidoses I. Direct
method for quantifying excessive urinary glycosaminoglycan
excretion. Clin Chem 1989;35:374–9.
14 Pennock CA. A review and selection of simple laboratory
methods used for the study of glycosaminoglycan excretion
and diagnosis of the mucopolysaccharidoses. J Clin Pathol
1976;29:111–23.
15 Hesse A, Wuzel H, Vanlensieck W. The excretion of
glycosaminoglycans in the urine of calcium-oxalate-stone
patients and healthy persons. Urology 1986;41:81–7.
16 Goldberg JM, Catlier E. Specific isolation and analysis of
mucopolysaccharides (glycosaminoglycan) from human
urine. Clin Chem Acta 1984;41:19–21.

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Urinary glycosaminoglycan excretion in
urolithiasis
Tülay Akçay, Díldar Konukoglu and Yildiz Dínçer
Arch Dis Child 1999 80: 271-272
doi: 10.1136/adc.80.3.271
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