|
|
|
||||||
|
|
|||||||
|
URINARY HEPARAN SULPHATE IN DIABETES URINARY HEPARAN SULPHATE IS INCREASED IN NORMOALBUMINURIC DIABETIC PATIENTS ALICIA E. ELBERT1, ANA MARIA PAGLIONE2, JULIO C. BRAGAGNOLO1,
HECTOR A. MAINETTI1, CARLA D. BONAVITA2, MAXIMINO RUIZ1 División Diabetología, Hospital de Clínicas José de San Martín. Facultad de Medicina, Universidad de Buenos Aires; 2 Laboratorio de Lípidos y Lipoproteínas, Departamento de Bioquímica Clínica. Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires Resumen Aumento de heparan sulfato urinario en diabéticos normoalbuminúricos. Se estudiaron 49 pacientes diabéticos: 22 hombres y 27 mujeres. Se determinaron: heparán sulfato (HS) urinario, albuminuria, creatininemia, creatininuria, clearance de creatinina, HbA1c y presión arterial (PA). Los pacientes se clasificaron en dos grupos: grupo 1, portadores de Diabetes Mellitus (DM) Tipo 1 (n = 16), y grupo 2, portadores de DM Tipo 2 (n = 33). Se utilizaron 24 sujetos sanos como controles, 12 hombres y 12 mujeres los cuales mostraron un valor medio ± DS de HS de 0.36 ± 0.18 mg/24 hs. Se hallaron diferencias significativas entre hombres y mujeres; 0.43 ± 0.15 versus 0.28 ± 0.17 respectivamente; ( p = 0.02). La población diabética dio un valor medio de 0.68 ± 0.44 el cual comparado con el de los controles mostró una significancia de p < 0.001. Los pacientes hombres tuvieron un valor medio de 0.82 ± 0.48 y las pacientes mujeres 0.54 ± 0.35; p < 0.02. No se encontraron datos significativamente diferentes entre los grupos 1 y 2. Los valores de HS no se correlacionaron con la edad, índice de masa corporal (IMC), tiempo transcurrido desde el comienzo de la diabetes, albuminuria, creatininemia, creatininuria, clearance de creatinina, HbA1c ni PA. Se concluye que: tanto en controles como diabéticos los hombres eliminan más HS urinario que las mujeres y que los pacientes diabéticos normoalbuminúricos eliminan más HS urinario que los controles, independientemente del tiempo transcurrido desde el inicio de la diabetes y la PA.
Abstract Forty-nine normoalbuminuric diabetic patients were studied: 22 males and 27 females, in whom urinary heparan sulphate (HS), albuminuria, creatininemia, creatininuria, creatinine clearance, HbA1c and arterial pressure (AP) were determined. Two groups were discerned: group 1, Type 1 DM, diabetic cases (n = 16); and group 2, Type 2 DM diabetic cases (n = 33). Patients were compared with 24 healthy controls: 12 men and 12 women, who showed a mean value ± SD of 0.36 ± 0.18 mg/24 h HS with significant differences between males and females (0.43 ± 0.15 versus 0.28 ± 0.17, respectively; p = 0.02). The total population of diabetic cases rendered a mean of 0.68 ± 0.44 and comparison with controls proved highly significant (p < 0.001). Globally, male patients had a mean of 0.82 ± 0.48 and females 0.54 ± 0.35, with p < 0.02. Group 1 and 2 values of HS were not significantly different. HS levels failed to correlate either with age, body mass index (BMI), time since onset of diabetes, albuminuria, creatininemia, creatininuria, creatinine clearance, HbA1c or arterial hypertension. To conclude: both normal and diabetic males eliminate a greater quantity of HS than females. Normoalbuminuric diabetic patients of both types eliminate a greater quantity of HS regardless of arterial pressure and time since onset of diabetes. Key words: heparan
sulphate, diabetes, normoalbuminuria Diabetic
nephropathy is the first cause of admission to supportive renal
treatment, and is represented by 30-40% of insulin-dependent (Type
1)1
and 5-10% of non-insulin-dependent (Type 2) patients in the course
of diabetes2. In
developed countries 30-40% of patients on hemodialysis are diabetic,
which carries an extremely high social and economic burden3,
4. Over
the last few years, attempts have been made to detect early markers
of nephropathy in order to initiate prompt treatment and thus avoid
the development of renal insufficiency, since patients with impaired
renal function present greater morbidity and mortality than the
general population5. In
spite of the efforts and multiple studies, to date it has not been
possible to identify reliably the factors that allow prevention of
this pathology. Microalbuminuria is considered an early marker of
renal lesion, and its determination is useful to decide the
initiation of treatment for incipient nephropathy6.
In order to optimise such therapy it has been attempted to detect
diverse even earlier markers than microalbuminuria, such as prorenin7
and Na/Li cotransport8. The
glomerular barrier is made up of fenestrated endothelium, basal
membrane and endothelial and epithelial cells. In turn, the
glomerular basal membrane (GBM) consists of collagen IV and V,
laminin, enactin, heparan and chondroitin sulphate proteoglycans9. This barrier is selectively permeable to the passage of molecules
(albumin, globulins), restricting the passage according to their
load, size and charge. Membrane heparan sulphate (HS) seems to play
a major role in such restriction, as confirmed by injecting
monoclonal antibodies against the glycosaminoglycan (GAG) chain of
HS10, 11,
with alteration of GMB contents and appea-rance of albuminuria12, 13. In
other studies, alterations have been observed in the selective
permeability of the membrane through urine analysis, where a
modification in the immunoglobulin/albumin ratio was found, as well
as differences between neutral immunoglobulins and those charged
negatively14. HS
is synthesised by several cell types: vascular endothelium, smooth
muscle and glomerular or mesangial epithelial cells. Its function is
to maintain the structural integrity of the GMB and its negative
charge. It binds to lipoproteinlipase and superoxide dismutase,
which produces a regulatory effect on lipids and thrombogenesis,
besides affecting the regulation of cellular growth15. The
hypothesis advanced at Steno suggests that diabetes produces a
modification in HS metabolism and that such alteration is the key to
renal and vascular pathology16. Several
experiments carried out in diabetic rats and humans show that
urinary excretion of GAGs and more specifically of HS are increased.
Reddi et al17
measured HS in diabetic rat glomeruli and observed a decrease in
level. In
Type 1 DM patients with microalbuminuria exceed-ing 200 mg/24 hours,
Vernier et al40
detected a 30-40% decrease in anionic sites digestible by heparinase
in the kidney. Bonavita et al.22
studied HS
urinary excretion in 15 diabetic juvenile, 15 diabetic adults and 44
normal controls.It was found that youngsters of both sexes excrete
more HS than adults and that both normal males and diabetic
juveniles eliminate more HS than females. Diabetic adults of both
sexes also eliminate a greater percentage of HS, more than normal
controls. Baggio et al.23
determined
urinary GAGs in 40 Type 1 patients with normoalbuminuria and found
that they excreted a greater quantity of GAGs. Perez Blanco et al.24 investigated
urinary GAG excretion in healthy individuals, patients with neither
hypertension nor microalbuminuria, those without hypertension but
with microalbuminuria and those with both microalbuminuria and
hypertension. The latter cases showed the highest values; in the
second group GAGs were greater than in controls and in the third
levels were significantly higher. In
the present work we attempted to determine urinary HS elimination in
normoalbuminuric Type 1 DM and Type 2 DM patients (albuminuria <
30 mg/24 hs), and its correlation with risk factors of diabetic
nephropathy, time since onset of diabetes, glycosylated hemoglobin
(HbA1c) and arterial hypertension. Patients
and Methods Forty-nine diabetic patients, all normoalbuminuric (albuminuria < 30
mg/24 hs), were studied to determine HS in urine. There were 27
females and 22 males. Two groups were discerned according to the
American Diabetes Association25:
Group I (Type 1 DM n = 16); and Group 2 (Type 2 DM n = 33). Patients
were compared with 24 healthy controls, comprising 12 males and 12
females. Diabetic
patients were free of any associated pathology, liver or thyroid
disease, decompensated cardiac insufficiency or infectious process.
Pregnant women were excluded. Clinical
features of controls and diabetic patients are shown in Table 1.
Twenty four hour urine was collected without addition of preserving
agents and cooled to 4°C. Sample collection was contraindicated in
patients who were febrile, while relative impediments included
physical exercise and liquid intake during the night. Briefly,
HS was determined as follows22:
40 ml aliquots were precipitated with
bromo-hexadecyltrimethylammonium (cetrimide USP, Sigma). The
precipitate was removed by repeated ethanol washes. Material
insoluble in ethanol was dissolved in 2.5 ml of water, centrifuged
and passed through a 1 x 4 cm 50 x 2, H+
Dowex column, and thereafter eluted with distilled water. The acidic
effluent was neutralised with 0.5 N NaOH and lyophilised; then an
aliquot of the lyophilised material reconstituted with distilled
water was taken and an equal volume of 1 N HCI was added, heating
the mixture at 110°C during 2 h; duplicate aliquots were then drawn
to measure glucosamine. For this reaction, desulphation and
deamination with nitrous acid was performed according to Lagunoff
and Warren26;
lastly, produced 2-5 anhydrohexose was quantified with
chloro-3-methyl-2-benzothiazolone according to Smith and Gilkerson27, using a glucosamine standard. Results are expressed in mg of
glucosamine per 24 hours. Method CV% was 19.4%. Albuminuria
was determined in 24 hours urine by radioimmunoassay using a
commercial device (Albumin RIA, DPC, USA)28.
In this technique the patient’s urinary albumin competes with 125I-labeled
urine for the binding sites of an antibody. After incubation free
and bound fractions were separated with an anti gamma globulin
diluted with polyethy-lenglycol. The bound fraction was precipitated
by centrifugation and quantified in a gamma counter. Urine sample
concentrations were determined in comparison with a calibration
curve (from 0 to 60 µg/ml) that was processed simultaneously.
Method detection limit is 0.3 µg/ml. Cut-off value of
normoalbuminuria was taken as 30 mg/24 hs. method
CV% was 2.8%. Besides,
creatinine in blood was determined by a reaction with picric acid in
alkaline medium to render a coloured complex which is quantified by
spectrophotometric reading29. method CV% was 2.4%. For
determination in urine, samples were diluted 1/10 and 1/50 in
distilled water. HbA1c
was determined with a DCA 2000 commercial kit from Bayer
Diagnostics, which used a system based on the inhibition of the
agglutination of latex particles coated with a specific monoclonal
antibody against HbA1c. Reference values were 4.3-5.7% of total
hemoglobin. Method CV% was 3.3. Arterial
pressure was taken with a sphingomanometer after 5 minuts resting
and defined as greater than 140 mm Hg and greater than 90 mm Hg as
arterial hypertension. Statistical
analysis. Quantitative variables were expressed as means ± SD. SD
values between group pairs were analysed by Student’s t test for
independent samples or by the non-parametric Kruskal-Wallis method
when irregular distribution were present. The correlation between
quantitative variables was studied by Pearson’s linear regression
method. A
multiple linear regression with dependent variable HS was carried
out with the following independent variables: age, gender,
creatinine, arterial hypertension and time since onset of diabetes,
as well as 24 h microalbuminuria and albuminuria. Results Table
1 provides clinical and laboratory data for diabetic and control
groups. Figure 1 shows HS data distribution observed in diabetic
patients and controls. Mean HS value ± SD for controls was 0.36 ±
0.18 mg/24 hs and 0.68 ± 0.44 mg/24 hs for the total diabetic
population. Mean
HS value in the control group displayed a significant difference
according to the t test between males and females (0.43 ± 0.15
versus 0.28 ± 0.17 mg/24 hs respectively; p = 0.02) (Fig. 2). In
the diabetic group, males had a mean of 0.82 ± 0.48 mg/24 hs and
females 0.54 ± 0.35 mg/24 hs (p < 0.02). If
HS levels are compared in diabetic men versus male controls, the
significant difference in greater elimination in the former is
maintained (p = 0.001), as well as in diabetic women versus their
controls (p = 0.02), both according to Kruskal-Wallis (data not
shown). Figure
3 shows mean HS values in controls versus diabetic patients with a
highly significant p < 0.001 When
adjustment was made for age, diabetic patients presented a mean
value of 0.66 mg/24 hs and controls 0.35 mg/24 hs; with p = 0.002
(data not shown). On
comparing HS value in the Type 1 DM versus the Type 2 DM diabetic
group (0.75 ± 0.52 mg/24 hs and 0.60 ± 0.36 mg/24 hs,
respectively), the difference was not significant. There
was no correlation between HS level with factors such as age, time
since onset of diabetes, BMI, albumin level and HTA, or with
creatinine clearance, while a positive correlation with HbA1C was
not found. Discussion According
to Mogensen6 incipient nephropathy has its onset roughly towards the fifth year
following the development of microalbuminuria in the course of
diabetes. The features of its course are predictable, with
microalbuminuria progressing to proteinuria, then to azotemia and
finally to chronic renal insufficiency (CRI), with speeding up of
successive stages when the management of arterial pressure or
metabolic control is unsuitable. Over
the last few years, research has been focused on studying early
markers of chronic complications in order to determine accurately
which patients are prone to develop such disorders. From the
clinical point of view, microalbuminuria30
has been the component taken into account as a marker of renal
lesion31,
of nephropathy development32,
or cardiovascular risk and of generalized vascular lesion33. The
Steno hypothesis16, as well as work demonstrating the greater prevalence of renal
complications in familial groups34,
in addition to metabolic and haemodynamic factors, support the
existence of genetic factors in its development. Heparan
sulphate proteoglycan (HS-PG) is a major component of the glomerular
basal membrane (GBM) that plays a leading role as an organizing and
structural molecule35. The presence of this strongly negative molecule is essential to
maintain the selective permeability of GMB36.
The loss of HS has been associated with proteinuria in numerous
glomerulopathies37,
38.
Despite their normoalbuminuric status, kidney biopsy in patients
with diabetes during this period discloses alterations in the
extracellular matrix with GMB thickening and an increase in the
mesangial matrix. Changes in the extracellular matrix with loss of
the heparan sulphate proteoglycan seem to play a leading role in
protei- nuria35. Rohrabach
et al reported reduction in HS synthesis in diabetic rats36,
while others demonstrated a decrease in tissue by means of
antibodies directed against the HS-PG in the basal membrane10. The
results obtained in our study in urine from normoalbuminuric
diabetic patients show increased levels of HS, significantly greater
in men that in women, and such difference persists with age
adjustment. It may therefore be concluded that age difference seems
not to be a factor influencing the increase in urinary HS. Other
studies evaluating the elimination of diverse proteoglycan
components in normoalbuminuric patients also displayed some
differences. Craddock et al20
studied in urine the fraction designated Astrup, consisting mainly
of acid mucopolysaccharides with a predominance of hexuronic acid
and hexosamine. They also found that such fraction is excreted in
greater proportion by diabetic subjects with normal renal function
that by normal controls. Lubec
et al21
evaluated urinary GAGs by means of the reaction with carbazol in
diabetic youths, to find that they excreted more GAGs as compared
with controls. They also determined HbA1c and recorded a highly
significant correlation with GAGs (r = 0.7; p < 0.01). Reddi17
studied Wistar rats, to find that the group treated with
streptozotocine presented lower glomerular GAG and HS concentrations
as evidenced by reduced incorporation of 35S
sulphate, which would indicate decreased glomerular synthesis.
Furthermore, these rats presented greater urinary HS elimination
than controls. The correlation of urinary HS versus proteinuria was
significantly positive. The author concluded that glomerular HS
synthesis was reduced concomitantly with increased urinary
elimination of the proteoglycan. The
relationship between HS excretion and albuminuria remain unclear. McAuliffe
et al.18 measured
urinary HS excretion in diabetic and non diabetic subjects with
varying degree of albuminuria. The urine was collected with the
subject at rest over a 2.5 h period after a 300 ml water load.
Categorizing for albuminuric status shows that the diabetic micro
and macroalbuminuric groups have a significantly higher HS excretion
rate than non diabetic subjects. In
Shield et al19
article both groups of diabetic patients with and without
microalbuminuria (15 µg/min in at least two of three consecutive
urine collections) had significantly elevated excretion HS when
compared to normal individual, there was no difference in HS
excretion between diabetic subjects with and without
microal-buminuria. It
has been debated whether the mechanism of HS decrease is due to an
alteration in synthesis, to a change in its composition or to the
loss of the membrane proteoglycan by urine; however, in every case
there is a decrease in HS concentration at the glomerular barrier. The
correlation in our study between albuminuria and HS excretion was
not significant. Interestingly, though, the albuminuria level
(significantly different in diabetic patients vs controls, with p
< 0.001), while remaining within ranges considered normal, should
be regarded as an influential factor in the elevation of urinary HS
elimination. Converting
enzyme inhibitors have been widely used for their multiple effects
in delaying the progression of renal function decline. Reddi et al35
observed that these drugs increased GBM HS and reduced its
elimination, a mechanism perhaps operative on the GBM which favours
its effect on microalbuminuria, also supporting the major role
played by HS in the development of diabetic nephropathy. Heparan
sulphate is a strong inhibitor of mesangial growth and its reduction
has been demonstrated in diabetic patients with mesangial expansion
and clinical nephropathy39. In other work in humans a negative correlation was demonstrated between
GBM anionic groups and albumin excretion40. No
correlation was found in our study with risk factors such as
arterial pressure levels. The high pressure seems related with the
deterioration in renal function but its increase is described when
urinary albumin exceeds 100 mg/24 hs40.
Our patients had less albuminuria. In
other studies performed in normoalbuminuric patients high arterial
pressure levels were only found when evaluation was carried out by
more sensitive methods such as pressurometry41, not employed here. In
our population, there was no correlation between HS and the time
since onset of diabetes and no positive correlation with HbA1c was
found. In all likelihood, other as yet undetermined variables,
particularly genetic factors, influence urinary HS elimination. On
the other hand, there was a trend that failed to reach statistical
significance with creatinine values. The
leading role of the glycosaminoglycan in the pathogenesis of the
nephropathy would gain support from work showing the possibility of
reducing microalbuminuria by means of oral therapy with
glycosaminoglycan42. The
increase in urinary HS in our study failed to correlate with any of
the risk parameters considered for the development of the diabetic
nephropathy such as time since onset of diabetes, arterial
hypertension or HbA1c. This finding agrees with the theory of Steno
suggesting that extracellular matrix impairment alters endothelial
GBM in a subgroup of diabetic patients with greater genetic
susceptibility to develop the renal complication43.
The answer could be obtained through a study with a greater number
of patients followed up prospectively, confirming the subsequent
appearance of microalbu-minuria in those presenting increased HS. The
loss of HS-PG modifies capillary permeability leading to an increase
in albuminuria levels. Perhaps the early appearance of HS in urine
could predict patients likely to present microalbuminuria, but this
would have to be studied prospectively during follow-up of patients
showing increased HS levels. In
conclusion, our results show that both normal and diabetic males
eliminate a greater quantity of HS than females, normoalbuminuric
diabetic patients of both types eliminate a greater quantity of HS
without any correlation with the evaluated risk factors. The
increased HS in longitudinal follow-up of this subgroup and
confirmation of their progress towards microalbuminuria would
support the leading role of glycosaminoglycan in the pathogenesis of
diabetic nephropathy. Acknowledgements:
This work has been supported by grant FA 081, 1995-1997 Program,
from the University of Buenos Aires. References 1. Deckert T, Poulsen JE, Larsen M. Prognosis of diabetics with diabetes onset before the age of thirty one. Diabe-tologia 1987; 14: 371-4. 2.
Viberti GC. Recent
advances in understanding mechanisms and natural history of diabetic
renal disease. Diabetes Care 1988; 11 (Suppl I): 3-9.
3. The USRDS
1995 Annual Data. Report III. Incidence and causes of treated ESRD. Am
J Kidney Dis 1995; 26 (Suppl 2): 539-50.
4. The USRDS
1995 Annual Data. Report II. Prevalence of ESRD therapy. Am J
Kidney Dis 1995; 26 (Suppl 2): 530-8.
5. Almad T,
Kirsten N, Feldt-Rasmussen B, Deckert T. The predictive value of
microalbuminuria in IDDM. Diabetes Care 1994; 17:
120-4.
6. Mogensen
CE. Microalbuminuria as a predictor of clinical diabetic
nephropathy. Kidney Int 1987; 31: 673-89.
7. Luetscher
JA, Kralmer F. Microalbuminuria and increased plasma prorenin. Arch
Int Med 1988; 148: 937-41.
8. Trevisan
R, Nosadini R, Fioretto P, Semplicini A, Donado V, Doria A, et al. Clustering
of risk factors in hypertensive insulin dependent diabetics with
high sodium-lithium countertransport. Kidney Int 1992; 41:
855-61.
9. Craig
Tisher C, Madsen KM. Anatomy of the kidney. In: Brenner BM, Rector
FC, eds. The Kidney. 5th edn. Philadelphia: WB Saunders, 1996; 1-71. 10.
Tamsma JT, Van Der Bom JF, Bruijn AJ, Assmann JK, Weening JJ.
Expression of glomerular extracellular matrix components in human
diabetic nephropathy: decrease of heparan sulphate in glomerular
basement membrane. Diabetologia 1994; 37: 313-20. 11.
Van Der Bom J, Lambert PWJ, Bakker M, Veerkamp HJ. A
monoclonal antibody against GBM heparan sulfate induces an acute
selective proteinuria in rats. Kidney Int 1992; 41: 115-23. 12. Olgemoller L, Schwaabe S, Gertitz D, Schleicher L. Elevated glucose decreases the content of a basement membrane associated heparan sulphate proteoglycan in proliferating cultured porcine mesangial cells. Diabetologia 1992; 35: 183-6. 13. Tamsma
JT, Van Der Woude FJ, Lemkes HHPJ. Effect
of sulphated glycosaminoglycans on albuminuria in patients with
overt diabetic (type 1) nephropathy. Nephrol Dial Transplant
1996; 11: 182-5. 14.
Deckert T, Feldt Rasmussen B, Djurup R, Deckert M. Glomerular
size and charge selectivity in insulin dependent diabetes mellitus. Kidney
Int 1988; 33: 100-6. 15.
Saxena U, Klein MG, Goldberg IJ. Identification and
characterization of the endothelial cell surface lipoprotein lipase
receptor. J Biol Chem 1991; 266: 17516-21. 16.
Deckert T, Feldt Rasmussen B, Borch Johnsen K, Jensen T,
Kofoed-Enevoldsen A. Albuminuria reflects widespread vascular
damage. The Steno Hypothesis. Diabetologia 1989, 32: 219-26. 17. Reddi AS. Glomerular and urinary glycosaminoglycans in diabetic rats. Clinica Chimica Acta 1990; 189: 211-20. 18. McAuliffe
VV, Fisher EJ, McLennan SV, Yue DK, Turtle JR. Urinary
glycosaminoglycan excretion in NIDDM subjects: its relationship to
albuminuria. Diabet Med 1996; 13: 758-63. 19.
Shield JH, Carradus M, Stone JE, Hunt LP, Baum JD, Pennock
CA. Urinary heparan sulphate proteoglycan excretion is abnormal in
insulin dependent diabetes. Ann Clin Biochem 1995; 32:
557-60. 20.
Craddock JG, Kerby GP, Durham NC. Urinary excretion of acid
mucopolysaccharides by diabetic patients. J Lab and Clinic Med
1955; 46: 193-8. 21. Lubec G, Legenstein E, Pollak A, Meznik E. Glomerular basement membrane charges, Hb A1c and urinary excretion of acid glycosaminoglycans in children with diabetes mellitus. Clinica Chimica Acta 1980; 103: 45-9. 22.
Bonavita N, Reed P, Donnelly PV, Hill LL, Differante N. The
urinary excretion of heparan sulfate by juvenile and adult onset
diabetic patients. Connective Tissue Research 1984; 13: 83-7. 23. Baggio
B, Briani G, Cicerello E, Gambaro G, Bruttomesso D, Tiengo A, et al.
Urinary
glycosaminoglycans, sialic acid and lysosomal enzymes increase in
nonalbuminuric diabetic patients. Nephron 1986; 43: 187-90. 24.
Perez Blanco F, Moreno Terribas G, Cantero Hinojosa J,
Rodriguez Cuartero A. Urinary excretion of glycosa-minoglycans in
patients with early diabetic nephropathy (Letter). Nephron
1996; 73: 344-5. 25.
Report of the Expert Committe on the diagnosis and
classification of Diabetes Mellitus. The Expert Committee on the
diagnosis and classification of Diabetes Mellitus. Diabetes Care
1998; 21 (Suppl 1) S5-S19. 26.
Lagunoff D, Warren G. Determination of
2-deoxy-2-sulfoaminohexose content of mucopolysaccharides. Arch
Bioch & Biophys 1962; 99: 392-400. 27.
Smith RL, Gilkerson E. Quantitation of glycosaminoglycan
hexosamine using 3-methyl-2-benzothiazolone hydrazone hydrochloride.
Analyt Biochem 1979; 98: 478-80. 28.
Viberti GC, Mogensen CE, Keen H, Jacobsen FK, Jarrett RJ,
Christensen CK. Urinary excretion of albumin in normal man: the
effect of water loading. Scand J Clin Lab Invest 1982; 42:
147-57. 29.
Stroten A. A micromethod for creatinine using resin to remove
interfering substances. J Clin Lab Technol 1968; 25: 240-4. 30.
Kofoed-Enevoldsen A, Borch Johnsen K, Kreiner S, Nerup J,
Deckert T. Declining incidence of proteinuria in type I diabetic
patients in Denmark. Diabetes 1987; 2: 205-9. 31.
Mogensen CE, Keane WF, Bennett PM, Jerums G, Parving HH,
Passa P, et al. Prevention of diabetic renal disease with special
reference to microalbuminuria. Lancet 1995; 346: 1080-4. 32.
Mathiesen ER, Oxenboll B, Johanden K, Svenchsen PA, Deckert
T. Incipient nephropathy in Type 1 (insulin-dependent) diabetes.
Diabetologia 1984; 26: 406-10. 33.
Borch Johnsen K, Krag Andersen P, Deckert T. The effect of
proteinuria on relative mortality in IDDM. Diabetologia 1985;
28: 590-6. 34.
Barbosa J. Concordance for nephropathy in diabetic siblings:
Evidence of genetic susceptibility to diabetic kidney disease
(Abstract). Diabetes 1987; 36 (suppl): 105A. 35. Kanwar YS, Veis A, Kumura JM, Jakuboswsky ML. Characterization of heparan sulfate proteoglycan of glomerular basement membranes. Proc Natl Acad Sci USA 1984; 81: 762-6. 36.
Rohrbach DH, Hassell JR, Kleinman H, Martin GR. Alterations
in the basement membrane (heparan sul-
fate) proteoglycan in diabetic mice. Diabetes 1982;
31: 185-8. 37.
Reddi AS, Ranamurth M, Miller S, Dhuper S, Lasker N.
Enalapril improves albuminuria by preventing glomerular loss of
heparan in diabetic rats. Biochem Med Metab Biol 1991; 45:
119-31. 38.
Mitsuhashi H, Tsukada Y, Ono K, Yano S, Naruse T. Urine
glycosaminoglycans and heparan sulfate excretions in adult patients
with glomerular disease. Clin Nephrol 1993; 39: 231-7. 39.
Tarsio JF, Wigness B, Rhade TD, Rupp WM, Buchwald H, Furcht
LT. Non enzymatic glycation of fibronectin and alteration in
molecular association of cell matrix and basement membrane
components in diabetes mellitus. Diabetes 1985; 34: 477-84. 40.
Vernier RL, Steffes MW, Sisson Ross S, Mauer SM. Heparan
proteoglycan in glomerular basement membrane in type I diabetes
mellitus. Kidney Int 1992; 41: 1070-80. 41.
Mathiesen ER, Ronn B, Yensen T, Storm B, Deckert T.
Microalbuminuria precedes elevation in blood pressure in diabetic
nephropathy (Abstract). Diabetologia 1988; 31: 519A. 42.
Velussi M, Cemigoi AM, Dapas F, De Monte A.
Glyco-saminoglycans oral therapy reduces microalbuminuria, blood
fibrinogen levels and limb arteriopathy clinical signs in patients
with non-insulin dependent diabetes mellitus. Diab Nutr Metab
1996; 9: 53-8. 43.
Kofoed-Enevoldsen A. Heparan sulphate in pathogenesis of
diabetic nephropathy. Diabetes Metab Reviews 1995; 11:
137-60. Received: 18-III-1999 Accepted:
4-XI-1999 Postal
address: Bioq. Ana María
Paglione, Corrientes 3985, 1194 Buenos Aires, Argentina Fax:
(54-11) 4508-3989
e-mail: ampaglione@dbc.ffyb.uba.ar TABLE
1.– Clinical features and laboratory findings in control
subjects and diabetic patients
Feature
Controls Diabetic patients
(n=24)
(n=49) Sex
ratio (F/M) 12/12
27/22 Age
in yearsa
40.61 ± 17.09
53.44 ± 19.02
Range in years
18-73 17-83 BMI
kg/m2
b
25.2 ± 3.1
27.1 ± 4.6 Albuminuria
mg/dayc
5.54 ± 2.87
12.0 ± 11.8 HbA1c%d
6.5 ± 2.8
7.3 ± 2.3 Blood
creatinine mg/dle
0.87 ± 0.21
0.96 ± 0.19 Creatinine
clearance %f
92.12 ± 27.0
82.97 ± 30.24 Laboratory
data are expressed in mean values ± SD and SEM. a
p < 0.008; b NS; c p
< 0.001; d
NS; e
p < 0.025; f NS Fig.
1.– a. Heparan sulphate distribution in 24 control subjects. Fig.
1.– b. Heparan sulphate distribution in 49 diabetic patients. Fig.
2.– a. Urinary excretion of heparan sulphate (HS) in female (F)
and in male control subjects (M). Data are expressed as mean values
± SD and SEM in mg/24 hours of urinary HS, with p = 0.02. Fig.
2.– b. Urinary excretion of heparan sulphate (HS) in diabetic
females (F) and in males (M). Data are expressed as mean values ±
SD and SEM in mg/24 hours of urinary HS, with p < 0.02. Fig.
3.– Urinary excretion of heparan sulphate (HS) in healthy controls
(C) and in normoalbuminuric diabetic patients (D). Data are
expressed as mean values ± SD and SEM in mg/24 hours of urinary HS,
with p = 0.001. -
- - - [...]
Most people really waste the time of enforced idleness which is
such an inevitable accompaniment of war. It is greatly to the credit
of the contributors here that they have fought mental lethargy to
the point of creating this valuable composite statements upon the
cell. The spirit is indeed hard to crush: the weakness and strength
of the flesh rest upon organized chemistry. [...] La mayoría de las personas realmente desperdicia el tiempo de forzada inactividad que es inevitable acompañamiento de la guerra. Es grande el mérito de los que contribuyeron aquí que han combatido el letargo mental hasta el punto de crear esta valiosa combinación de exposiciones sobre la célula. El espíritu es verdaderamente duro de aplastar; la debilidad y la fuerza de la carne se apoyan sobre la química organizada. R.
A Peters Foreword
(Prefacio) de Cytology and Cell Physiology, edited by
Geoffrey Bourne. Oxford:
OU Press, 1942
|
|||||||