Skip to content

Advertisement

You're viewing the new version of our site. Please leave us feedback.

Learn more

BMC Ophthalmology

Open Access
Open Peer Review

This article has Open Peer Review reports available.

How does Open Peer Review work?

Erytrocyte membrane anionic charge in type 2 diabetic patients with retinopathy

  • Yasemin Budak1Email author,
  • Hakan Demirci2,
  • Muberra Akdogan3 and
  • Dilek Yavuz4
BMC Ophthalmology20044:14

https://doi.org/10.1186/1471-2415-4-14

Received: 26 May 2004

Accepted: 08 October 2004

Published: 08 October 2004

Abstract

Background

The Steno hypothesis states that changes in basement membrane anionic charge leads to diabetic microvascular complications. In diabetic nephropathy, loss of basement membrane glycosaminoglycans and the association between glomerular basement membrane heparan sulphate and proteinuria has been documented. A correlation between erythrocyte surface and the glomerular capillary wall charges has also been observed.

The aim of this study is to evaluate the relationship between retinopathy and erythrocyte anionic charge and urinary glycosaminoglycan excretion in type 2 diabetic patients.

Methods

49 subjects (58 ± 7 yrs, M/F 27/22) with type 2 diabetes with proliferative retinopathy (n = 13), nonproliferative retinopathy (n = 13) and without retinopathy (n = 23) were included in the study. 38 healthy subjects were selected as control group (57 ± 5 yrs, M/F 19/19). Erythrocyte anionic charge (EAC) was determined by the binding of the cationic dye, alcian blue. Urinary glycosaminoglycan and microalbumin excretion were measured.

Results

EAC was significantly decreased in diabetic patients with retinopathy (255 ± 30 ng alcian blue/106 RBC, 312 ± 30 ng alcian blue/106 RBC for diabetic and control groups respectively, p < 0.001). We did not observe an association between urinary GAG and microalbumin excretion and diabetic retinopathy. EAC is found to be negatively corralated with microalbuminuria in all groups.

Conclusions

We conclude that type 2 diabetic patients with low erythrocyte anionic charge are associated with diabetic retinopathy. Reduction of negative charge of basement membranes may indicate general changes in microvasculature rather than retinopathy. More prospective and large studies needs to clarify the role of glycosaminoglycans on progression of retinopathy in type 2 diabetic patients.

Background

Diabetic retinopathy is the leading cause of blindness in diabetic adults [1, 2]. During the first two decades of the disease, nearly all patients with type 1 diabetes and > 60 % of type 2 diabetes have retinopathy. The duration of the diabetes is probably the strongest predictor for development and progression of retinopathy [3]. The incidence of retinopathy positively correlates with HbA1c [4].

Diabetic patients with evidence of nephropathy are characterized by a 5 to 10 times higher incidence of proliferative retinopathy [5]. Albuminuria not only is associated with kidney disease but, is also a strong predictor of cardiovascular disease and proliferative retinopathy, suggesting it reflects a generalized vascular disease [6]. Thus, the coincidence of generalized vascular dysfunction, albuminuria, mesangial expansion, proliferative retinopathy and accelerated development of atherosclerosis suggests a common cause of abnormalities in susceptible diabetic patients.

The Steno hypothesis proposed that increased loss of anionic charge from basement membranes leads to diabetic microangiopathy [7]. This hypothesis has been proven for diabetic nephropathy but it's not clear for diabetic retinopathy.

Anionic content of basement membranes predominantly produced by glycosaminoglycan molecules (GAG). They constitute the charge selective barrier of the glomerul basement membrane [8]. Several clinical studies indicate that loss of the GAG is associated with diabetic albuminuria in diabetic and experimental models [9, 10].

Although abnormal GAG metabolism is a well known phenomen in diabetic nephropathy, it is still debated whether it has a pathogenic role in diabetic retinopathy. The aim of this study was to evaluate the relationship between diabetic retinopathy and erythrocyte anionic charge as well as the urinary glycosaminoglycan excretion in type 2 diabetic patients.

Methods

49 outpatients (27 male, 22 female) with type 2 diabetes mellitus diagnosed after the age of 30, were included and divided into 3 subgroups according to severety of retinopathy. Diabetes was diagnosed according to American Diabetes Association criteria. The onset of diabetes was after the age of 30 in these patients.

The study was approved by the institutional ethics committee and patients gave written informed consent. Exclusion criteria included hypertension, myocardial infarction, cerebrovascular disease, heart failure, treatment with antiaggregants, steroids or other drugs that effect blood pressure and glucose, serum creatinine > 200 μmol/l, connective tissue disorders and other systemic disease.

Renal and bladder infection diseases of patients and controls were excluded biochemically and microbiologically.

The ocular fundus was examined by an ophthalmologist after dilation of the pupils and classified. The patients were divided into three groups according to the severity of retinopathy. Group 1 consisted of 23 patients without retinopathy (R0). Group 2 consisted of 13 patients with nonproliferative retinopathy (R1-hemorrhages, microaneurysms, cotton-wool exudates). The 13 patients in group 3 (R2) had proliferative retinopathy (R2-neovascularisations, vitreous hemorrhages, ablatio of the retina).

Age and sex matched 38 healthy volunteers were included as healthy controls. Demographic characteristics of the groups are shown in Table 1.
Table 1

Demographic characteristics of the study group

 

Control group

Group R0

GroupR1

Group R2

Age(yrs)

57 ± 5

59 ± 8

56 ± 5

57 ± 5

M/F

19/19

13/10

7/6

7/6

BMI(kg/m2 )

28 ± 5

27.8 ± 6

29.6 ± 5

28.5 ± 8

OAD/insulin

-

22/1

12/1

12/1

Duration of diabetes (yrs)

-

10 ± 5

11 ± 6

12 ± 8

Venous samples were collected after an overnight fast and 24 hour urine samples collected.

The charge on erythrocytes (RBC) was measured by means of cationic dye alcian blue 8GX (Sigma catalogue no: A 5268) according to the method of Levin with minor modifications as follows: [11]. From citrated venous blood samples, platelets and leukocytes were removed using the method of Beutler et al. [12]. After removal of platelets and leucocytes erythrocytes were washed 3 times in phosphate buffered saline (PBS), and subsequently a fraction of RBC was resuspended in PBS containing alcian blue at a final concentration of 250 mg/l. After a 30 min incubation at 37°C, the RBC suspension was centrifuged, and the remaining alcian blue concentration was measured in the supernatant with a Shimadzu UV 1201 V spectrophotometer (Shimadzu, Japan) at a wavelength of 650 nm. Each determination represents the mean of 2 assays. The quantity of alcian blue bound to RBCs was expressed as nanograms of alcian blue per 106 RBCs. In our experimental conditions the intraassay and interassay coefficients of variation were 5.8 and 7.6 % for 100 nanograms of alcian blue per 106 RBCs.

Urinary glycosaminoglycan (UGAG) excretion was determined in 24 hr urine samples spectrophotometrically at 520 nm by the addition of dimethylmethylene blue (Aldrich Chem Co., USA) and bovine kidney heparan sulfate as standard (Sigma catalogue no H 7640) [13]. In our experimental conditions the intraassay and interassay coefficients of variation were 1.5 and 2.4 % for 10 mg/l glycosaminoglycan, respectively.

Serum sialic acid was measured in serum with a colorimetric method described by Svennerholm with a Shimadzu UV-1201 spectrophotometer (Shimadzu, Japan) with a wavelength of 525 nm. Using N-acetyl neuraminic acid (Sigma catalogue no: A 3007) as standard [14]. Intra assay and interassay coefficients of variations were 6.6 and 9.2 % for 1 mmol/l sialic acid respectively.

Urinary albumin excretion was measured by an immunoturbidemetric assay (Roche diagnostics, USA) on automated clinical chemistry analyzer (Hitachi 902). In all patients, the annual level was determined as the mean of urinary albumin excretions in three 24 h urine collections taken at home during normal physical activity. The intraassay and interassay coefficients of variation were 1.3 and 4.3 % for a mean value of 25 mg/l albumin concentration.

Serum glucose, cholesterol, triglyceride and HDL (Dade Behring, USA) were evaluated with enzymatic methods (Dade Behring, USA) on an automated clinical chemistry analyzer (Dimension RxL). Creatinin was assayed by the Jaffe reaction on an automated clinical chemistry analyzer (Dimension RxL).

HbA1C (reference range 4.4–6.0 %) was measured by a turbidimetric inhibition immunoassay technique (Roche Diagnostics, USA) on automated clinical chemistry analyzer (Hitachi 902). The intraassay and interassay coefficients of variation were 1.8 and 3.0 % for a mean value of 10.5 % HbA1C concentration.

The analysis of the data was performed with a PC compatible Instat-II programme. Paired t test and ANOVA were used where appropriate. Correlation analysis was performed with Spearman rank test. The differences were considered significant when the probability was p < 0.05. The results were given as mean ± SD.

Results

The clinical characteristics of the study groups are shown in Table 1. There were no difference in age and BMI between the groups. Duration of diabetes mellitus was slightly longer, although not significantly different than that of the patients with proliferative retinopathy.

Results of laboratory parameters are shown in Table 2. Patients with proliferative retinopathy had higher concentrations of triglyceride and lower concentrations of HDL cholesterol as compared to diabetics without evidence of retinopathy and control group. Serum cholesterol level was the same in all study groups. Blood glucose and HbA1C levels were not different in subgroups of our diabetic population.
Table 2

Laboratory parameters of the study groups

 

Control group

Group R0

GroupR1

Group R2

Fasting glucose (mg/dl)

100 ± 26

229 ± 70

234 ± 89

236 ± 120

HbA1c (%)

 

7.6 ± 0.7

8.0 ± 0.7

8.0 ± 0.8

Cholesterol (mg/dl)

192 ± 44

187 ± 33

197 ± 41

193 ± 40

Triglycerides (mg/dl)

131 ± 55

153 ± 84

199 ± 71

228 ± 143

HDL (mg/dl)

45 ± 9

39 ± 9

37 ± 7

36 ± 10

Serum sialic acid (mmol/l)

1.81 ± 0.3

2.0 ± 0.5

2.1 ± 0.3

2.3 ± 0.5

EAC (ng alcian blue 106 erythrocyte)

312 ± 30

281 ± 30

259 ± 23

252 ± 37

UGAG (mg/24 h)

5.0 ± 2.4

8.3 ± 4.1

8.7 ± 4.8

8.4 ± 3.3

Microalbumin (mg/24 h)

10 ± 6

73 ± 40

75 ± 21

85 ± 45

Distribution of erythrocyte anionic charges of all study groups are demonstrated in figure 1. EAC was 312 ± 30 ng alcian blue/106 RBC in healthy controls. Diabetic patients both with nonproliferative and proliferative retinopathy had a significantly lower alcian blue binding to RBCs compared with diabetic patients without retinopathy (p < 0.001).
Figure 1

Distribution of erythrocyte anionic charges of all study groups.

A marked (p < 0.05) difference was found between type 2 diabetic patients (8.3 ± 4.1, 8.7 ± 4.8, 8.4 ± 3.3 mg/24 h, Table 2) and controls (5.0 ± 2.4 mg/24 h) regarding urinary GAG excretion. We did not observe a correlation between urinary GAG excretion and diabetic retinopathy.

A significant difference (p < 0.05) was observed in serum sialic acid values in diabetic patients compared with healthy controls. Serum sialic acid level was higher in the nonproliferative retinopathy group, but the difference was statistically insignificant (p > 0.05).

Microalbumin level was significantly p < 0.05 higher in diabetic patients (73 ± 40 [35–250], 75 ± 21 [50–202], 85 ± 45 [56–224] mg/24 h, Table 2) compared to healthy controls (10 ± 6 mg/24 h). But there wasn't a correlation between the severity of retinopathy and microalbuminuria (p > 0.05).

Results of the Spearman rank correlation test are shown on table 3. EAC is found to be negatively corralated with microalbuminuria in all groups.
Table 3

Correlation analysis between erythrocyte anionic charge (EAC) and other parameters.

 

EAC (ng AB/ 106 erythrocyte)

 
 

r

P

HbA1c (%)

-0.251

0.073

Microalbumin (mg/24 h)

-0.393

0.032*

UGAG (mg/24 h)

-0.226

0.094

Serum sialic acid (mmol/L)

0.095

0.546

Duration of diabetes (yrs)

0.097

0.478

* Statistically significant at 0.05 level.

Discussion

Our study demonstrates a statistically significant quantitative reduction of alcian blue binding to RBCs in diabetic patients than healthy subjects, which is in accordance with literature [15, 16]. Previously we have shown a statistically significant quantitative reduction of alcian blue binding to RBCs in streptozosin diabetic rats [17]. Alcian blue is a complex amphoteric molecule that binds to acidic glycoproteins which represent a large class of cell surface anionic molecules [18]. As alcian blue binding is an expression of the anionic charge on the cell surface, this result indicates that RBC anionic charge is reduced in diabetic patients.

Erythrocyte anionic charge by itself is unlikely to be important in the pathogenesis of diabetic retinopathy but reduced RBC alcian blue binding was found to be associated with the loss of glomerular basement membrane anionic charges in diabetic rats and patients [19]. Changes in the composition of basement membranes are likely to be responsible for functional disturbances and hence for the development of capillary disease. Chakrabarti et al. reported a decreased density of anionic sites in the retinal basement membrane of BB rats [20]. A similar decrease in anionic density was also demonstrated in the Bruch's membrane of BB and streptozosin diabetic rats [21].

Several authors have questioned the validity of the alcian blue binding assay. The major objections concern the incomplete alcian blue dissolution in PBS buffer and its tendency to precipitate with time [22, 23]. We prepared a fresh alcian blue solution for each experiment. Each experiment was considered valid when no change in the optical density of the blank alcian blue solution occurred over the duration of the experiment. In addition, the experiments were done in batches with cells from a healthy person. Therefore any flaw in the methodology would not affect only one group and thus distort the results.

Diabetic retinopathy is characterized by gradually progressive alterations in the retinal microvasculature leading to vascular hyperpermeability, capillary occlusion and ultimately neovascularization. Thickening of the retinal microvascular basement membrane and loss of anionic content are well documented morphological features of diabetic retinopathy [20, 21, 24]. These changes causes breakdown of the blood-retinal barrier resulting in capillary hyperpermeability and leakage of proteins into the deep and superficial layers of the retina [1, 2].

Glycosaminoglycan primarily heparan sulfate has been implicated in permeability properties of basement membrane of glomerular basement membrane as well as the retinal basement membrane [25]. Kahaly et al. reported that urinary GAG excretion is increased in diabetic patients with microangiopathy [26]. Williamson et al and Ginn et al. have shown that albumin permeation is increased into the diabetic compared to normal retina [27, 28]. Loss of heparan sulfate from the retinal basement membrane in addition to its thickening may cause increased vascular permeation of albumin and other substances into the retina and possibly into the optic nerve.

Systemic treatment of IDDM patients with proteinuria with danaparoid sodium, a glycosaminoglycan, not only reduced proteinuria, but also decreased the number of hard exudates in the retina [29]. Studies in diabetic rats have shown reduced activity of the key enzyme in the biosynthesis of heparan sulfate, N-deacetylase, which results in impaired heparan sulfate biosynthesis in experimental diabetes [30, 31].

In this study we found that urinary GAG excretion is increased in diabetic patients than healthy controls, but there was no difference in GAG excretion between nonproliferative and proliferative retinopathy patients. We couldn't find a correlation between GAG excretion and EAC but we found a correlation between albumin excretion and EAC. Although our working hypothesis is that the reduction in anionic charge in basement membranes may be due to an abnormal GAG metabolism, we couldn't confirm it with this study.

Our study did not directly address the pathophysiology of diabetic retinopathy but a reduction in erythrocyte charge perhaps connected with the properties of retinal basement membrane.

Several reports suggest that elevated serum sialic acid levels in type 1 and type 2 diabetic patients indicate microvascular damage [32, 33]. In our study, diabetic patients were found to have elevated serum sialic acid levels in comparison with healthy controls. Therefore, the increase in serum sialic acid level may indicate a coincidence between the structural alterations in the retina and general vascular damage.

Conclusions

We conclude that type 2 diabetic patients with low erythrocyte anionic charge are associated with diabetic retinopathy. Reduction of negative charge of basement membranes may indicate general changes in microvasculature rather than retinopathy. More prospective and large studies needs to clarify the role of glycosaminoglycans on progression of retinopathy in type 2 diabetic patients.

List of abbrevations

BMI: 

Body mass index

EAC: 

Erythrocyte anionic charge

GAG: 

Glycosaminoglycan

PBS: 

Phosphate buffered saline

RBC: 

Erythrocytes

UGAG

Urinary glycosaminoglycan

Declarations

Authors’ Affiliations

(1)
SSK Sevket Yilmaz Hospital, Section of Clinical Chemistry
(2)
SSK Sevket Yilmaz Hospital, Section of Family Medicine
(3)
SSK Sevket Yilmaz Hospital, Section of Ophthalmology
(4)
Marmara University Medical School, Section of Endocrinology and Metabolism

References

  1. Merimee TJ: Diabetic retinopathy: a synthesis ofperspectives. N Engl J Med. 1990, 322: 978-983.View ArticlePubMedGoogle Scholar
  2. D'Amico DJ: Diseases of the retina. N Engl J Med. 1994, 331: 95-106. 10.1056/NEJM199407143310207.View ArticlePubMedGoogle Scholar
  3. Fong DS, Aiello L, Gardner TW, King GL, Blankenship G, Cavallerano JD, Ferris FL, American Diabetes Association, Klein R: Diabetic retinopathy. Diabetes Care . 2003, 26 (Suppl 1): S99-S102. 3View ArticlePubMedGoogle Scholar
  4. Chaturvedi N, Sjoelie AK, Porta M, Aldington SJ, Fuller JH, Songini M, Kohner EM, EURODIAB Prospective Complications Study : Markers of insulin resistance are strong risk factors for retinopathy incidence in type 1 diabetes. Diabetes Care. 2001, 24: 284-289.View ArticlePubMedGoogle Scholar
  5. Kofoed-Enevoldsen A, Jensen T, Borch-Johnson K, Deckert T: Incidence of retinopathy in type 1 (insulin dependent) diabetes: association with clinical nephropathy. J Diabet Comp. 1987, 1: 96-99.Google Scholar
  6. Jensen T: Albuminuria: a marker of renal and generalized vascular disease in insulin dependent diabetes mellitus. Dan Med Bull. 1991, 38: 134-144.PubMedGoogle Scholar
  7. Deckert T, Feldt-Rasmussen B, Burch-Johnsen K, Jensen T, Kofoed-Enevoldsen A: Albuminuria reflects widespread vascular damage. The Steno hypothesis. Diabetologia. 1989, 32: 219-226. 10.1007/BF00285287.PubMedGoogle Scholar
  8. Karwar YS, Linker A, Farquhar MG: Increased permeability of the glomerular basement membrane to ferritin after removal of glycosaminoglycan (heparan sulfate) by enzyme digestion. J Cell Biol. 1980, 86: 688-693. 10.1083/jcb.86.2.688.View ArticleGoogle Scholar
  9. Van den Born J, Van Kraats AA, Bakker MAH, Assmann KJM, Van den Heuvel LPWJ, Berden JHM: Selective proteinuria in diabetic nephropathy in the rat is associated with a relative decrease in glomerular basement membrane heparan sulfate. Diabetologia. 1995, 38: 161-172. 10.1007/s001250050266.View ArticlePubMedGoogle Scholar
  10. Gambaro G, Cicerello E, Mastrosimone S, Lavagninit BB: High urinary excretion of glycosaminoglycan:a possible marker of glomerular involvement in diabetes. Metabolism. 1989, 38: 419-420. 10.1016/0026-0495(89)90190-X.View ArticlePubMedGoogle Scholar
  11. Gambaro G, Baggio B, Cicerello E, Mastrosimone S, Marzaro G, Borsatti A, Crepaldi G: Abnormal erythrocyte charge in diabetes mellitus. Link with microalbuminuria. Diabetes. 1988, 37: 745-748.View ArticlePubMedGoogle Scholar
  12. Beutler E, West C, Blume KG: The removal of leucocytes and platelets from whole blood. J Lab Clin Med. 1976, 88: 328-333.PubMedGoogle Scholar
  13. Jong JGN, Wavera RA, Laarakkers C, Poorthhula BJHM: Dimethylene blue-based spectrophotometry of glycosaminoglycans in untreated urine: a rapid screening procedure for mucopolysaccharidosis. Clin Chem. 1989, 35: 1472-1477.PubMedGoogle Scholar
  14. Crook M, Haq M, Tutt P: Evaluation of three assays for the determination of serum sialic acid. Clin Biochem. 1993, 26: 449-454.View ArticlePubMedGoogle Scholar
  15. Bernard A, Amor AO, Goemare-Vanneste J, Antoine JL, Lauweys I, Vandeleene B, Lambert A: Urinary proteins and red blood cell membrane negative charges in diabetes mellitus. Clin Chim Acta. 1990, 190: 249-262. 10.1016/0009-8981(90)90178-U.View ArticlePubMedGoogle Scholar
  16. Estivi P, Cavallo-Perin P, Pagano G: Electrical anionic charges on red blood cells are reduced in insulin dependent diabetic patients. J Diabet Complications. 1989, 3: 45-48. 10.1016/0891-6632(89)90010-X.View ArticlePubMedGoogle Scholar
  17. Yavuz D, Ersöz Ö, Küçükkaya B, Budak Y, Ahiskali R, Ekicioğlu G, Emerk K, Akalın S: The effect of losartan and captopril on glomerular basement membrane anionic charge in a diabetic rat model. J Hyper. 1999, 17: 1217-1223. 10.1097/00004872-199917080-00023.View ArticleGoogle Scholar
  18. Spicer SS, Schulte BA: Diversity of cell glycocongugates shown histochemically: a perspective. J Histochem Cytochem. 1992, 40: 1-38.View ArticlePubMedGoogle Scholar
  19. Gambaro G, Baggio B, Cicerello E, Mastrosimone S, Marzaro G, Borsatti A, Crepaldi G: Abnormal erythrocyte charge in diabetes mellitus. Link with microalbuminuria. Diabetes. 1988, 37: 745-748.View ArticlePubMedGoogle Scholar
  20. Chakrabarti S, Ma N, Sima AAF: Anionic sites in diabetic basement membranes and their possible role in diffusion barrier abnormalities in the BB-rat. Diabetologia. 1991, 34: 301-306.View ArticlePubMedGoogle Scholar
  21. Caldwell RB, Slapnick SM, McLaughin BJ: Decreased anionic sites in Bruch's membrane of spontaneous and drug-induced diabetes. Invest Ophtalmol Vis Sci. 1986, 27: 1691-1697.Google Scholar
  22. Swell RF, Brendey PEC: Red cell surface charge in glomerular disease. Lancet. 1986, 2: 635-636.Google Scholar
  23. Swell RF, Short CD: Minimal change nephropathy: how does the immune system affect the glomerulus. Nephrol Dialysis Transplant. 1993, 8: 108-112.Google Scholar
  24. Carlson EC, Audette JL, Veitenheimer NJ, Risan JA, Laturnus DI: Ultrastructural morphometry of capillary basement membrane thickness in normal and transgenic diabetic mice. Anat Rec. 2003, 271: 332-341. 10.1002/ar.a.10038.View ArticleGoogle Scholar
  25. Das A, Frank RN, Zhang NL , Turezyn TJ: Ultrastructural localization of extracellular matrix components in human retinal vessels and Bruch's membrane. Arch Ophthalmol. 1990, 108: 421-429.View ArticlePubMedGoogle Scholar
  26. Kahaly G, Hansen C, Otto E, Forster G, Beyer J, Homme IG: Diabetic microangiopathy and urinary glycosaminoglycans. Exp Clin Endocrinol Diabetes. 1997, 105: 145-151.View ArticlePubMedGoogle Scholar
  27. Williamson JR, Chang K, Tilton RG, Prater C, Jeffrey JR, Weigel C, Sherman WR, Eades DM, Kilo C: Increased vascular permeability in spontaneously diabetic BB/W rats with mild versus severe streptozocin-induced diabetes. Prevention by aldose reductase inhibitors and castration. Diabetes. 1987, 36: 813-821.View ArticlePubMedGoogle Scholar
  28. Gin T, Joon TL, Panagiotopoulos S, Cooper M, Taylor H, Jerums G: Organ specificity of antihypertansive therapy on ocular albumin vascular clearence and albuminuria in hypertensive diabetic rat. Invest Ophthalmol Vis Sci. 1996, 37: 281-288.PubMedGoogle Scholar
  29. Van der Pill , van der Woude FJ, Swat W, Van Es LA, Lemkes HH: Effect of danaparoid sodium on hard exudates in diabetic retinopathy. Lancet. 1997, 350: 1743-1745. 10.1016/S0140-6736(97)07126-2.View ArticleGoogle Scholar
  30. Kjellen L, Bielefeld D, Hook M: Reduced sulfatation of liver heparan sulfate in experimentally diabetic rats. Diabetes. 1983, 32: 337-342.View ArticlePubMedGoogle Scholar
  31. Kofoed-Enewoldsen A: Inhibition of glomerular glucosaminyl N- deacetylase in diabetic rats. Kidney Int. 1992, 41: 763-767.View ArticleGoogle Scholar
  32. Crook MA, Pickup JJC, Lumb PJ, Georgino F, Webb DJ, Fuller JH: Relationship between plasma sialic acid concentration and microvascular complications in type 1 diabetes. Diabetes Care. 2001, 24: 316-322.View ArticlePubMedGoogle Scholar
  33. Chen JW, Gall MA, Yokoyama H, Jensen JS, Deckert M, Parving HH: Raised serum sialic acid concentration in NIDDM patients with and without diabetic nephropathy. Diabetes Care. 1996, 19: 130-134.View ArticlePubMedGoogle Scholar
  34. Pre-publication history

    1. The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2415/4/14/prepub

Copyright

© Budak et al; licensee BioMed Central Ltd. 2004

This article is published under license to BioMed Central Ltd. This is an open-access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Advertisement