Potential Value Of Glycopatterns In Urinary Protein On The Differential Diagnosis Of Diabetic Nephropathy And Non-diabetic Renal Disease

Background:Diabetic nephropathy(DN)is the most common and serious microvascular complication of diabetes.To date,the gold standard for identifying DN and non-diabetic renal disease(NDRD)is a renal biopsy;however,there is currently no reliable diagnostic marker to identify DN and NDRD in a noninvasive manner.This study aimed to investigate the different glycopatterns in urine specimens of DN patients and NDRD patients for a differential diagnosis.Methods:In total,19 DN patients and 18 NDRD patients who underwent renal biopsies between March 2015 and March 2016 at the Chinese People's Liberation Army General Hospital were enrolled in this study.A lectin microarray was used to investigate the glycopatterns in the urinary protein of the 37 patients.Ratio analysis and one-way analysis of variance(ANOVA)were used to screen altered glycopatterns.Then,the altered glycopatterns between the DN and NDRD groups were verified by a urinary protein microarray among another 32 patients(15 with DN,17 with NDRD),and receiver operating characteristic(ROC)curve analysis was used to determine the diagnostic value of the altered glycopatterns in differentiating DN and NDRD.Finally,lectin blotting was used to evaluate the altered glycosylation in protein level.Results:The result of lectin microarrays revealed that the glycan profile exhibited difference between DN and NDRD groups.The relative abundance of the(?-1,4)-linked N-acetyl-D-glucosamine(G1cNAc)recognized by lectin Datura stramonium agglutinin(DSA)was significantly higher in urinary protein in DN patients than that in NDRD patients(fold change>1.50,p<0.001).Meanwhile,in the MN patients,the relative abundance of glycopatterns of ?-Gal identified by lectin RCA120 was higher than others.In IgAN patients,the relative abundance of the terminal GlcNAc recognized by STL was higher than others.In order to verify the finding of lectin bllotting,urinary protein microarray was performed to evaluate the difference between DN and NDRD patient.Subsequently,the results of urinary protein microarrays revealed that the relative abundance of the(?-1,4)-linked GlcNAc was significantly different between the DN and NDRD groups,and it was consistent with lectin microarrays(p<0.05).Furthermore,the ROC curve showed that glycopatterns recognized by lectin DSA could effectively distinguish DN from NDRD patients(area under the ROC curve[AUC]=0.94,p<0.001).DSA SDS-PAGE and lectin blotting showed that the glycoproteins,with a molecular weight of approximately 50,000,demonstrate a difference in urine samples between DN patients and NDRD patients,and the molecular weight of(?-1,4)-linked GlcNAc is 50000,the result further reveled that the relative abundance of the(?-1,4)-linked GlcNAc was higher in DN groups than in NDRD groups.Conclusions:The relative abundance of(?-1,4)-linked GlcNAc recognized by lectin DSA and urinary glycoprotein with a molecular weight of approximately 50,000 are significantly different between DN and NDRD patients,indicating that the glycopatterns could be used as potential biomarkers for a differential diagnosis.