BACKGROUND: Additional urinary biomarkers for diabetic nephropathy (DN) are needed, providing early and reliable diagnosis and new insights into its mechanisms. Rigorous selection criteria and homogeneous study population may improve reproducibility of the proteomic approach. METHODS: Long-term type 1 diabetes patients without metabolic comorbidities were included, 11 with sustained microalbuminuria (MA) and 14 without MA (nMA). Morning urine proteins were precipitated and resolved by 2D electrophoresis. Principal component analysis (PCA) and Projection to latent structures discriminatory analysis (PLS-DA) were adopted to assess general data validity, to pick protein fractions for identification with mass spectrometry (MS), and to test predictive value of the resulting model. RESULTS: Proteins (n = 113) detected in more than 90% patients were considered representative. Unsupervised PCA showed excellent natural data clustering without outliers. Protein spots reaching Variable Importance in Projection score above 1 in PLS (n = 42) were subjected to MS, yielding 33 positive identifications. The PLS model rebuilt with these proteins achieved accurate classification of all patients (R2X = 0.553, R2Y = 0.953, Q2 = 0.947). Thus, multiple earlier recognized biomarkers of DN were confirmed and several putative new biomarkers suggested. Among them, the highest significance was met in kininogen-1. Its activation products detected in nMA patients exceeded by an order of magnitude the amount found in MA patients. CONCLUSIONS: Reducing metabolic complexity of the diseased and control groups by meticulous patients' selection allows to focus the biomarker search in DN. Suggested new biomarkers, particularly kininogen fragments, exhibit the highest degree of correlation with MA and substantiate validation in larger and more varied cohorts.

• Keywords
• Diabetes, Kallikrein-kinin system, Nephropathy, Proteomics,
• MeSH
• Albuminuria diagnosis   urine   MeSH
• Biomarkers urine   MeSH
• Early Diagnosis MeSH
• Diabetic Nephropathies diagnosis   urine   MeSH
• Adult MeSH
• Kallikrein-Kinin System * MeSH
• Humans MeSH
• Nephrons metabolism   MeSH
• Proteins metabolism   MeSH
• Reproducibility of Results MeSH
• Sensitivity and Specificity MeSH
• Check Tag
• Adult MeSH
• Humans MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Observational Study MeSH
• Names of Substances
• Biomarkers MeSH
• Proteins MeSH