Differential Proteome Profiling Using iTRAQ in Microalbuminuric and Normoalbuminuric Type 2 Diabetic Patients

Abstract

Diabetic nephropathy (DN) is a long-term complication of diabetes mellitus that leads to end-stage renal disease. Microalbuminuria is used for the early detection of diabetic renal damage, but such levels do not reflect the state of incipient DN precisely in type 2 diabetic patients because microalbuminuria develops in other diseases, necessitating more accurate biomarkers that detect incipient DN. Isobaric tags for relative and absolute quantification (iTRAQ) were used to identify urinary proteins that were differentially excreted in normoalbuminuric and microalbuminuric patients with type 2 diabetes where 710 and 196 proteins were identified and quantified, respectively. Some candidates were confirmed by 2-DE analysis, or validated by Western blot and multiple reaction monitoring (MRM). Specifically, some differentially expressed proteins were verified by MRM in urine from normoalbuminuric and microalbuminuric patients with type 2 diabetes, wherein alpha-1-antitrypsin, alpha-1-acid glycoprotein 1, and prostate stem cell antigen had excellent AUC values (0.849, 0.873, and 0.825, resp.). Moreover, we performed a multiplex assay using these biomarker candidates, resulting in a merged AUC value of 0.921. Although the differentially expressed proteins in this iTRAQ study require further validation in larger and categorized sample groups, they constitute baseline data on preliminary biomarker candidates that can be used to discover novel biomarkers for incipient DN.

Prediction of development and maintenance of high albuminuria during chronic renin–angiotensin suppression by plasma proteomics

“…Interestingly, elevation of CP and ORM1, which predicts development of high albuminuria (Fig. 6B), is detected earlier in plasma than in urine in view of the fact that a recent study homologous carried out by Gonzalez-Calero et al found elevated urine CP in individuals with de novo high albuminuria (data submitted), as well as a previous study that found higher urine levels of ORM1 in diabetic patients with high albuminuria [30]. Our results suggest that routine analysis of the plasma levels of the hereby reported proteins by turdibimetry could be useful to evaluate renal function in hypertensive patients with no clinical signs of renal impairment.…”

A pharmacoproteomic study confirms the synergistic effect of chondroitin sulfate and glucosamine

“…Data were normalized for loading error by bias and the background correction was calculated using the Pro Group™ algorithm (ABSciex). To determine the cutoff value for significant fold changes, coefficient of variation and cumulative frequency were calculated for the common proteins employing the R statistical package58. Using this tool, we considered statistically significant only those changes with a p value ≤ 0.05 and a ratio ≥ 1.4 (or ≤0.7).…”

Quantitative mass spectrometry of urinary biomarkers

“…The use of this powerful technique is gradually becoming the method of choice in the field of biomarker discovery [3, 76–78]. This method allowed discovering P- and E-cadherins as urinary biomarkers of idiopathic nephrotic syndrome [76].…”