iTRAQ-Based Quantitative Protein Expression Profiling and MRM Verification of Markers in Type 2 Diabetes

Abstract

The pathogenesis of Type 2 diabetes mellitus (T2DM) is complex owing to molecular heterogeneity in the afflicted population. Current diagnostic methods rely on blood glucose measurements, which are noninformative with respect to progression of the disease to other associated pathologies. Thus, predicting the risk and development of T2DM-related complications, such as cardiovascular disease, remains a major challenge. We have used a combination of quantitative methods for characterization of circulating serum biomarkers of T2DM using a cohort of nondiabetic control subjects (n = 76) and patients diagnosed with T2DM (n = 106). In this case-control study, the samples were randomly divided as training and validation data sets. In the first step, iTRAQ (isobaric tagging for relative and absolute quantification) based protein expression profiling was performed for identification of proteins displaying a significant differential expression in the two study groups. Five of these protein markers were selected for validation using multiple reaction-monitoring mass spectrometry (MRM-MS) and further confirmed with Western blot and QPCR analysis. Functional pathway analysis identified perturbations in lipid and small molecule metabolism as well as pathways that lead to disruption of glucose homeostasis and blood coagulation. These putative biomarkers may be clinically useful for subset stratification of T2DM patients as well as for the development of novel therapeutics targeting the specific pathology.

Metformin, but Not Rosiglitazone, Attenuates the Increasing Plasma Levels of a New Cardiovascular Marker, Fibulin-1, in Patients With Type 2 Diabetes

“…Thus, plasma fibulin-1 appears as a new biomarker for vascular damage, which is in line with the results from four subsequent studies, where plasma fibulin-1 was found to be associated with diabetes, hypertension, and kidney damage (19)(20)(21). Interestingly, two of these studies used non-hypothesisdriven proteomic approaches to search for new markers (20,21). In the current study, we could confirm the previously observed associations between plasma fibulin-1 and age, glycemic status, and pulse pressure, although the correlations were not as strong as previously reported (18).…”

“…In addition, plasma fibulin-1 concentrations independently predicted mortality in a 15-year follow-up period (18). Thus, plasma fibulin-1 appears as a new biomarker for vascular damage, which is in line with the results from four subsequent studies, where plasma fibulin-1 was found to be associated with diabetes, hypertension, and kidney damage (19)(20)(21). Interestingly, two of these studies used non-hypothesisdriven proteomic approaches to search for new markers (20,21).…”

“…Moreover, we demonstrated an association between increased levels of plasma fibulin-1 and arterial stiffness, blood pressure, and glycemic status in a cohort of 305 welldescribed patients with T2D (18). These results, concerning relations between plasma fibulin-1, glycemic status, and hypertension, have been confirmed in three subsequent studies (19)(20)(21). In the initial paper, we also presented a 15-year follow-up study, showing that high plasma fibulin-1 concentrations among patients with T2D were independently predictive of mortality in T2D with a hazard ratio similar to that of plasma cholesterol (18).…”

First-trimester multimarker prediction of gestational diabetes mellitus using targeted mass spectrometry

“…2) [37][38][39]. The best independent predictive apolipoproteins were Apo M, Apo D and Apo L1 (Tables 2 and 3); the performance of Apo M and Apo L1 depended on BMI grouping.…”

Quantitative analysis of the murine lipid droplet-associated proteome during diet-induced hepatic steatosis

“…It is not immediately clear why extracellular proteins would localize intracellularly to LDs or why their localization would be affected by hepatic steatosis; however, many of the identifi ed extracellular proteins have links to hepatic steatosis, obesity, and diabetes (72)(73)(74)(75)(76). Additionally, it is not clear whether these extracellular proteins are in the secretory pathway or are being endocytosed.…”