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. 2021 Oct 22;26(21):6399.
doi: 10.3390/molecules26216399.

N-Glycosylation Profiling of Human Blood in Type 2 Diabetes by Capillary Electrophoresis: A Preliminary Study

Rebeka Torok  1 , Klaudia Horompoly  1 , Marton Szigeti  1 , Andras Guttman  1   2 , Marta Vitai  3 , Laszlo Koranyi  3 , Gabor Jarvas  1
Affiliations

N-Glycosylation Profiling of Human Blood in Type 2 Diabetes by Capillary Electrophoresis: A Preliminary Study

Rebeka Torok et al. Molecules. .

Abstract

Currently, diagnosing type 2 diabetes (T2D) is a great challenge. Thus, there is a need to find rapid, simple, and reliable analytical methods that can detect the disease at an early stage. The aim of this work was to shed light on the importance of sample collection options, sample preparation conditions, and the applied capillary electrophoresis bioanalytical technique, for a high-resolution determination of the N-glycan profile in human blood samples of patients with type 2 diabetes (T2D). To achieve the profile information of these complex oligosaccharides, linked by asparagine to hIgG in the blood, the glycoproteins of the samples needed to be cleaved, labelled, and purified with sufficient yield and selectivity. The resulting samples were analyzed by capillary electrophoresis, with laser-induced fluorescence detection. After separation parameter optimization, the capillary electrophoresis technique was implemented for efficient N-glycan profiling of whole blood samples from the diabetic patients. Our results revealed that there were subtle differences between the N-glycan profiles of the diabetic and control samples; in particular, two N-glycan structures were identified as potential glycobiomarkers that could reveal significant changes between the untreated/treated type 2 diabetic and control samples. By analyzing the resulting oligosaccharide profiles, clinically relevant information was obtained, revealing the differences between the untreated and HMG-CoA reductase-inhibitor-treated diabetic patients on changes in the N-glycan profile in the blood. In addition, the information from specific IgG N-glycosylation profiles in T2D could shed light on underlying inflammatory pathophysiological processes and lead to drug targets.

Keywords: N-glycan; biomarker; blood collecting tubes; capillary electrophoresis; type 2 diabetes.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
The effect of different types of blood-collection tubes on the resulting N-glycan profiles of whole blood (healthy) after two months with 5 intermittent re-freezing cycles. Separation conditions: 30 cm total (20 cm effective) length, 50 µm i.d. bare fused silica capillary at 20 °C, HR-NCHO separation gel buffer, two-stage injection: 5.0 psi for 5 s water plug → 6.0 kV for 3 s sample.
Figure 2
Figure 2
Comparison of the N-glycome of healthy and type 2 diabetic serum and blood samples. Panel (A)—healthy pooled vs. T2 diabetic serum sample, Panel (B)—healthy blood vs. healthy serum sample, (C)—healthy pooled vs. T2 diabetic blood sample with the indication of potential biomarker peaks by the arrows. Separation conditions were the same as in Figure 1. Structures corresponding to peaks: 1: FA4BG4S4, 2: A2G2S2, 3: A2BG2S2, M3, 4: FA2G2S2, 5: FA2BG2S2, 6: FA2G1S1, 7: A2G2S1, 8: A2BG2S1, 9: FA2G2S1, 10: FA2BG2S1, 11: A4G4S2, 12: FA2, M6, 13: FA2[6]G1, M7, 14: FA2[3]G1, FA2BG1, M8, 15: FA2G2.
Figure 3
Figure 3
Comparison of the N-glycosylation of healthy (n = 10), treated T2D (n = 8) and untreated T2D (n = 1) blood samples. Separation conditions were the same as in Figure 1.
See this image and copyright information in PMC

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