Chromatographic profiles of blood plasma free oligosaccharides in patients with cardiovascular disease
Authors: Pismenetskaya I. U. , Butters T. D. 
Published: May 2016

Free oligosaccharides (FOS) are unbound structural analogs of glycans of glycoconjugates. There are several sources of them inside the cell: 1) multistep pathways of N-glycosylation, 2) the cell quality control and endoplastic reticulum-associated degradation of mis-glycosylated and/or misfolded glycoproteins, 3) lysosomal degradation of mature glycoconjugates. Some of these FOS are the earliest indicators of potential glycosylation alterations that would be revealed in the course of the cell quality control and the endoplastic reticulum-associated degradation. Ischemia and hypertension cause stress of intracellular organelles leading to disruption of their functions. The main objective of the work was the characterization of free oligosaccharides (FOS) in plasma obtained from patients with cardiovascular diseases compared to those from healthy subjects to evaluate the potential of these compounds for diagnostics. Chromatographic profiles of FOS composed of 4–12 monosaccharides were obtained and analyzed for quantitative and qualitative differences between the samples. After plasma deproteinization and FOS purification the oligosaccharides were labelled with anthranilic acid (2-AA), separated into the neutral and charged with QAE Sephadex (Q25-120) chromatography and analysed using high-performance liquid chromatography (HPLC). Glucose unit values were determined following comparison with a 2-AA-labelled glucose oligomer ladder derived from a partial hydrolysate of dextran as an external standard. The data were collected and processed using Empower software. The charged FOS were digested with the sialidase from Arthrobacter ureafaciens. 2-AA – labelled free oligosaccharides from transferrin were used as an external standard for the structure decoding. The profiles obtained were compared with intracellular free oligosaccharides of known structures and with the glycan structures and their descriptions in the databases GlycoBase and EUROCarbDB. These approaches allowed predicting a range of glycan structures for each of the main peaks of HPLC profiles of plasma free oligosaccharides and managing ways for their future experimental analysis. In the case of cardiovascular disorders, HPLC profiles of FOS revealed a changing pattern of heterogeneity, depending on the severity of the disease. Three main enlarged glycan species in the netral fraction and one peak in the charged fraction distinguished the FOS of the patients from those of the healthy volunteers. It has been revealed that the neutral marker peaks were represented by polimannose glycans with 5–7 mannose residues and 1–2 residues of N-acetylglucosamine, and one of the major peaks of the charged fraction – by two-antennary complex N-glycan with two sialic acid residues. The study of free oligosaccharides of blood plasma is a new field of glycobiology allowing an evaluation of an organism state at the level of the cell organelle functional status and openning up broad prospects for finding early diagnostic and prognostic markers of cardiac insufficiency.