Levels of Urinary Trypsin Inhibitor and Structure of Its Chondroitin Sulphate Moiety in Type 1 and Type 2 Diabetes.

Antonio Junior Lepedda,Nadia Ucciferri,Pierina De Muro,Marilena Formato,Gabriele Nieddu,Silvia Rocchiccioli,Michela Idini

doi:10.1155/2018/9378515

Abstract

Background Diabetes mellitus is a global health problem representing the fifth leading cause of mortality and a major risk factor for cardiovascular diseases. In the last years, we reported an association among urinary trypsin inhibitor (UTI), a small proteoglycan that plays pleiotropic roles in many inflammatory processes, and both type 1 and 2 diabetes and developed a method for its direct quantitation and structural characterization. Methods Urine from 39 patients affected by type 1 diabetes, 32 patients with type 2 diabetes, and 52 controls were analysed. UTI was separated from the main glycosaminoglycans physiologically present in urine by anion exchange chromatography, treated for chondroitin sulphate (CS) chain complete depolymerisation, and analysed for both UTI content and CS structure. UTI identification was performed by nano-LC-MS/MS analysis. Results We evidenced increased UTI levels, as well as reduced sulphation of its CS moiety in association with diabetes, regardless of both age and medium-term glycaemic control. Furthermore, no association between UTI and albumin excretion rate was found. Conclusions Evidences suggest that UTI levels are not directly correlated with renal function or, otherwise, that they may increase before the onset of renal impairment in diabetes, representing a potential marker for the underlying inflammatory condition.

Highlights

Urinary trypsin inhibitor (UTI) is a small proteoglycan (PG), with inhibitory activity against serine proteases, resulting from excretion of plasma bikunin into urine [1]
As the purpose of this work was to analyse at a structural level urinary trypsin inhibitor (UTI)-chondroitin sulphate (CS) moiety, we performed two elution steps that allowed the separation of UTI fraction from the highly charged GAG fraction, containing HS and a normosulphated CS with different origin from UTI-CS
A half of each sample was subjected to SDS-PAGE analysis for UTI quantitation and identification by nano-LCMS/MS analysis (Table 1, see Supplementary Material 1 for complete MS information), while the other one, following AMAC derivatization, was analysed by means of fluorophoreassisted carbohydrate electrophoresis (FACE) for CS content and structure

Summary

Introduction

Urinary trypsin inhibitor (UTI) is a small proteoglycan (PG), with inhibitory activity against serine proteases, resulting from excretion of plasma bikunin into urine [1]. Despite that UTI was purified in 1955 for the first time [2] and several studies on its structure and levels have been performed since the 1950s, its biological function has not been fully understood yet [3] It is composed of a protein moiety, a lowsulphated chondroitin sulphate (CS) chain, O-linked to the serine 10, as well as an oligosaccharide, N-linked to the asparagine 45 of the protein moiety (Figure 1). We reported an association among urinary trypsin inhibitor (UTI), a small proteoglycan that plays pleiotropic roles in many inflammatory processes, and both type 1 and 2 diabetes and developed a method for its direct quantitation and structural characterization. Evidences suggest that UTI levels are not directly correlated with renal function or, otherwise, that they may increase before the onset of renal impairment in diabetes, representing a potential marker for the underlying inflammatory condition

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Conclusion