Serum levels of endothelial glycocalyx constituents in women at 20 weeks’ gestation who later develop gestational diabetes mellitus

Abstract

Serum concentration of Heparan Sulfate, Chondroitin Sulfate, Hyaluronan, and Syndecan-1, along with maternal age, BMI, and GDM status of each participant. Study Participants, Definition of GDM, and Matching Criteria: We identified 20 participants without any hypertensive complications from the prospective cohort Screening for Pregnancy Endpoints (SCOPE) study (http://www.scopestudy.net/) in Auckland New Zealand who developed GDM by existing New Zealand criteria defined as a fasting glucose > 5.5mmol/L and/or 2h > 9.0 mmol/L after a 75g Oral Glucose Tolerance Test (OGTT). The GDM subjects were matched with participants who had uncomplicated pregnancies and negative screening for GDM (using the definition of GDM above) and matched for (a) ethnicity; (b) maternal age (age ± 5 years); and, (c) BMI (matched to ± 3 kg/m2). Serum Sample Collection: Venipuncture was performed at 20±1 weeks’ gestation in nonfasting participants. Serum samples were collected into BD plain serum vacutainer tubes, placed on ice and centrifuged at 2400g at 4°C according to a standardized protocol. Serum was stored in 250 μl aliquots at -80°C within 4 hours of collection. Experimental Methods: To assess shedding of the EGL in the circulation in GDM and non-GDM women, we quantified the concentration of the main components of the EGL: syndecan-1, heparan sulfate, hyaluronan, and chondroitin sulfate, by quantitative enzyme linked immunosorbent assay (ELISA) measurements. For each EGL constituent, its concentration was determined using commercially available ELISA kits, as per manufacturer’s instructions (Syndecan-1, 950.640.096, Diaclone, Besancon Cedex, France; Heparan Sulfate, CSB-E09585h, CusaBio Biotech, Hubei Province, P.R. China; Hyaluronan, DHYAL0, RD Chondroitin Sulfate, CSB-E09587h, CusaBio Biotech, Hubei Province, P.R. China). For each target, all samples were run in triplicate, while standards were run in duplicate; samples were randomly assigned to a triplicate block on the ELISA plate. GDM cases and their matched controls were run on the same ELISA plate. All laboratory staff performing the ELISA was blinded to GDM status and participant matches. Note: an entry of '999' denotes missing data ColumnHeadings: Ethnicity: 1 = Asian, 2 = European, 3 = Other (including African), 4 Polynesian, 5 = Indian; Maternal Age (years); BMI (kg/m^2); GDM Status (1 = GDM, 0 = No GDM); Heparan Sulfate Concentration (ng/ml); Chondroitin Sulfate Concentration (ng/ml); Hyaluronan Concentration (ng/ml); Syndecan-1 Concentration (ng/ml);