Comprehensive Biomarker Profiling in Diabetes

HbA_1c was determined by high-performance liquid chromatography (Menarini Diagnostics, Firenze, Italy). Total serum cholesterol, triglycerides, HDL cholesterol, and LDL cholesterol were measured using standard enzymatic methods. Estimated glomerular filtration rate (eGFR) was calculated using the Chronic Kidney Disease-Epidemiology (CKD-EPI) equation on the basis of serum creatinine, age, sex and race [18 (link)]. Serum 25(OH)D was measured using an ELISA kit (BioVendor, Brno, Czech Republic). Plasma FGF-23 was measured using an ELISA kit (Immunotopics Inc. San Clemente, CA, USA). Intra- and inter-assay coefficients of variation were 5% and 7.8% for 25(OH)D and 2.4% and 4.7% for FGF-23, respectively. hsCRP was determined by immunonephelometry (Siemens, Munich, Germany). IL-6 was determined by ELISA (R&D Systems, Oxon, UK), as well as sTNFαR1 (HyCult Biotech, Uden, The Netherlands) and sTNFαR2 (R&D Systems, Oxon, UK). Concentrations of sICAM-1, sVCAM-1 and E-selectin were also determined by ELISA (Wuhan Boster Biological Technology, Ltd., Wuhan, China) as previously described [15 (link),19 (link)].