Analytical Studies
Blood samples were collected in Ethylene-Diamine-Tetra-Acetic Acid and centrifuged at 2,500 g for 10 minutes and plasma was stored at -80°C. Patients were seen every year at their hospital.
The investigators who performed the laboratory determinations were unaware of the clinical data. Plasma concentrations of MCP-1 and galectin-3 were determined in duplicate using commercially available enzyme-linked immunosorbent assay kits (DCP00, R&D Systems and BMS279/2 Bender MedSystems respectively) following the manufacters’ instructions. Hs-CRP was assessed by latex-enhanced immunoturbidimetry (ADVIA 2400 Chemistry System, Siemens, Germany), NT-pro-BNP by immunoassay (VITROS, Orthoclinical Diagnostics, U.S.A.), and hsTnI was assessed by direct quimioluminiscence (ADVIA Centaur, Siemens, Germany). Plasma calcidiol levels were quantified by chemiluminescent immunoassay (CLIA) on the LIAISON XL analyzer (LIAISON 25OH-Vitamin D total Assay DiaSorin, Saluggia, Italy). Lipids, glucose, and creatinin were determined by standard methods (ADVIA 2400 Chemistry System, Siemens, Germany).