Measurement of serum inflammatory markers

Blood draws (20 mL) were taken from the antecubital vein of each participant at 1:00 p.m. at each of the 3 testing sessions (baseline, 1 month, and 3 months). Following extraction, the whole blood was allowed to clot for 30 minutes followed by centrifugation at 1000 × g for 15 minutes. Serum was extracted and immediately stored at −80°C until later analysis. Inflammatory mediators of interest included the pro-inflammatory cytokines IL-2, IL-1β, IL-6, TNF-α, and IFN-γ; the acute phase protein CRP; the anti-inflammatory cytokines IL-4, IL-10, and IL-1RA; and the pro-inflammatory eicosanoid PGE2. Analysis of pro- and anti-inflammatory cytokines was performed in triplicate via the Magpix Multiplex system (EMD Millipore, MA) and analyzed using Luminex software. CRP and PGE2 were analyzed in triplicate and quantified via enzyme-linked immunosorbent assay (R&D Systems, Minneapolis, MN).