Biochemical Measurements

Fasting plasma glucose, plasma lipids, and creatinine were measured with enzymatic-colorimetric assays (Biosystems, Spain). Glycated hemoglobin A1c (HbA1c) was determined using a National Glycohemoglobin Standardization Program-certified boronate affinity technique (NycoCard^® Reader II, Alere Technologies, Norway).

Myonectin was measured with the Aviscera Bioscience Myonectin ELISA (Cat# SK00393-10, dynamic range 8–5,000 ng/mL, intra-assay CV: 6.0–8.0%, interassay CV: 12.0–14.0%), myostatin with the R&D systems GDF-8/Myostatin Quantikine ELISA (Cat#DGDF80, dynamic range 31.3–2,000 pg/mL, intra-assay CV: 1.8–5.4%, interassay CV: 3.1–6.0%) and FGF-21 with the R&D Systems Human FGF-21 Quantikine ELISA (Cat #DF2100, dynamic range 31.3–2,000 pg/mL, intra-assay CV: 2.9–3.9%, interassay CV: 5.2–10.9%). hsCRP was measured using R&D Systems Human C-Reactive Protein/CRP Immunoassay (CAT# DCRP00, dynamic range 0.010–50 ng/mL, intra-assay CV: 3.8–8.3%, interassay CV: 6.0–7.0%). All biomarker immunoassays were based on double-sandwich or competitive-ELISA techniques. Absorbance determinations were done in a BioTek Synergy HT Reader^® using the appropriate wavelength according to protocol specifications. All measurements were performed in duplicate. Procedures were executed at the Diabetes, Lipids and Metabolism laboratory of Universidad de Los Andes, following current institutional biosafety protocols.