Effect of physical fitness and endurance exercise on indirect biomarkers of growth hormone and insulin misuse: Immunoassay-based measurement in urine samples.

Indirect biomarkers of recombinant human growth hormone (rhGH), insulin-like growth factor-I (IGF-I), insulin-like growth factor-II (IGF-II), insulin-like growth factor binding proteins (IGFBP-2 and IGFBP-3) and insulin (C-peptide) were measured together with urinary parameters of renal damage (beta(2)-microglobulin and proteinuria) by immunoassays, in house validated for the purpose, in 61 subjects (36 elite athletes, 18 recreational athletes and 7 sedentary individuals) with different levels of physical fitness and endurance exercise. Validation parameters were good for the evaluated assays, excluding a high inter-assay imprecision and inaccuracy of 24 and 26% obtained for GH assay. The range of concentrations found in urine samples under investigation was generally covered by the calibration curves of the studied immunoassays. However, for the samples below or above the calibration curve, opportune dilution or concentration were performed. Particularly, C-peptide samples had to be diluted 1:5 and beta(2)-microglobulin ones assayed using a triple sample volume, to fall within the calibration range. Urinary C-peptide was the only biomarker statistically higher in samples of elite athletes when compared to recreational athletes and sedentary individuals. Among elite athletes, tae-kwon-do athletes showed the highest IGF-II basal values while weightlifting athletes showed the lower IGF-I and IGFBP-3 basal values. The trend observed in weightlifters' basal samples was confirmed in their training samples: IGF-I, IGF-II, IGFBP-3 and beta(2)-microglobulin were lower in with respect to those from synchronised swimming. Over the training season, within athlete variability was observed for IGFBP-3 for weightlifting athletes. In the studied subjects, no direct associations were found between biomarkers of GH or insulin misuse and urinary parameters of renal damage, eventually due to high-workload endurance training. The variations observed in different biomarkers should be taken in consideration in the hypothesis of setting reference concentration ranges for doping detection.