Hepcidin and Erythroferrone Complement the Athlete Biological Passport in the Detection of Autologous Blood Transfusion.

PURPOSE: We investigated whether hepcidin and erythroferrone (ERFE) could complement the athlete biological passport (ABP) in indirectly detecting a 130-mL packed red blood cells (RBC) autologous blood transfusion. Endurance performance was evaluated. METHODS: Forty-eight healthy men ( n = 24) and women ( n = 24) participated. Baseline samples were collected weekly followed by randomization to a blood transfusion (BT, n = 24) or control group (CON, n = 24). Only the BT group donated 450 mL whole blood from which 130 mL red blood cell was reinfused 4 wk later. Blood samples were collected 3, 7, 14, 21, and 28 d after donation, and 3, 6, and 24 h and 2, 3, and 6 d after reinfusion. In the CON group samples were collected with the same frequency. Endurance performance was evaluated by a 650-kCal time trial ( n = 13) before and 1 and 6 d after reinfusion. RESULTS: A time-treatment effect existed ( P < 0.05) for hepcidin and ERFE. Hepcidin was increased ( P < 0.01) ~110 and 89% 6 and 24 h after reinfusion. Using an individual approach (99% specificity, e.g., allowing 1:100 false-positive), sensitivities, i.e., true positives, of 30% and 61% was found for hepcidin and ERFE, respectively. For the ABP, the most sensitive marker was Off-hr score ([Hb] (g·L -1 ) - 60 × âˆšRET%) ( P < 0.05) with a maximal sensitivity of ~58% and ~9% after donation and reinfusion, respectively. Combining the findings for hepcidin, ERFE, and the ABP yielded a sensitivity across all time-points of 83% after reinfusion in BT. Endurance performance increased 24 h (+6.4%, P < 0.01) and 6 d after reinfusion (+5.8%, P < 0.01). CONCLUSIONS: Hepcidin and ERFE may serve as biomarkers in an antidoping context after an ergogenic, small-volume blood transfusion.

An Untargeted Urine Metabolomics Approach for Autologous Blood Transfusion Detection.

PURPOSE: Autologous blood transfusion is performance enhancing and prohibited in sport but remains difficult to detect. This study explored the hypothesis that an untargeted urine metabolomics analysis can reveal one or more novel metabolites with high sensitivity and specificity for detection of autologous blood transfusion. METHODS: In a randomized, double-blinded, placebo-controlled, crossover design, exercise-trained men (n = 12) donated 900 mL blood or were sham phlebotomized. After 4 wk, red blood cells or saline were reinfused. Urine samples were collected before phlebotomy and 2 h and 1, 2, 3, 5, and 10 d after reinfusion and analyzed by ultraperformance liquid chromatography-quadrupole time-of-flight mass spectrometry. Models of unique metabolites reflecting autologous blood transfusion were attained by partial least-squares discriminant analysis. RESULTS: The strongest model was obtained 2 h after reinfusion with a misclassification error of 6.3% and 98.8% specificity. However, combining only a few of the strongest metabolites selected by this model provided a sensitivity of 100% at days 1 and 2 and 66% at day 3 with 100% specificity. Metabolite identification revealed the presence of secondary di-2-ethylhexyl phtalate metabolites and putatively identified the presence of (iso)caproic acid glucuronide as the strongest candidate biomarker. CONCLUSIONS: Untargeted urine metabolomics revealed several plasticizers as the strongest metabolic pattern for detection of autologous blood transfusion for up to 3 d. Importantly, no other metabolites in urine seem of value for antidoping purposes.

Autologous Blood Transfusion Enhances Exercise Performance-Strength of the Evidence and Physiological Mechanisms.

This review critically evaluates the magnitude of performance enhancement that can be expected from various autologous blood transfusion (ABT) procedures and the underlying physiological mechanisms. The review is based on a systematic search, and it was reported that 4 of 28 studies can be considered of very high quality, i.e. placebo-controlled, double-blind crossover studies. However, both high-quality studies and other studies have generally reported performance-enhancing effects of ABT on exercise intensities ranging from ~70 to 100% of absolute peak oxygen uptake (VO2peak) with durations of 5-45 min, and the effect was also seen in well-trained athletes. A linear relationship exists between ABT volume and change in VO2peak. The likely correlation between ABT volume and endurance performance was not evident in the few available studies, but reinfusion of as little as 135 mL packed red blood cells has been shown to increase time trial performance. Red blood cell reinfusion increases endurance performance by elevating arterial oxygen content (CaO2). The increased CaO2 is accompanied by reduced lactate concentrations at submaximal intensities as well as increased VO2peak. Both effects improve endurance performance. Apparently, the magnitude of change in haemoglobin concentration ([Hb]) explains the increase in VO2peak associated with ABT because blood volume and maximal cardiac output have remained constant in the majority of ABT studies. Thus, the arterial-venous O2 difference during exercise must be increased after reinfusion, which is supported by experimental evidence. Additionally, it remains a possibility that ABT can enhance repeated sprint performance, but studies on this topic are lacking. The only available study did not reveal a performance-enhancing effect of reinfusion on 4 × 30 s sprinting. The reviewed studies are of importance for both the physiological understanding of how ABT interacts with exercise capacity and in relation to anti-doping efforts. From an anti-doping perspective, the literature review demonstrates the need for methods to detect even small ABT volumes.

Time Trial Performance Is Sensitive to Low-Volume Autologous Blood Transfusion.

PURPOSE: This study tested the hypothesis that autologous blood transfusion (ABT) of ~50% of the red blood cells (RBC) from a standard 450-mL phlebotomy would increase mean power in a cycling time trial. In addition, the study investigated whether further ABT of RBC obtained from another 450-mL phlebotomy would increase repeated cycling sprint ability. METHODS: In a randomized, double-blind, placebo-controlled crossover design (3-month wash-out), nine highly trained male subjects donated two 450-mL blood bags each (BT trial) or were sham phlebotomized (PLA trial). Four weeks later, a 650-kcal time trial (n = 7) was performed 3 d before and 2 h after receiving either ~50% (135 mL) of the RBC or a sham transfusion. On the following day, transfusion of RBC (235 mL) from the second donation or sham transfusion was completed. A 4 × 30-s all-out cycling sprint interspersed by 4 min of recovery was performed 6 d before and 3 d after the second ABT (n = 9). RESULTS: The mean power was increased in time trials from before to after transfusion (P < 0.05) in BT (213 ± 35 vs 223 ± 38 W; mean ± SD) but not in PLA (223 ± 42 vs 224 ± 46 W). In contrast, the mean power output across the four 30-s sprint bouts remained similar in BT (639 ± 35 vs 644 ± 26 W) and PLA (638 ± 43 vs 639 ± 25 W). CONCLUSIONS: ABT of only ~135 mL of RBC is sufficient to increase mean power in a 650-kcal cycling time trial by ~5% in highly trained men. In contrast, a combined high-volume transfusion of ~135 and ~235 mL of RBC does not alter 4 × 30-s all-out cycling performance interspersed with 4 min of recovery.