Exogenous erythropoietin increases hematological status, fat oxidation, and aerobic performance in males following prolonged strenuous training

Abstract

This study investigated the effects of EPO on hemoglobin (Hgb) and hematocrit (Hct), time trial (TT) performance, substrate oxidation, and skeletal muscle phenotype throughout 28 days of strenuous exercise. Eight males completed this longitudinal controlled exercise and feeding study using EPO (50 IU/kg body mass) 3×/week for 28 days. Hgb, Hct, and TT performance were assessed PRE and on Days 7, 14, 21, and 27 of EPO. Rested/fasted muscle obtained PRE and POST EPO were analyzed for gene expression, protein signaling, fiber type, and capillarization. Substrate oxidation and glucose turnover were assessed during 90-min of treadmill load carriage (LC; 30% body mass; 55 ± 5% V̇O₂peak) exercise using indirect calorimetry, and 6-6-[²H₂]-glucose PRE and POST. Hgb and Hct increased, and TT performance improved on Days 21 and 27 compared to PRE (p < 0.05). Energy expenditure, fat oxidation, and metabolic clearance rate during LC increased (p < 0.05) from PRE to POST. Myofiber type, protein markers of mitochondrial biogenesis, and capillarization were unchanged PRE to POST. Transcriptional regulation of mitochondrial activity and fat metabolism increased from PRE to POST (p < 0.05). These data indicate EPO administration during 28 days of strenuous exercise can enhance aerobic performance through improved oxygen carrying capacity, whole-body and skeletal muscle fat metabolism.

Keywords: V̇O2peak; hematocrit; hemoglobin; mitochondria; substrate oxidation.

FIGURE 1

Hematological adaptations to training and erythropoietin administration. (A) Hemoglobin (g/dL) assessed throughout the 28‐day intervention. (B) Hematocrit (%) assessed throughout the 28‐day intervention. Data represented as mean ± SD. Timepoints not sharing letters are different than each other. n = 8 males. Statistical test = mixed model ANOVA. Significance determined as p < 0.05.

FIGURE 2

Serum analytes in response to training and erythropoietin (EPO) administration. (a) Change in Interleukin‐6 (IL‐6, pg/mL) assessed prior to and 80 min into load carriage exercise at PRE and POST timepoints. (b) Hepcidin (ng/mL) assessed prior to and 80 min into load carriage exercise at PRE and POST timepoints. Dotted line at 0 represents the blood draw prior to load carriage. Data represented as mean ± SD. n = 8 males. Statistical test = A: paired t‐test, B: Wilcoxon matched‐pairs sign rank test.

FIGURE 3

Performance adaptations to training and erythropoietin administration. (A) Treadmill V̇O₂peak (mL/kg/min) assessed throughout the 28‐day intervention. (B) Five‐km treadmill time to completion (s) assessed throughout the 28‐day intervention. Individual data points represented as circles. Data represented as mean ± SD. Timepoints not sharing letters are different than each other. n = 8 males. Statistical test = mixed model ANOVA. Significance determined as p < 0.05.

FIGURE 4

Substrate dynamics in response to training and erythropoietin administration. (A) Carbohydrate Oxidation (g/min) during the load carriage exercise PRE and POST. (B) Fat Oxidation (g/min) during the load carriage exercise PRE and POST. (C) Glucose rate of appearance (mg/kg/min) during the load carriage exercise PRE and POST. (D) Glucose rate of disappearance (mg/kg/min) during the load carriage exercise PRE and POST. (E) Metabolic Clearance Rate (mL/kg/min) during the load carriage exercise PRE and POST. Data represented as mean ± SD. n = 8 males. Timepoints not sharing letters are different than each other; p < 0.05. Numeric p value = different than PRE. Statistical test = mixed model ANOVA.

FIGURE 5

Myofiber type and mitochondrial indices. (a) Representative image of observed myofiber isoforms. Red = Type I, green = IIA, dark green = IIX/IIAX, and red/green = I/IIA. (b) Myofiber type proportion PRE (fibers analyzed = 586 ± 157, n = 7). (c) Mean number of myofibers analyzed at PRE and POST (n = 5). (d) Myofiber type distribution PRE and POST (n = 5). (e) COXIV skeletal muscle protein signaling at PRE and POST timepoints (n = 8) (f) PPARGC1 gene expression at PRE and POST timepoints (n = 8). (g) TFAM gene expression at PRE and POST timepoints (n = 8). All data are represented as mean ± SD. Statistical test = paired t‐test. Blot images with the target protein and the molecular weight where membranes were cut for all participants can be found in Figure S3.

FIGURE 6

Capillary content and angiogenic gene expression. (a) Representative image of capillary stain. Red = Type I fibers, Black = Type II fibers, green = capillaries. (b) Capillary density PRE and POST (n = 5). (c) Capillary contacts by Type I, Type II, and total fibers PRE and POST (n = 5). (d) VEGFA skeletal muscle gene expression at PRE and POST timepoints (n = 8). All data are represented as mean ± SD. Statistical test = paired T‐test.

FIGURE 7

Skeletal muscle gene expression at PRE and POST timepoints. (a) COXIV. (b) HADHA. (c) FABP3. (d) CPT1A. (e) CD36. (f) FASN. (g) ACACA. Data expressed as Log(2^−ΔΔCT) relative to PRE. n = 8 males. Statistical test = paired t‐test.