Hemodynamic and hematological measures.

Whole blood, serum, and plasma samples were collected via venipuncture and centrifugation before exercise on day 1 and day 15 for hematological, blood chemistry, and liver function assessments. Hemoglobin concentration was determined from complete blood count (CBC) measurements using fresh whole blood. The oxygen partial pressure at which Hb is 50% saturated (p50) was determined from oxygen association curves using fresh whole blood from an arterial blood sample and reported at standard conditions: 37°C, 40 mmHg carbon dioxide partial pressure, and a pH of 7.40 (Hemox Analyzer, TCS Scientific Corporation, Southampton). In our laboratory, the coefficient of variation for historical data in a large cohort of healthy control subjects was 3.7% with a mean p50 of 28.6 ± 1.05 mmHg. Erythropoietin (EPO) concentration, blood chemistries, and liver function were assessed from serum and plasma samples according to guidelines (26).

An arterial catheter (FA-04020, Arrow International Inc., Reading, PA) was placed under local anesthesia (2% lidocaine) by an experienced anesthesiologist for arterial blood draws and continuous blood pressure recording. At rest and during submaximal exercise, an arterial blood sample was collected for blood gas analysis (IL-1620, Instrumentation Laboratories, Lexington), including arterial oxygen saturation (SaO2), partial pressure of arterial oxygen (PaO2) and carbon dioxide (PaCO2), pH, Hb, and carboxyhemoglobin (COHb). Arterial oxygen content (CaO2) was computed as CaO2 = (1.34 × Hb × SaO2) + (PaO2 × 0.0031). Venous oxygen content (CvO2) and the arterial-venous oxygen content difference (Ca-v O₂) were inferred by rearranging the Fick equation, and fractional O₂ extraction was calculated by dividing Ca-v O₂ difference by CaO2. Oxygen delivery (Do₂) was calculated as the product of cardiac output and CaO2. Estimates of regional tissue oxygenation (r-So₂) were continuously recorded from a sensor (SensSmart Model 8004CA, Nonin Medical, Inc., Plymouth, MN) placed on the midline of the Vastus Lateralis via near-infrared spectroscopy (Equanox Model 7600, Nonin Medical Inc, Minnesota).