Assessment of Hemoglobin Mass and Blood Volumes at High Altitude Using a Carbon Monoxide Rebreathing Technique

Introduction: Hemoglobin (Hb) and hematocrit (Hct) are conveniently used in clinical practice for diagnosis of anemia and polycythemia. However, altitude-associated physiological adaptations can complicate the interpretation of Hb and Hct values. Plasma volume decreases with altitude and red blood cell volume increases, meaning that increased hemoglobin concentration can be partially due to volume contraction. At moderate altitude, a true increase in the red cell mass is questioned and the increase in Hb may be largely due to volume contraction. Moreover, the WHO diagnostic criteria for polycythemia vera overlap with the reference ranges of Hb and Hct at altitude, resulting in unnecessary work-up of otherwise altitude-associated polycythemia. Direct assessment of Hb mass can overcome this limitation. While historical used methods for direct assessment of red cell mass are complicated and not available in clinical practice, the carbon monoxide (CO) rebreathing method widely used in physiological studies is easier to use, has been tested and recently approved for clinical use. In this study, we aimed to assess (Hb) mass and intravascular volumes (total blood volume [BV], red blood cell volume [RBCV], and plasma volume [PV]) at high altitude, and to explore if moderate altitude is associated with a true increase in Hb mass.

Method: Participants were recruited from a moderately high-altitude city (2,300 m above sea level). Hb mass and intravascular volumes were measured using (CO) rebreathing method. Lean body mass (LBM) was determined using dual-energy X-ray absorptiometry (DEXA). Hb mass and intravascular volumes (BV), (RBCV), and (PV) were reported as total, per body weight (BW), and per LBM, and were compared between genders. To explore an altitude effect, a comparison was made with predicted values generated from sea level dwellers cohort (Oberholzer et al., 2024).

Results: 94 participants (F:30, M:64) were recruited, with a mean age of 25.8±10.6 (18-64). Overall mean parameters were BMI (kg/m²), 27.2±6 (15-43); Hb (g/dL), F: 14.1±1.2, M: 16.5±1.3; and Hct (%), F: 41.4±3.7, M: 48.6±3.7. LBM measurements was available for 21 females and 28 males.

Hb mass and intravascular volumes were as follows: Hb mass, F: 594.9±130.4 g, 9.9±2.4 g/kg^BW, 17.6±3.8 g/kg^LBM; M: 922.8±195 g, 11.7 ±3.1 g/kg^BW, 18.5±3 g/kg^LBM.

Total blood volume, F: 4230±880.7 ml, 70.3±15.3 ml/kg^BW, 126.8±23.7 ml/kg^LBM; M: 5625±1209 ml , 72.3±22 ml/kg^BW, 111±15.3 ml/kg^LBM.

RBCV, F: 1750±388 ml, 29±7 ml/kg^BW, 52±11 ml/kg^LBM, M: 2721±567 ml total, 35±8.8 ml/kg^BW, 54±8.8 ml/kg^LBM.

Plasma volume, F: 2480±551 ml, 41±9 ml/kg^BW, 75±14 ml/kg^LBM; M: 2903±717 ml , 37±13 ml/kg^BW, 57±8 ml/kg^LBM.

Males have higher Hb mass, blood volume, and RBCV compared to females when reported as absolute values or normalized to body weight (p =0.001). However, the gender differences in Hb mass and red blood cell volume greatly diminished when normalized to LBM (p = 0.375). Meanwhile, women have higher total blood volume per LBM (p =0.007) due to higher plasma volume (p =0.0001). High altitude cohort has higher measured Hb mass (g) compared to predicted; F: 594.8±130 versus 523.3±84.6 (p =0.014), M:922.7 ±194 versus 859.6±84, (p =0.01) for body weight-based predication and F: 594.8 ±130 versus 476±54 (p =0.0001), M: 922.7±194 versus 759.7±87, (p =0.0001) for LBM based prediction. Finally, Hb and Hct showed significant but weak (Pearson) correlations with Hb mass and intravascular volumes at altitude.

Conclusion: Change in the Hb and Hct values at altitude does not reflect true change on Hb mass. When corrected to lean body mass, gender differences in Hb mass and red blood cell volume greatly diminished. Lean body mass is needed to reliably interpret Hb mass as measured by CO rebreathing. High-altitude residents have a higher Hb mass when compared with predicted values established from a sea-level cohort. Local sea level controls are needed to reliably establish true altitude-associated change in Hb mass. Assessment of red cell mass by CO rebreathing could be helpful in selecting patients for further evaluation of polycythemia and in those with masked polycythemia (increased red cell mass with normal Hb/HCT) and in differentiating between essential thrombocytosis and polycythemia vera. Establishment of a validated population and altitude-based reference range is an important requisite for applying this CO rebreathing method in clinical practice

No relevant conflicts of interest to declare.