Effects of simulated altitude (normobaric hypoxia) on cardiorespiratory parameters and circulating endothelial precursors in healthy subjects

Abstract

Background: Circulating Endothelial Precursors (PB-EPCs) are involved in the maintenance of the endothelial compartment being promptly mobilized after injuries of the vascular endothelium, but the effects of a brief normobaric hypoxia on PB-EPCs in healthy subjects are scarcely studied.

Methods: Clinical and molecular parameters were investigated in healthy subjects (n = 8) in basal conditions (T0) and after 1 h of normobaric hypoxia (T1), with Inspiratory Fraction of Oxygen set at 11.2% simulating 4850 mt of altitude. Blood samples were obtained at T0 and T1, as well as 7 days after hypoxia (T2).

Results: In all studied subjects we observed a prompt and significant increase in PB-EPCs, with a return to basal value at T2. The induction of hypoxia was confirmed by Alveolar Oxygen Partial Pressure (PAO2) and Spot Oxygen Saturation decreases. Heart rate increased, but arterial pressure and respiratory response were unaffected. The change in PB-EPCs percent from T0 to T1 was inversely related to PAO2 at T1. Rapid (T1) increases in serum levels of hepatocyte growth factor and erythropoietin, as well as in cellular PB-EPCs-expression of Hypoxia Inducible Factor-1alpha were observed.

Conclusion: In conclusion, the endothelial compartment seems quite responsive to standardized brief hypoxia, possibly important for PB-EPCs activation and recruitment.

Figure 1

Flow cytometry evaluation of circulating endothelial precursors cells (EPCs). (A) Representative panel showing the analysis gate used to exclude platelets and debris. (B) The gate used to exclude CD45-positive hematopoietic cells. (C, D) Representative panels showing the EPCs before (T0) and after (T1) hypoxia exposure. PerCP, peridin chlorophyll protein; PE, phycoerythrin; FITC, fluorescein isothiocyanate.

Figure 2

Regression plot showing the correlation between changes in PB-EPCs from T0 to T1 and levels of PAO₂ at T1 (r = 0.73; p = 0.03). PB-EPCs ΔT0-T1, peripheral blood endothelial precursors change from T0 to T1; PAO₂ T1, Alveolar Oxygen Partial Pressure at T1.

Figure 3

Molecular changes in serum and PB-EPCs after normobaric hypoxia. Normal subjects were examined before (T0) and 1 h (T1) after experimental hypoxia. Serum samples were used for HGF (A), Epo (C) and Et-1 (D) evaluation. HGF data are reported as relative fold-increases, calculated using the absolute values (T0 = 2.9 ± 0.3 ng/ml). All the data were analysed by ANOVA, and the values reported are the means ± S.E. of experiments performed in triplicate. A p value < 0.05 was considered significant. (B) PB-EPCs, prepared on slides, were used to examine HIF-1α and CXCR4 expression by immunofluorescence. Specific stains with anti-HIF-1α or anti-CXCR4 antibody followed by the appropriate secondary antibody (green, a); nuclear staining with DAPI (blu, b); merged image (c). Images were taken using fluorescence microscopy at 400 × magnification.