Effect of HR stress in PA or CON C57Bl/6 (C57; n = 7 per group) and SAD mice (n = 6 per group). Eight weeks of a voluntary wheel-running protocol were conducted in C57 and SCD SAD mice. Before euthanizing the animals, HR stress (4 hours hypoxic stress at 6.5% oxygen followed by 2 hours of reoxygenation in ambient air) was used to stimulate the pathophysiological parameters known to trigger the VOC in this mouse model. (A) P-selectin and (B) VCAM-1 immunostaining scores in the lung. Immunohistologic slides were blindly semiquantified by 3 experienced anatomists for VCAM-1 and P-selectin with the staining intensity. For each vessel (15 to 20 vessels per mouse), a score from 0 to 3 was attributed (0, no immunostaining; 1, <25% vessel staining; 2, <50% vessel staining; 3, >50% vessel staining). For each mouse, the immunostaining score was the mean of each vessel score from a section. (C) Representative staining for VCAM-1 and P-selectin in the blood vessel of lungs. Magnification ×40. (D) Nitrite/nitrate (NOx) concentrations in the lungs. The Griess method was used: the sum of nitrite and nitrate in the plasma is considered an index of NO production. (E) Plasma concentrations of lactate dehydrogenase (LDH). *P < .05; #, P < .05 vs C57CONHR; §, P < .05 vs C57PAHR; a, P = .07. Values are means ± standard deviation. All variables were tested for normality and variance homogeneity. P-selectin immunostaining and NOx concentrations were tested with nonparametric Kruskall-Wallis test followed by Mann-Whitney U test. The other variables were compared using factorial analysis of variance followed by planned comparisons. Nx, normoxic conditions.
