Figure 2.

Changes in intracellular redox state by treatment with t-BuOOH decrease RBC deformability and increase blood viscosity in a concentration-dependent fashion. (A) Changes in intracellular GSH and GSSG levels upon treatment of RBCs with different concentrations of t-BuOOH (0 M, 10⁻⁹ M, 10⁻⁷ M, 10⁻⁶ M, 10⁻⁴ M, 10⁻³ M, 3 × 10⁻³ M, 10⁻¹ M). Free/reduced GSH levels decreased with a simultaneous increase of GSSG, the oxidized form of glutathione (n = 4). GSH: one-way RM ANOVA p = 0.0218, ^**p < 0.01, Dunett's test vs. untreated control. GSSG: one-way RM ANOVA p < 0.0001, ^**p < 0.01, ^*** p < 0.001, Dunett's test vs. untreated control. (B) Ratio of free/reduced GSH and GSSG calculated from the values in (A). One-way RM ANOVA p < 0.0001, ^*** p < 0.001, Dunett's test vs. untreated control. (C) Relationship between elongation index (EI) and applied shear stress (0.30–53.33 Pa) of whole blood samples treated with increasing concentrations of t-BuOOH (3–7 mM) showing that t-BuOOH impairs RBC deformability in a concentration-dependent fashion (n = 6). Two-way RM ANOVA p < 0.001 and Dunett's test vs. untreated control; ^*p ≤ 0.05, ^***p < 0.001 (statistically significant for 3 mM from shear stresses between 0.3 and 9.48 Pa; for 5 and 7 mM over the whole range of shear stresses). (D) Analysis of blood viscosity of the same samples described in (C) over a range of different shear rates (n = 6). Two-way ANOVA p = 0.0004 and Dunnett's test vs. untreated control, ^* p ≤ 0.05 (for 5 mM statistically significant from shear rates between 0.5 and 3.98 s⁻¹ and for 7 mM from shear rates between 0.5 and 11.22 s⁻¹). (E) Comparison of changes in viscosity and EI at matching shear rates show inverse relationship between the two parameters.