Abstract
Red blood cell (RBC) deformability is an essential component of microcirculatory function that appears to be enhanced by physiological shear stress, while being negatively affected by supraphysiological shears and/or free radical exposure. Given that blood contains RBCs with non-uniform physical properties, whether all cells equivalently tolerate mechanical and oxidative stresses remains poorly understood. We thus partitioned blood into old and young RBCs which were exposed to phenazine methosulfate (PMS) that generates intracellular superoxide and/or specific mechanical stress. Measured RBC deformability was lower in old compared to young RBCs. PMS increased total free radicals in both sub-populations, and RBC deformability decreased accordingly. Shear exposure did not affect reactive species in the sub-populations but reduced RBC nitric oxide synthase (NOS) activation and intriguingly increased RBC deformability in old RBCs. The co-application of PMS and shear exposure also improved cellular deformability in older cells previously exposed to reactive oxygen species (ROS), but not in younger cells. Outputs of NO generation appeared dependent on cell age; in general, stressors applied to younger RBCs tended to induce S-nitrosylation of RBC cytoskeletal proteins, while older RBCs tended to reflect markers of nitrosative stress. We thus present novel findings pertaining to the interplay of mechanical stress and redox metabolism in circulating RBC sub-populations.
Highlights
Effective microcirculatory function requires highly deformable red blood cells (RBCs), a property classically shown to be governed by the cell’s exceptional surface area-to-volume ratio, cytoplasmic viscosity being greater than the surrounding fluid, and the visco-elastic properties of the plasma membrane [1]
Application of shear stress to phenazine methosulfate (PMS)-treated old RBCs resulted in a further increase in cytosolic reactive oxygen species (ROS)/RNS (ES = 0.4, p < 0.05), while no significant effect of shear stress was observed in young RBCs
Exposure of PMS-treated old sub-fractions to shear stress significantly increased Total antioxidant capacity (TAC) compared to old non-sheared cells (ES = 0.6, p < 0.05; Figure 1C), while no effect was observed in young RBC fractions
Summary
Effective microcirculatory function requires highly deformable red blood cells (RBCs), a property classically shown to be governed by the cell’s exceptional surface area-to-volume ratio, cytoplasmic viscosity being greater than the surrounding fluid, and the visco-elastic properties of the plasma membrane [1]. Increased NO bioavailability is generally beneficial for RBCs, as NO may be oxidized to nitrite and nitrate [21,22] and/or may transiently bind to exposed thiols within active cysteine residues in a process termed S-nitrosylation [23]. This post-translational modification has been observed in the major RBC cytoskeletal proteins α- and β-spectrin, which are pivotal in governing the flexibility of the cell membrane, and it is thought that this process contributes to the NO-dependent increase in RBC deformability [24]. It is clear that the impact of NO on cellular mechanics of RBC is complex and dependent upon the local environment
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