Figure 8. Mechanical perturbation improves nitrosylation levels and viability of RBCs in stored blood.

Human blood from 3 healthy volunteers was collected and stored in static or continuous rocker condition (15 rpm) for 0–24 h at 4 °C. Other stored samples were vortexed (800 rpm for 20 seconds) during storage at 3 time points (0, 12 hr, and 24 hr). (a) Nitrosylation of RBCs in stored blood samples subjected to vortex or rocker conditions was investigated by immunostaining with anti-nitrocysteine antibody. RBCs were imaged by OlympusIX71 microscope, 60× magnification and oil-immersion objective lenses with a numerical aperture (NA) of 1.42. (b) Nitrocysteine levels gradually decreased in static RBC stored for 24 hours compared to 0 h (n = 3; ^#p = 0.025). At 0 hours, RBC in vortexed blood showed higher nitrocysteine levels compared to static RBC (n = 3; *p < 0.001). After 12 hrs and 24 hrs of storage, RBC pre-conditioned by rocker or vortex treatment showed higher nitrocysteine levels than that observed in static blood stored for 12 hr and 24 hr (n = 3; **p < 0.03; and ***p < 0.004, respectively). (c) LDH activity in plasma at 0, 24 hr, 48 hr. LDH activity increased in static blood stored at 24 hr and 48 hr compared with rocked blood (n = 3; p = 0.04; p = 0.007). Inhibition of eNOS by caveolin peptide in stored blood subjected to rocker, decreased RBC viability and lead to increased plasma LDH when compared to similarly treated blood without caveolin peptide at 48 hr (n = 3; p = 0.022).