Previous Cryopreservation Alters the Natural History of the Red Blood Cell Storage Lesion

Background: During storage, packed red blood cells (pRBCs) undergo a number of biochemical, metabolic, and morphologic changes, collectively known as the "storage lesion." We aimed to determine the effect of cryopreservation on the red blood cell storage lesion compared with traditional 4°C storage. Methods: Previously cryopreserved human pRBCs were compared with age-matched never-frozen pRBCs obtained from the local blood bank. The development of the red cell storage lesion was evaluated after 7, 14, 21, 28, and 42 days of storage at 4°C in AS-3 storage medium. We measured physiological parameters including cell counts, lactic acid, and potassium concentrations as well as signs of eryptosis including loss of phosphatidylserine (PS) asymmetry, microparticle production, and osmotic fragility in hypotonic saline. Results: Compared with controls, previously cryopreserved pRBC at 7 days of storage in AS-3 showed lower red cell counts (3.7 vs. 5.3 × 10 6 cells/μL, P < 0.01), hemoglobin (Hgb) (12.0 vs. 16.5 g/dL, P < 0.01), hematocrit (33.0% vs. 46.5%, P < 0.01), and pH (6.27 vs. 6.72, P < 0.01). Over 28 days of storage, storage cryopreserved pRBC developed increased cell-free Hgb (0.7 vs. 0.3 g/dL, P < 0.01), greater PS exposure (10.1% vs. 3.3%, P < 0.01), and microparticle production (30,836 vs. 1,802 MP/μL, P < 0.01). Previously cryopreserved cells were also less resistant to osmotic stress. Conclusion: The red blood cell storage lesion is accelerated in previously cryopreserved pRBC after thawing. Biochemical deterioration of thawed and deglycerolized red cells suggests that storage time before transfusion should be limited to achieve similar risk profiles as never-frozen standard liquid storage pRBC units.