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Red blood cell (RBC) transfusions have been a life-saving procedure following trauma for decades. Ex vivo storage of RBCs is limited to 42 days, as the RBCs degrade during storage. Recent evidence suggests that transfusions of stored RBCs are associated with adverse events and increased mortality. Blood shortages are frequent and are expected to increase as the population ages. My research explored new methods to prolong ex vivo RBC storage, and alternatives to RBCs. A novel improvement to RBC storage is storage under anaerobic conditions to minimize oxidative stress. Anaerobically stored RBCs improved RBC recovery and resuscitation from hemorrhagic shock compared to conventionally stored RBCs. However, anaerobic storage only slowed the degradation of RBCs, without further extending the RBC storage period. Therefore, we explored hemoglobin (Hb)-based O2 carriers (HBOCs) as an alternative to RBCs. HBOCs are advantageous relative to blood, as they are shelf-stable, not limited by donors for supply, and do not require blood-type matching before infusion. Clinical trials of previous generations of HBOCs failed due to unforeseen side-effects. The next generation of HBOCs involves high molecular weight Polymerized Hb (PolyHb), which is the most promising and scalable HBOC formulation that alleviates the side effects of previous HBOCs. We compared fresh and stored PolyHb to RBCs in a hemorrhagic shock resuscitation model. The results confirm that PolyHb was as effective as fresh RBCs to recover from shock and significantly superior to stored RBCs, and that PolyHb’s properties or efficacy did not change during long term storage. Finally, we assessed the safety of PolyHb in guinea pigs, as they are more human-relevant than other preclinical models. This study demonstrated that increasing the molecular size of PolyHb alleviates side-effects of previous HBOC formulations, and that PolyHb toxicity is transient. Future work will focus on the safety of PolyHb, particularly in vulnerable patient populations.

BACKGROUND: Hemoglobin (Hb) based oxygen carriers (HBOCs) have been proposed as alternatives to blood for decades. Previous studies demonstrated that large molecular diameter HBOCs based on polymerized bovine Hb (PolybHb) attenuate Hb side-effects and toxicity. The objective of this study was to test the safety and efficacy of tense state PolybHb after long-term storage. METHODS AND RESULTS: PolybHb was subjected to diafiltration to remove low molecular weight (< 500 kDa) species and stored for 2 years. PolybHb was studied in parallel with blood, collected from rats and stored leukodepleted under blood bank conditions for 3 weeks. Rats were hemorrhaged and resuscitated to 90% of the blood pressure before the hemorrhage with fresh blood, stored blood, fresh PolybHb, or two-year-stored PolybHb. Hemorrhagic shock impaired oxygen delivery and cardiac function. Resuscitation restored blood pressure and cardiac function, but stored blood required a significantly larger transfusion volume to recover from shock compared to fresh blood and PolybHb (fresh and stored). Stored blood transfusion elevated markers of organ damage compared to all other groups. CONCLUSIONS: These studies indicate that large molecular diameter PolybHb is as efficacious as fresh blood in restoring cardiac function and confirm the lack of degradation of PolybHb’s safety or efficacy during long-term storage.

Author(s): Williams, Alexander T; Jani, Vivek P; Nemkov, Travis; Lucas, Alfredo; Yoshida, Tatsuro; Dunham, Andrew; D'Alessandro, Angelo; Cabrales, Pedro | Abstract: BackgroundResuscitation from hemorrhagic shock (HS) by blood transfusion restores oxygen (O2) delivery and provides hemodynamic stability. Current regulations allow red blood cells (RBCs) to be stored and used for up to 42 days. During storage, RBCs undergo many structural and functional changes. These storage lesions have been associated with adverse events and increased mortality after transfusion, increasing the need for improved RBC storage protocols. This study evaluates the efficacy of anaerobically stored RBCs to resuscitate rats from severe HS compared with conventionally stored RBCs.Methods and resultsRat RBCs were stored under anaerobic, anaerobic/hypercapnic, or conventional conditions for a period of 3 weeks. Hemorrhage was induced by controlled bleeding, shock was maintained for 30 min, and RBCs were transfused to restore and maintain blood pressure near the prhemorrhage level. All storage conditions met current regulatory 24-h posttransfusion recovery requirements. Transfusion of anaerobically stored RBCs required significantly less RBC volume to restore and maintain hemodynamics. Anaerobic or anaerobic/hypercapnic RBCs restored hemodynamics better than conventionally stored RBCs. Resuscitation with conventionally stored RBCs impaired indices of left ventricular cardiac function, increased hypoxic tissue staining and inflammatory markers, and affected organ function compared with anaerobically stored RBCs.ConclusionsResuscitation from HS via transfusion of anaerobically stored RBCs recovered cardiac function, restored hemodynamic stability, and improved outcomes.