Multiple genetic engineering of porcine red blood cells improves compatibility with human blood.

To address critical challenges in clinical blood shortages and limited preservation periods, particularly affecting emergency transfusions, transgenic pig red blood cells (RBCs) emerge as a promising solution. We performed multi-gene editing on pig RBCs by knocking out three Xeno antigen genes (GGTA1, B4GALNT2, CMAH) and introducing high expression of human CD55 and CD47, generating 5-gene edited (5GE) RBCs. After co-incubating with human blood, we assessed their survival rate, along with physicochemical indicators related to hemolytic reactions and inflammatory responses. We further compared the physicochemical properties between transgenic pig blood and human blood. 5GE RBCs showed no significant agglutination, only mild hemolysis, and markedly reduced cytotoxicity. Binding by human antibodies and complement was significantly diminished. This study demonstrates the strong potential of genetically engineered porcine RBCs for emergency human transfusion, offering a viable alternative to alleviate blood shortages, especially when human RBCs are unavailable.