Evaluation of Physiological Integrity in Six-Gene-Edited Bama Miniature Pigs as a Model for Xenotransplantation.

Genetically engineered pigs exhibit significant potential as a solution to organ scarcity in xenotransplantation. Nonetheless, a formidable challenge lies in overcoming the rejection of porcine organs by the human immune system. In this study, we generated a six-gene-edited pig model by simultaneously knocking out three major xenoantigens, GGTA1, CMAH, and β4GalNT2, along with the incorporation of coagulation regulatory factor THBD, and two complement regulatory proteins, hCD55 and hCD46, in Bama miniature pigs. These pigs exhibit genetic modifications designed to reduce xenograft rejection while maintaining normal physiology. Assessments of vital organ structure and function, including the heart, liver, spleen, lungs, and kidneys in the gene-edited pigs, showed no abnormalities. The pigs with human transgene knock-in THBD and complement genes exhibited no significant alterations in coagulation function and immune performance. These healthy features hold promise for further xenotransplantation. In conclusion, we successfully constructed six-gene-edited miniature pigs. This work serves as a valuable reference for gene editing strategies for xenotransplantation pig models.