Humanized skeletal muscle in MYF5/MYOD/MYF6-null pig embryos

Because post-mortem human skeletal muscle is not viable, autologous muscle grafts are typically required in tissue reconstruction after muscle loss due to disease or injury. However, the use of autologous tissue often leads to donor-site morbidity. Here, we show that intraspecies and interspecies chimaeric pig embryos lacking native skeletal muscle can be produced by deleting the MYF5, MYOD and MYF6 genes in the embryos via CRISPR, followed by somatic-cell nuclear transfer and the delivery of exogenous cells (porcine blastomeres or human induced pluripotent stem cells) via blastocyst complementation. The generated intraspecies chimaeras were viable and displayed normal histology, morphology and function. Human:pig chimaeras generated with TP53-null human induced pluripotent stem cells led to higher chimaerism efficiency, with embryos collected at embryonic days 20 and 27 containing humanized muscle, as confirmed by immunohistochemical and molecular analyses. Human:pig chimaeras may facilitate the production of exogenic organs for research and xenotransplantation. Human:pig embryos can be produced by deleting the MYF5, MYOD and MYF6 genes in pig embryos via CRISPR and somatic-cell nuclear transfer, followed by the delivery of TP53-null human induced pluripotent stem cells via blastocyst complementation.

Automated summary

Through CRISPR technology, deletion of MYF5, MYOD and MYF6 genes in pig embryos can lead to the production of intraspecies and interspecies chimaeric pig embryos lacking native skeletal muscle, followed by somatic-cell nuclear transfer and the delivery of exogenous cells through blastocyst complementation. The generated chimaeric pig embryos exhibited normal histology, morphology and function, showing potential for research and xenotransplantation of exogenic organs. Furthermore, human:pig chimaeras created with TP53-null human induced pluripotent stem cells demonstrated higher chimaerism efficiency, with humanized muscle observed in embryos at different embryonic stages.