Non-viral, tumor-free induction of transient cell reprogramming in mouse skeletal muscle to enhance tissue regeneration

Overexpression of Oct3/4, Klf4, Sox2 and c-Myc (OKSM) transcription factors can de-differentiate adult cells in vivo. While sustained OKSM expression triggers tumorigenesis through uncontrolled proliferation of toti- and pluripotent cells, transient reprogramming induces pluripotency-like features and proliferation only temporarily, without teratomas. We sought to transiently reprogram cells within mouse skeletal muscle with a localized injection of plasmid DNA encoding OKSM (pOKSM) and hypothesized that the generation of proliferative intermediates would enhance tissue regeneration after injury. Intramuscular pOKSM administration rapidly upregulated pluripotency (Nanog, Ecat1, Rex1) and early myogenesis genes (Pax3) in the healthy gastrocnemius of various strains. Mononucleated cells expressing such markers appeared in clusters among myofibers, proliferated only transiently and did not lead to dysplasia or tumorigenesis for at least 120 days. Nanog was also upregulated in the gastrocnemius when pOKSM was administered 7 days after surgically sectioning its medial head. Enhanced tissue regeneration after reprogramming was manifested by the accelerated appearance of centro-nucleated myofibers and reduced fibrosis. These results suggest that transient in vivo reprogramming could develop into a novel strategy towards the acceleration of tissue regeneration after injury, based on the induction of transiently-proliferative, pluripotent-like cells in situ. Further research to achieve clinically meaningful functional regeneration is warranted.