Deciphering in vivo reprogramming and its rejuvenating potential

Resumo

The manipulation of cell fates through reprogramming is one of the most exciting advances in recent years. The most striking breakthrough in the field occurred when Yamanaka first illustrated the possibility to convert differentiated cells into pluripotent stem cells by the ectopic expression of four transcription factors, Oct4, Sox2, Klf4 and c-Myc or OSKM. Our laboratory has demonstrated that transient expression of these four Yamanaka factors leads to teratoma formation in mice indicative of in vivo reprogramming. We were interested in deciphering the intermediate events of de-differentiation induced by OSKM in vivo before reaching pluripotency. For this purpose, we focused on the pancreas which we found to be the organ with the highest reprogramming efficiency in vivo. We demonstrate that in vivo reprogramming leads to the loss of pancreatic acinar identity and the acquisition of an atypical tubular morphology different from normal pancreatic ducts or from the metaplastic ducts produced by pancreatitis. Interestingly, the OSKM-induced atypical tubules are characterized by the presence of keratin 14 (KRT14), a protein that it is normally absent from pancreas, except in the context of rare squamous pancreatic cancers. Moreover, these atypical KRT14-positive cells emerge in many different tissues and primary cells upon OSKM activation, with Klf4 overexpression being the driving force in their emergence, at least in fibroblasts. Single cell RNA sequencing of partially reprogrammed pancreas confirms the presence of extensive changes in cell identity both in acinar and ductal cells. Finally, cells derived from partially reprogrammed pancreas efficiently form organoids in vitro with a transcriptional profile that resembles primitive embryonic stages. In a complementary experimental approach, we have investigated the potential of partial reprogramming to rejuvenate tissues. For this, OSKM was transiently induced and then switched off to allow the restoration of normal histology. Interestingly, we found that transient OSKM activation in old mice rejuvenates transcriptomic alterations associated to aging. Moreover, at an epigenetic level, OSKM transient expression reverses the methylation status of some age-associated differentially methylated regions, including promoters and enhancers, highlighting its beneficial window towards rejuvenation. Overall, unraveling the cellular plasticity induced by transient OSKM activation is of great importance considering that a shift in equilibrium could lead either to extensive tissue remodeling and tumor formation, or to rejuvenation