Multi-omic rejuvenation of human cells by maturation phase transient reprogramming

Abstract Ageing is the gradual decline in organismal fitness that occurs over time leading to tissue dysfunction and disease. At the cellular level, ageing is associated with reduced function, altered gene expression and a perturbed epigenome. Somatic cell reprogramming, the process of converting somatic cells to induced pluripotent stem cells (iPSCs), can reverse these age-associated changes. However, during iPSC reprogramming somatic cell identity is lost, and can be difficult to reacquire as re-differentiated iPSCs often resemble foetal rather than mature adult cells. Recent work has demonstrated that the epigenome is already rejuvenated by the maturation phase of reprogramming, which suggests full iPSC reprogramming is not required to reverse ageing of somatic cells. Here we have developed the first “maturation phase transient reprogramming” (MPTR) method, where reprogramming factors are expressed until this rejuvenation point followed by withdrawal of their induction. Using dermal fibroblasts from middle age donors, we found that cells reacquire their fibroblast identity following MPTR, possibly as a result of persisting epigenetic memory at enhancers. Excitingly, our method substantially rejuvenated multiple cellular attributes including the transcriptome, which was rejuvenated by around 30 years as measured by a novel transcriptome clock. The epigenome, including H3K9me3 histone methylation levels and the DNA methylation ageing clock, was rejuvenated to a similar extent. The magnitude of rejuvenation instigated by MTPR is substantially greater than that achieved in previous transient reprogramming protocols. MPTR fibroblasts produced youthful levels of collagen proteins, suggesting functional rejuvenation. Overall, our work demonstrates that it is possible to separate rejuvenation from pluripotency reprogramming, which should facilitate the discovery of novel anti-ageing genes and therapies. Highlights We developed a novel method by which human fibroblasts are reprogrammed until the maturation phase of iPSCs and are then returned to fibroblast identity DNA methylation memory in fibroblast enhancers may allow recovery of cell identity when fibroblast gene expression programmes are already extinct Molecular measures of ageing including transcriptome and DNA methylation clocks and H3K9me3 levels reveal robust and substantial rejuvenation Functional rejuvenation of fibroblasts by MPTR is suggested by reacquisition of youthful levels of collagen proteins