The Effect of Senolytic (BI01) Administration on DNA Methylation Clock Age and the Methylome in Aged and Regenerated Skeletal Muscle

Background: Senescence cells emerge with aging and injury. The contribution of senescent cells to DNA methylation age (DNAmAGE) in vivo is uncertain. Furthermore, stem cell therapy can mediate “rejuvenation”, but how tissue regeneration controlled by resident stem cells affects whole tissue DNAmAGE is unclear. Methods: We assessed DNAmAGE with or without senolytics (BI01) in aged male mice (24-25 months) 35 days following muscle healing (BaCl2-induced regeneration versus non-injured). Young injured mice (5-6 months) without senolytics were comparators.Results: DNAmAGE was decelerated by up to 68% after injury in aged muscle. DNAmAGE was modestly but further significantly decelerated by injury recovery with senolytics. ~1/4 of measured CpGs were altered by injury then recovery regardless of senolytics in aged muscle. Specific methylation changes caused by senolytics included differential regulation of Col, Hdac, Hox, and Wnt genes, which likely contributed to improved regeneration. Altered extracellular matrix remodeling using histological analysis aligned to the methylomic findings with senolytics. Without senolytics, regeneration had a contrasting effect in young mice and tended not to influence or modestly accelerate DNAmAGE. Comparing young to old injury recovery without senolytics using methylome-transcriptome integration, we found a more coordinated molecular profile in young and differential regulation of genes implicated in muscle stem cell performance: Axin2, Egr1, Fzd4, Meg3, and Spry1. Conclusions: Muscle injury and senescent cells affect DNAmAGE and aging influences the transcriptomic-methylomic landscape after resident stem cell-driven tissue reformation. Our data have implications for understanding muscle plasticity with aging and developing therapies aimed at collagen remodeling and senescence. We aimed to profile changes in DNA methylation age (DNAmAGE) and global methylation 35 days after induced chemical injury in the skeletal muscle of young and old C57BL/6J mice. Furthermore, old mice underwent an additional condition in which one group received oral administeration of a senolytic compound and one group served as a vehicle control. To determine DNAmAGE and global methylation, we utilized the Horvath 320k methylation assay. This custom mammalian array combines a prior custom mammalian array with the 285k Illumina Mouse Methylation BeadChip. This was completed on a cohort of 20 old mice (24-25 months) and 5-6 young mice (5-6 months). .