Longitudinal DNA methylation in serially passaged BJ Fibroblasts to investigate the latent aging of cells

As epigenetic clocks have evolved from powerful estimators of chronological aging to predictors of mortality and disease risk, it begs the question of what role DNA methylation plays in the aging process. We hypothesize that while it has the potential to serve as an informative biomarker, DNA methylation could also be a key to understanding the  biology entangled between aging, (de)differentiation, and epigenetic reprogramming. Here we use an unsupervised approach to analyze time associated DNA methylation from both in vivo and in vitro samples to measure an underlying signal that ties these phenomena together. We identify a methylation pattern shared across all three, as well as a signal that tracks aging in tissues but appears refractory to reprogramming, suggesting that aging and reprogramming may not be fully mirrored processes. BJ fibroblasts with and without hTert expression were serially passaged, with samples being frozen every other passage for potential DNA methylation analysis until senescence. Senescent cells were maintained in culture and passaged to check cell counts, lastly being harvested after 200 days of senescence. From the initial vial, two sets of flasks were passaged independently, called Trajectory A and B. Wild-type serially passaged BJ Fibroblasts were done at separate locations from different original vials from ATCC.