Heterogeneity in premature senescence by oxidative stress correlates with differential DNA damage during the cell cycle

The development of cellular senescence both by replication and by oxidative stress is not homogenous in cultured primary human fibroblasts. To investigate whether this is due to the heterogeneity in the susceptibility of DNA in different phases of the cell cycle, we subjected synchronised cells to oxidative stress and examined the extent of DNA damage and its long-term effects on the induction of cellular senescence. Here, we first show marked heterogeneity in DNA damage as detected by markers of double strand breaks caused by oxidative stress in an asynchronous human fibroblast culture. Cell cycle synchronization followed by oxidative stress demonstrated that DNA in S-phase is most susceptible to oxidative stress whereas DNA in the quiescent phase is most resistant. DNA repair is an ongoing process after sensing DNA damage; reparable DNA damage is repaired even in cells that contain persistent DNA damage. The extent of persistent DNA damage is tightly correlated with permanent cessation of DNA replication and SA-β-gal activity. Oxidative stress encountered by cells in S-phase resulted in more persistent DNA damage, more permanent cell cycle arrest and the induction of premature senescence.