The free radical theory of aging suggests that reactive oxygen species (ROS) are a driving force of the aging process. In many postmitotic tissues with high mitochondrial activity, the mitochondria are considered as main sources of ROS, which arise as by-products of oxidative metabolism. However, in recent years it has been shown that depending on the tissue type also nonmitochondrial enzymes, such as NADPH oxidases and other oxidative enzymes, can contribute significantly to the cellular load of ROS. The emerging picture suggests that various sources of ROS contribute in a different way to aging phenotypes in various tissues. Using senescence of human endothelial cells as a model system, we reported previously that endogenous levels of NADPH oxidase Nox4 are major drivers of human endothelial cell senescence, by inducing a DNA damage response (Lener et al., 2009). We have addressed molecular mechanisms by which Nox4 may induce DNA damage and found an unanticipated link to the mitochondria, which seem to act as important signal amplifiers. We also found that the hydrogen sulphide (H2S)-producing enzyme cystathione beta-synthase (CBS) regulates senescence of human endothelial cells (Albertini et al., 2012); the role of H2S in cellular senescence will be discussed.