Under physiological circumstances, there is an exquisite balance between reactive oxygen species (ROS) production and ROS degradation, resulting in low steady-state ROS levels. ROS participate in normal cellular function and in cellular homeostasis. Oxidative stress is the state of a transient or a persistent increase of steady-state ROS levels leading to disturbed signaling pathways and oxidative modification of cellular constituents. It is a key pathophysiological player in pathological hypertrophy, pathological remodeling, and the development and progression of heart failure. The heart is the metabolically most active organ and is characterized by the highest content of mitochondria of any tissue. Mitochondria are the main source of ROS in the myocardium. The causal role of oxidative stress in heart failure is highlighted by gene transfer studies of three primary antioxidant enzymes, thioredoxin, and heme oxygenase-1, and is further supported by gene therapy studies directed at correcting oxidative stress linked to metabolic risk factors. Moreover, gene transfer studies have demonstrated that redox-sensitive microRNAs constitute potential therapeutic targets for the treatment of heart failure. In conclusion, gene therapy studies have provided strong corroborative evidence for a key role of oxidative stress in pathological remodeling and in the development of heart failure.