INTRODUCTION: Chronic oxidative stress is one of the major factors playing an important role in radiation-induced normal tissue injury. However, the role of oxidative stress in radiation-induced erectile dysfunction (ED) has not been fully investigated. Aims.  To investigate role of oxidative stress after prostate-confined irradiation in a rat model of radiation-induced ED. METHODS: Fifty-four young adult male rats (10-12 weeks of age) were divided into age-matched sham radiotherapy (RT) and RT groups. Irradiated animals received prostate-confined radiation in a single 20 Gy fraction. MAIN OUTCOME MEASURES: Intracavernous pressure (ICP) measurements with cavernous nerve electrical stimulation were conducted at 2, 4, and 9 weeks following RT. The protein expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits (Nox4 and gp91(phox)), markers of oxidative DNA damage (8-hydroxy-2'-deoxyguanosine [8-OHdG]), lipid peroxidation (4-hydroxynonenal [4HNE]), and inflammatory response including inducible nitric oxide synthase, macrophage activation (ED-1), and nitrotyrosine, and endogenous antioxidant defense by nuclear factor erythroid 2-related factor (Nrf2) were evaluated in irradiated prostate tissue and corpora cavernosa (CC). In addition, we investigated the relationships between results of ICP/mean arterial pressure (MAP) ratios and expression level of oxidative stress markers. RESULTS: In the RT group, hemodynamic functional studies demonstrated a significant time-dependent decrease in ICP. Increased expression of Nox4, gp91(phox), 8-OHdG, and 4HNE were observed in the prostate and CC after RT. Similarly, expressions of inflammatory markers were significantly increased. There was a trend for increased Nrf2 after 4 weeks. ICP/MAP ratio negatively correlated with higher expression level of oxidative markers. CONCLUSION: NADPH oxidase activation and chronic oxidative stress were observed in irradiated prostate tissue and CC, which correlated with lower ICP/MAP ratio. Persistent inflammatory responses were also found in both tissues after RT. These findings suggest that oxidative stress plays a crucial role in the development of radiation-induced ED.