ORIGINAL ARTICLE Role of oxidative stress-induced systemic and cavernosal molecular alterations in the progression of diabetic erectile dysfunction Angela CASTELA, 1,2 Pedro GOMES, 1,3 Valentina F. DOMINGUES, 4 Paula PAÍGA, 4 Raquel COSTA, 1 Pedro VENDEIRA 5 and Carla COSTA 1 Departments of 1 Biochemistry (U38/FCT) and 3 Pharmacology and Therapeutics, Center for Medical Research, Faculty of Medicine of the University of Porto, 2 Institute for Molecular and Cell Biology of the University of Porto (IBMC-UP), 4 Requimte, Instituto Superior de Engenharia, Instituto Politécnico do Porto, and 5 Clínica Saúde Atlântica, Clínica Urológica Vendeira, Porto, Portugal Correspondence Carla Costa, Faculty of Medicine of the University of Porto, Department of Biochemistry, Al. Prof Hernâni Monteiro, 4200-319 Porto, Portugal. Tel: +351 225513654 Fax: +351 225513655 Email: carcosta@med.up.pt Received 10 February 2014; revised 13 May 2014; accepted 1 June 2014. doi: 10.1111/1753-0407.12181 Abstract Background: Erectile dysfunction (ED) is a prevalent complication of dia- betes, and oxidative stress is an important feature of diabetic ED. Oxidative stress-induced damage plays a pivotal role in the development of tissue altera- tions. However, the deleterious effects of oxidative stress in the corpus caver- nosum with the progression of diabetes remain unclear. The aim of this study was to evaluate systemic and penile oxidative stress status in the early and late stages of diabetes. Methods: Male Wistar streptozotocin-diabetic rats (and age-matched con- trols) were examined 2 (early) and 8 weeks (late) after the induction of diabe- tes. Systemic oxidative stress was evaluated by urinary H2O2 and the ratio of circulating reduced/oxidized glutathione (GSH/GSSG). Penile oxidative status was assessed by H2O2 production and 3-nitrotyrosine (3-NT) forma- tion. Cavernosal endothelial nitric oxide synthase (eNOS) was analyzed by quantitative immunohistochemistry. Dual immunofluorescence was also per- formed for 3-NT and α-smooth muscle actin (α-SMA) and eNOS–α-SMA. Results: There was a significant increase in urinary H2O2 levels in both diabetic groups. The plasma GSH/GSSG ratio was significantly augmented in late diabetes. In cavernosal tissue, H2O2 production was significantly increased in late diabetes. Reactivity for 3-NT was located predominantly in cavernosal smooth muscle (SM) and was significantly reduced in late diabetes. Quantitative immunohistochemistry revealed a significant decrease in eNOS levels in cavernosal SM and endothelium in late diabetes. Conclusions: The findings indicate that the noxious effects of oxidative stress are more prominent in late diabetes. Increased penile protein oxidative modi- fications and decreased eNOS expression may be responsible for structural and/or functional deregulation, contributing to the progression of diabetes- associated ED. Keywords: 3-nitrotyrosine, diabetes, endothelial nitric oxide synthase, erectile dysfunction, oxidative stress. Significant findings of the study: The noxious effects of oxidative stress are more prominent in advanced diabetes and include an imbalance in systemic oxidative stress mechanisms, as well as increased penile protein oxidative modifications and decreased endothelial nitric oxide synthase expression. What this study adds: Deleterious oxidative events induced by diabetes contribute to erectile dysfunction, which strengthens the relevance of preventative antioxidant therapy. Journal of Diabetes 7 (2015) 393–401 393 © 2014 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Wiley Publishing Asia Pty Ltd