Ureteral obstruction is associated with oxidative stress and fibrosis development of the kidney parenchyma. Apurinic/apyrimidinic endonuclease I (APE1) is an essential DNA repair enzyme for repair of oxidative DNA lesions and regulates several transcription factors. The aim of this study was to investigate whether APE1 is regulated by acute (24 hours) and chronic (7 days) unilateral ureteral obstruction (UUO). APE1 was expressed in essentially all kidney cells with the strongest expression in proximal tubuli. After 24h UUO APE1 mRNA was induced in cortex, inner stripe of outer medulla (ISOM) and inner medulla (IM). In contrast, APE1 protein level was not regulated in IM and ISOM and only slightly increased in cortex. APE1 DNA repair activity was not significantly changed. A different pattern of regulation was observed after 7 days UUO with APE1 mRNA increase in cortex but not in ISOM and IM. APE1 protein level in cortex, ISOM and IM increased significantly. Importantly, we observed a significant increase in APE1 DNA repair activity in cortex and IM. To confirm our model we investigated HO-1, collagen I, fibronectin I, and α-SMA levels. In vitro we found the transcriptional regulatory activity of APE1 to be involved in upregulation of the pro-fibrotic factor connective tissue growth factor (CTGF). In summary, APE1 is regulated at different levels after acute and chronic UUO. Thus, our results suggest that DNA repair activity is regulated in response to progressive (7 days) obstruction and that APE1 potentially could play a role for development of fibrosis in kidney disease.