Objective: To evaluate the expression of Wnt/β-catenin and NF-κB pathway in hypopspadiac male rats treated by utero exposure to di-n-butyl phthalate(DBP). Methods:Twenty pregnant rats were randomly divided into two groups, and treated group was given DBP by gastric intubation at a dose of 800 mg/kg from gestation day(GD) 14 to GD18. On GD19,GT samples of hypospadiac male rats(from DBP treated group)and normal rats(from control group)were collected. Furthermore,Western blot analysis was used to quantify expression of β-catenin,GSK-3β and NF-κB in genital tubercle(GT) of hypospadiac male rats. Results:The body weight and the anogenital distance(AGD) of fetal male rats significantly decreased in DBP treated group. Moreover,the expression of β-catenin was significantly decreased, and an increased expression of GSK-3β and NF-κB in GT was observed(P < 0.05) in hypospadiac male rats. Conclusion:DBP could possibly affect the development of GT by regulating Wnt/β-catenin and NF-κB pathway in fetal male rats.