Establishment of TP-induced BPH rat model

The rats were acclimatized for 1 week prior to randomized division into 6 groups (6 rats/group): the normal control (NC) group, which received an oral administration of PBS and a subcutaneous injection of corn oil; the BPH group, which received an oral administration of PBS and a subcutaneous injection of 3 mg/kg TP; the finasteride (Fina) group, used as a positive control, which received an oral administration of 10 mg/kg Fina and a subcutaneous injection of 3 mg/kg TP; and three HU-033 groups, which received an oral administration of 50, 150, and 300 mg/kg HU-033, respectively, and a subcutaneous injection of 3 mg/kg TP. All rats received the indicated treatments daily for 4 weeks and body weight was measured once per week. At the end of the experiment, the rats were fasted overnight, anesthetized by the intraperitoneal injection of pentobarbital (100 mg/kg), and dissected to obtain blood samples from the caudal vena cava. The whole blood samples were centrifuged at 1,000 g for 10 min to obtain serum, which was stored at −80°C until further analysis. The prostate of each rat was also carefully recovered and weighed. The prostatic index and percentage inhibition were calculated from the following equation:

(Patil et al., 2016). The ventral prostate of each rat was fixed overnight in 10% neutral buffered formalin (NBF) and the rest of the prostate was snap-frozen in liquid nitrogen for protein assays.