Objective To observe the expression of dipeptidyl peptidase 3 (DPP3) and heme oxygenase 1 (HO-1) in cardiac tissues of ovariectomzied (OVX) rats, and to explore the role of DPP3 in 17β-estradiol (E2)-improving stress. Methods OVX operated postmenopausal rats were used as experimental animal models, supplemented with E2 for 4 weeks as the main intervention. The rats were randomly divided into 4 groups according to the presence or absence of OVX and supplementary E2. They were named sham operation control group (Sham)＋Vehicle (Veh) group, Sham＋E2 group, OVX＋Veh group and OVX＋E2 group, with 5 rats in each group. The ratio of heart weight to body weight, mean arterial pressure (MAP), heart rate (HR), maximum rate of rise of left ventricular pressure (dp/dt_max), and the maximum rate of drop of left ventricular pressure (－dp/dt_max) of each group were measured. The levels of oxidative stress in cardiac tissues were detected by enzyme-linked immunosorbent assay (ELISA), and the expression levels of DPP3 and HO-1 were detected by Western blotting. Results Compared with the Sham+Veh group, the ratio of heart weight to body weight, MAP and HR were significantly increased, and the absolute values of maximum rate of rise of left ventricular pressure dp/dt_max and －dp/dt_max were significantly decreased in the OVX＋Veh group (all P<0.05); after E2 supplementary treatment, the ratio of heart weight to body weight, MAP and HR were significantly decreased, and the absolute values of dp/dt_max and －dp/dt_max were significantly increased in the OVX＋E2 group than in the OVX＋Veh group (all P<0.05). Compared with the Sham＋Veh group, the activities of antioxidant enzymes (catalase and superoxide dismutase) and the expression levels of DPP3 and HO-1 were significantly decreased, and the levels of reactive oxygen species and malondialdehyde were significantly increased in the OVX＋Veh group (all P<0.05); after 4 weeks of E2 supplement, the effects induced by OVX were improved (all P<0.05). Conclusion The expression of DPP3 and HO-1 is decreased in cardiac tissues of OVX rats, and can be increased by E2 supplement, showing a cardioprotective effect.