Objective To investigate the role and underlying mechanism of cathepsin B in myocar-dial injury in mice with diabetic cardiomyopathy(DCM).Methods Twenty 8-week-old male SPF C57BL/6 mice were randomly divided into wild-type(WT)group and WT DCM group,with 10 mice in each group.Another 20 8-week-old male SPF-grade mice with cathepsin B knockout(KO)were randomly and equally assigned to KO group and KO DCM group.HE staining was used to observe morphological changes,Prussian blue staining was employed to detect iron deposition,while immunohistochemical staining with 4-hydroxynonenal(4-HNE)was used to assess lipid peroxidation level in the myocardial tissues.Western blotting was performed to detect the expression of heme oxygenase-1(HO-1),superoxide dismutase 2(SOD2),and nuclear factor E2-related factor 2(Nrf2),while RT-PCR was applied to evaluate the expressions of Nrf-2,HO-1,and phospholipid hydroperoxide glutathione peroxidase 4(GPX4).Results Compared to the WT DCM group,the KO DCM group presented improved cell arrangement in cardiac tissues and sig-nificant reduction in inflammatory cell infiltration.Furthermore,the KO DCM group displayed a significant decrease in iron deposition compared to the WT DCM group.Additionally,the KO DCM group exhibited a significant reduction in 4-HNE expression compared to the WT DCM group.The protein levels of Nrf2,SOD2,and HO-1 were significant increased in the KO DCM group than the WT DCM group(0.68±0.21 vs 0.39±0.13,0.59±0.10 vs 0.28±0.09,1.03±0.10 vs 0.48±0.04,P＜0.05).Moreover,elevated mRNA levels of GPX4,Nrf2 and HO-1 were also observed in the KO DCM group than the WT DCM group(0.65±0.09 vs 0.40±0.10,0.61±0.11 vs 0.34±0.11,0.62±0.12 vs 0.39±0.09,P＜0.05).Conclusion Cathepsin B exacerbates myocardial injury in DCM mice through ferroptosis.