Doxorubicin (DOX) is a widely used antitumor drug that causes severe cardiotoxicity in patients; no effective strategy yet exists to address this problem. We previously reported that 8-formylophiopogonanone B (8-FOB), a natural isoflavone in Ophiopogon japonicas, antagonizes paraquat-induced hepatotoxicity. Here, we explored the mechanisms underlying DOX-induced cardiotoxicity as well as whether 8-FOB can alleviate DOX-induced cardiotoxicity. Acute cardiotoxicity was established by injecting C57BL/6J mice with a single dose of DOX (20 mg/kg, intraperitoneal). To elucidate the mechanisms underlying DOX-induced cardiotoxicity, differentially expressed genes between hearts from DOX-treated and control mice were identified from the Gene Expression Omnibus (GEO) database via GEO2R. Using the Cytoscape software plugin cytoHubba, five hub genes associated with DOX-induced cardiotoxicity were identified: CD68, PTEN, SERPINE1, AIF1, and HMOX1. However, of these, only HMOX1 protein expression levels were significantly increased after DOX treatment. We also confirmed that HMOX1-dependent myocardial inflammation and fibrosis were closely associated with DOX-induced cardiotoxicity. More importantly, 8-FOB protected against DOX-cardiotoxicity by ameliorating cardiac injury and dysfunction, reducing cardiac fibrosis and inflammatory cytokine release, and inhibiting HMOX1 expression. In conclusion, our results suggest that inhibition of HMOX1-dependent myocardial inflammatory insults and fibrosis is essential for 8-FOB to ameliorate DOX-caused cardiotoxicity.