Psychosocial stress exacerbates doxorubicin-induced cardiotoxicity in adult C57BL/6N mice

Psychosocial stress is an established cardiovascular risk factor, yet its influence on chemotherapy-induced cardiotoxicity remains poorly understood. Doxorubicin (DOX), a widely used chemotherapeutic agent, is known to induce cardiotoxicity. However, whether concurrent psychosocial stress exacerbates this effect is unclear. This study aimed to determine the impact of chronic subordination stress (CSS) on DOX-induced cardiotoxicity using a clinically relevant ‘two-hit’ mouse model. Twelve-week-old male C57BL/6N mice were subjected to CSS for 26 days. DOX (8 mg/kg/week) or vehicle was administered during the final 3 weeks of CSS. Cardiac function was evaluated using echocardiography, while myocardial fibrosis was assessed histologically. Bulk RNA sequencing was conducted to identify differentially expressed genes (DEGs), with key genes validated by real-time PCR. Neither CSS nor DOX alone induced significant cardiac dysfunction. However, the combination of CSS and DOX led to both systolic and diastolic dysfunction, myocardial fibrosis, and increased mortality. Expression of cardiac stress markers Nppa and Nppb was significantly elevated by DOX, with CSS further amplifying Nppa expression. RNA sequencing revealed upregulation of pro-fibrotic genes ( Lgals3, Sprr1a ) and the pro-inflammatory cytokine Il6 under combined CSS and DOX exposure. Gene set enrichment analysis showed dysregulation in metabolic, inflammatory, and cell cycle-related pathways. Psychosocial stress significantly worsens DOX-induced cardiotoxicity by promoting cardiac dysfunction, fibrosis, and maladaptive gene expression. This study highlights psychosocial stress as a critical risk factor for adverse cardiovascular outcomes in cancer patients receiving potentially cardiotoxic chemotherapy.