Abstract Body: Introduction: Doxorubicin-induced cardiotoxicty (DIC) is a key preclinical model for studying chemotherapy-related cardiac complications. While C57BL/6 mice are widely used, sub-strain dependent differences between C57BL/6J and C57BL/6N susceptibility to DIC remains unexamined. This study addresses this gap by evaluating sub-strain dependent differences to improve preclinical reproducibility and translation.
Methods:In this study, 15-week-old male C57BL/6N (6N) and C57BL/6J (6J) mice were used. Mice from each sub-strain were randomly assigned to DOX or control groups. The DOX group received intraperitoneal injections of 8 mg/kg DOX or an equal volume of sterile saline once weekly for three weeks. Body weights were monitored weekly, followed by echocardiography to assess cardiac function one week after the last dose of DOX. Bulk RNA sequencing identified differentially expressed genes, which were then validated by real-time PCR.
Results: DOX treatment caused higher mortality in 6J (33.3%) than 6N (12.5%) and significant weight loss in 6N but not 6J. Echocardiography showed a significant decline in ejection fraction in 6J mice only, while cardiac output, stroke volume, and cardiac atrophy were affected in both sub-strains. Gene expression analysis revealed significant upregulation of atrial natriuretic peptide in both sub-strains and B-type natriuretic peptide in 6N, and myosin heavy chains (Myh6, Myh7) in both, indicating cardiac remodeling. Transcriptomic analysis identified 518 upregulated and 203 downregulated genes in 6N, and 406 upregulated and 445 downregulated in 6J, with 113 and 115 commonly upregulated and downregulated genes, respectively. Gene set enrichment analysis revealed upregulation of protein secretion, MYC signaling, and heme metabolism in 6N, while inflammatory and developmental pathways were suppressed. In contrast, 6J exhibited upregulation of oxidative phosphorylation, DNA repair, and stress response pathways, with suppression of inflammatory and immune signaling.
Conclusions:The study highlights sub-strain dependent differences in DOX-induced cardiotoxicity, with 6J more susceptible than 6N. Differences in mortality, cardiac function, gene expression, and pathway enrichment, which underscores the impact of genetic background on phenotypic responses to DOX. These findings emphasize the need to consider sub-strain variability to improve the reproducibility and translational relevance of preclinical cardio-oncology models.