Abstract：
The clinical use of the DNA damaging anticancer drug doxorubicin (DOX) is limited by irreversible cardiotoxicity, which depends on the cumulative dose. The RAS-homologous (RHO) small GTPase RAC1 contributes to DOX-induced DNA damage formation and cardiotoxicity. However, the pathophysiological relevance of other RHO GTPases than RAC1 and different cardiac cell types (i.e., cardiomyocytes, non-cardiomyocytes) for DOX-triggered cardiac damage is unclear. Employing diverse in vitro and in vivo models, we comparatively investigated the level of DOX-induced DNA damage in cardiomyocytes versus non-cardiomyocytes (endothelial cells and fibroblasts), in the presence or absence of selected RHO GTPase inhibitors. Non-cardiomyocytes exhibited the highest number of DOX-induced DNA double-strand breaks (DSB), which were efficiently repaired in vitro. By contrast, rather low levels of DSB were formed in cardiomyocytes, which however remained largely unrepaired. Moreover, DOX-induced apoptosis was detected only in non-cardiomyocytes but not in cardiomyocytes. Pharmacological inhibitors of RAC1 and CDC42 most efficiently attenuated DOX-induced DNA damage in all cell types examined in vitro. Consistently, immunohistochemical analyses revealed that the RAC1 inhibitor NSC23766 and the pan-RHO GTPase inhibitor lovastatin reduced the level of DOX-induced residual DNA damage in both cardiomyocytes and non-cardiomyocytes in vivo. Overall, we conclude that endothelial cells, fibroblasts and cardiomyocytes contribute to the pathophysiology of DOX-induced cardiotoxicity, with RAC1- and CDC42-regulated signaling pathways being especially relevant for DOX-stimulated DSB formation and DNA damage response (DDR) activation. Hence, we suggest dual targeting of RAC1/CDC42-dependent mechanisms in multiple cardiac cell types to mitigate DNA damage-dependent cardiac injury evoked by DOX-based anticancer therapy.
摘要：
DNA损伤抗癌药物阿霉素（DOX）的临床使用受到不可逆心脏毒性的限制，这取决于所应用的累积剂量。RAS同源（RHO）小GTP酶RAC1有助于DOX诱导的DNA损伤形成和心脏毒性。然而,比RAC1和不同心脏细胞类型的其他RHOGTP酶的病理生理相关性(即，心肌细胞，非心肌细胞）用于DOX触发的心脏损伤尚不清楚。采用不同的体外和体内模型，我们比较研究了DOX诱导的心肌细胞与非心肌细胞（内皮细胞和成纤维细胞）的DNA损伤水平，在选择的RHOGTP酶抑制剂存在或不存在的情况下。非心肌细胞表现出最高数量的DOX诱导的DNA双链断裂（DSB），在体外有效修复。相比之下,在心肌细胞中形成相当低水平的DSB，然而，这在很大程度上仍未修复。此外，仅在非心肌细胞中检测到DOX诱导的细胞凋亡，而在心肌细胞中未检测到。RAC1和CDC42的药理学抑制剂在体外检查的所有细胞类型中最有效地减弱DOX诱导的DNA损伤。始终如一，免疫组织化学分析显示，RAC1抑制剂NSC23766和pan-RHOGTP酶抑制剂洛伐他汀可降低体内心肌细胞和非心肌细胞中DOX诱导的残留DNA损伤水平。总的来说,我们得出结论，这两种内皮细胞，成纤维细胞和心肌细胞有助于DOX诱导的心脏毒性的病理生理学，RAC1-和CDC42调节的信号通路与DOX刺激的DSB和DNA损伤反应（DDR）激活特别相关。因此，我们建议在多种心脏细胞类型中双重靶向RAC1/CDC42依赖性机制,以减轻基于DOX的抗癌治疗引起的DNA损伤依赖性心脏损伤.
