Using cultured canine cardiac slices to model the autophagic flux with doxorubicin

Chemotherapy-induced impairment of autophagy is implicated in cardiac toxicity induced by anti-cancer drugs. Imperfect translation from rodent models and lack of in vitro models of toxicity has limited investigation of autophagic flux dysregulation, preventing design of novel cardioprotective strategies based on autophagy control. Development of an adult heart tissue culture technique from a translational model will improve investigation of cardiac toxicity. We aimed to optimize a canine cardiac slice culture system for exploration of cancer therapy impact on intact cardiac tissue, creating a translatable model that maintains autophagy in culture and is amenable to autophagy modulation. Canine cardiac tissue slices (350 μm) were generated from left ventricular free wall collected from euthanized client-owned dogs (n=7) free of cardiovascular disease at the Foster Hospital for Small Animals at Tufts University. Cell viability and apoptosis were quantified with MTT assay and TUNEL stainin..., Cell viability was performed with MTT analysis.  Cell apoptosis was analyzed with TUNEL staining.,

化疗诱导的自噬（autophagy）功能受损与抗癌药物引发的心脏毒性密切相关。啮齿类动物模型的转化应用效果欠佳，且缺乏毒性相关体外（in vitro）模型，这一现状限制了对自噬流（autophagic flux）失调的研究，进而阻碍了基于自噬调控的新型心脏保护策略的开发。开发源自转化模型的成年心脏组织培养技术，将有助于推动心脏毒性相关研究。本研究旨在优化犬心脏切片培养体系，以探索癌症治疗对完整心脏组织的影响，构建一种可在培养过程中维持自噬水平、且可进行自噬调控的可转化模型。实验样本取自塔夫茨大学福斯特小动物医院的7只无心血管疾病的客户饲养家犬（n=7），处死后采集其左心室游离壁组织，制备成350 μm厚度的犬心脏组织切片。采用MTT比色法（MTT assay）与末端脱氧核苷酸转移酶介导的dUTP缺口末端标记染色（TUNEL staining）分别定量检测细胞活力与细胞凋亡；其中细胞活力通过MTT分析完成，细胞凋亡则通过TUNEL染色进行分析。