Hu Liufang, Sun Jianhui, Li Hongmei, Wang Lifang, Wei Yuna, Wang Ying, Zhu Yaying, Huo Hairu, Tan Yuqing
Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China.
Toxicol In Vitro. 2016 Dec;37:97-105. doi: 10.1016/j.tiv.2016.09.002. Epub 2016 Sep 5.
The clinical use of arsenic trioxide (AsO) for treating acute promyelocytic leukemia (APL) is limited due to its severe cardiotoxicity. The possible mechanisms of AsO-induced cardiotoxicity include DNA fragmentation, reactive oxygen species (ROS) generation, cardiac ion channel changes and apoptosis. The present study is designed to investigate the protective effects of imperatorin and sec-O-glucosylhamaudol and to explore their mechanistic involvement in AsO-induced cytotoxicity.
Cell viability assay, Lactate dehydrogenase (LDH) release, Acridine orange/ethidium bromide (AO/EB) double staining, Caspase-3 activity assay, ROS generation, cellular calcium levels, mRNA expression levels by qRT-PCR and protein expression levels by Western blotting were measured in H9c2 cells in combination with AsO and imperatorin or sec-O-glucosylhamaudol.
We observed that H9c2 cells treated with imperatorin or sec-O-glucosylhamaudol were more resistant to AsO-induced cell death. Both imperatorin and sec-O-glucosylhamaudol reduced H9c2 cell apoptosis, but both imperatorin and sec-O-glucosylhamaudol had no effects on Caspase-3 activity and intracellular calcium accumulation. Furthermore, imperatorin was capable of suppressing ROS generation, while sec-O-glucosylhamaudol did not show this effect. Moreover, imperatorin and sec-O-glucosylhamaudol triggered Nrf2 activation, which resulted in upregulation of downstream phase II metabolic enzymes and antioxidant protein/enzyme, probably offering cellular protection to AsO-induced cardiotoxicity via the Nrf2 signal pathway.
Imperatorin and sec-O-glucosylhamaudol can ameliorate AsO-induced cytotoxicity and apoptosis in H9c2 cells, the mechanisms probably related to antioxidation. AsO in combination with imperatorin or sec-O-glucosylhamaudol could be considered as a novel strategy to expand the clinical application of AsO.
三氧化二砷（AsO）用于治疗急性早幼粒细胞白血病（APL）的临床应用因严重的心脏毒性而受限。AsO诱导心脏毒性的可能机制包括DNA片段化、活性氧（ROS）生成、心脏离子通道改变和细胞凋亡。本研究旨在探讨欧前胡素和sec-O-葡萄糖基哈马豆醇的保护作用，并探讨它们在AsO诱导的细胞毒性中的作用机制。
在H9c2细胞中联合使用AsO与欧前胡素或sec-O-葡萄糖基哈马豆醇，检测细胞活力测定、乳酸脱氢酶（LDH）释放、吖啶橙/溴化乙锭（AO/EB）双重染色、半胱天冬酶-3活性测定、ROS生成、细胞内钙水平、通过qRT-PCR检测mRNA表达水平以及通过蛋白质印迹法检测蛋白质表达水平。
我们观察到用欧前胡素或sec-O-葡萄糖基哈马豆醇处理的H9c2细胞对AsO诱导的细胞死亡更具抗性；欧前胡素和sec-O-葡萄糖基哈马豆醇均能减少H9c2细胞凋亡，但对半胱天冬酶-3活性和细胞内钙积累均无影响；此外，欧前胡素能够抑制ROS生成，而sec-O-葡萄糖基哈马豆醇未显示出这种作用；而且，欧前胡素和sec-O-葡萄糖基哈马豆醇触发Nrf2激活，导致下游II期代谢酶和抗氧化蛋白/酶上调，可能通过Nrf2信号通路为AsO诱导的心脏毒性提供细胞保护。
欧前胡素和sec-O-葡萄糖基哈马豆醇可改善AsO诱导的H9c2细胞毒性和凋亡，其机制可能与抗氧化作用有关；AsO与欧前胡素或sec-O-葡萄糖基哈马豆醇联合使用可被视为扩大AsO临床应用的新策略。
