Lv Runxiao, Du Lili, Lu Chunwen, Wu Jinhui, Ding Muchen, Wang Chao, Mao Ningfang, Shi Zhicai
Department of Rehabilitation Medicine, Shengjing Hospital of China Medical University, Shenyang, Liaoning 110004, P.R. China.
Department of Pathophysiology, College of Basic Medical Science, China Medical University, Shenyang, Liaoning 110122, P.R. China.
Exp Ther Med. 2017 Sep;14(3):2053-2059. doi: 10.3892/etm.2017.4725. Epub 2017 Jul 9.
Allicin is a major bioactive ingredient of garlic and has a broad range of biological activities. Allicin has been reported to protect against cell apoptosis induced by HO in human umbilical vein endothelial cells. The present study evaluated the neuroprotective effect of allicin on the HO-induced apoptosis of rat pheochromocytoma PC12 cells and explored the underlying mechanism involved. PC12 cells were incubated with increasing concentrations of allicin and the toxic effect of allicin was measured by MTT assay. The cells were pretreated for 24 h with low dose (L-), medium dose (M-) and high dose (H-) of allicin, followed by exposure to 200 µM HO for 2 h, and the cell viability was examined by MTT assay. In addition, cell apoptosis rate was analyzed by Annexin V-FITC/PI assay, while intracellular reactive oxygen species (ROS) and mitochondrial transmembrane potential (∆ψm) were measured by flow cytometry. Bcl-2, Bax, cleaved-caspase-3 and cytochrome (Cyt C) in the mitochondria were also examined by western blotting. The results demonstrated that 0.01 µg/ml (L-allicin), 0.1 µg/ml (M-allicin) and 1 µg/ml (H-allicin) were non-toxic doses of allicin. Furthermore, HO reduced cell viability, promoted cell apoptosis, induced ROS production and decreased ∆ψm. However, allicin treatment reversed the effect of HO in a dose-dependent manner. It was also observed that HO exposure significantly decreased Bcl-2 and mitochondrial Cyt C, while it increased Bax and cleaved-caspase-3, which were attenuated by allicin pretreatment. The results revealed that allicin protected PC12 cells from HO-induced cell apoptosis via the mitochondrial pathway, suggesting the potential neuroprotective effect of allicin against neurological diseases.
大蒜素是大蒜的一种主要生物活性成分，具有广泛的生物活性。据报道，大蒜素可保护人脐静脉内皮细胞免受HO诱导的细胞凋亡。本研究评估了大蒜素对HO诱导的大鼠嗜铬细胞瘤PC12细胞凋亡的神经保护作用，并探讨了其潜在机制。用不同浓度的大蒜素孵育PC12细胞，通过MTT法检测大蒜素的毒性作用。细胞分别用低剂量（L-）、中剂量（M-）和高剂量（H-）的大蒜素预处理24小时，然后暴露于200μM HO中2小时，通过MTT法检测细胞活力。此外，通过Annexin V-FITC/PI法分析细胞凋亡率，同时通过流式细胞术检测细胞内活性氧（ROS）和线粒体跨膜电位（∆ψm）。还通过蛋白质印迹法检测线粒体中的Bcl-2、Bax、裂解的caspase-3和细胞色素C（Cyt C）。结果表明，0.01μg/ml（L-大蒜素）、0.1μg/ml（M-大蒜素）和1μg/ml（H-大蒜素）为大蒜素的无毒剂量。此外，HO降低了细胞活力，促进了细胞凋亡，诱导了ROS产生并降低了∆ψm。然而，大蒜素处理以剂量依赖性方式逆转了HO的作用。还观察到，HO暴露显著降低了Bcl-2和线粒体Cyt C，而增加了Bax和裂解的caspase-3，大蒜素预处理可减弱这些变化。结果表明，大蒜素通过线粒体途径保护PC12细胞免受HO诱导的细胞凋亡，提示大蒜素对神经疾病具有潜在的神经保护作用。
