Silibinin sensitizes TRAIL-mediated apoptosis by upregulating DR5 through ROS-induced endoplasmic reticulum stress-Ca(2+)-CaMKII-Sp1 pathway.
Silibinin sensitizes TRAIL-mediated apoptosis by upregulating DR5 through ROS-induced endoplasmic reticulum stress-Ca(2+)-CaMKII-Sp1 pathway.
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DOI：
10.18632/oncotarget.23129
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发表时间：
2018-02-13
期刊：
影响因子：
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通讯作者：
Choi YH
中科院分区：
文献类型：
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作者：
Dilshara MG;Jayasooriya RGPT;Molagoda IMN;Jeong JW;Lee S;Park SR;Kim GY;Choi YH
在这项研究中，我们解决了如何水飞蓟宾增强肿瘤坏死因子相关凋亡诱导配体（TRAIL）介导的各种癌细胞凋亡。水飞蓟宾和TRAIL联合处理（silibinin/TRAIL）诱导凋亡，伴随着caspase-3、caspase-8、caspase-9和Bax的活化以及细胞色素c的胞浆积累。抗凋亡蛋白如Bcl-2、IAP-1和IAP-2也被抑制。水飞蓟宾还通过上调死亡受体5（DR 5）触发TRAIL诱导的A549细胞凋亡。DR 5/Fc嵌合蛋白和DR 5靶向的小干扰RNA（siRNA）预处理可显著阻断水飞蓟宾/TRAIL介导的A549细胞凋亡。此外，水飞蓟宾增加了活性氧（ROS）的产生，这导致通过DR 5上调诱导TRAIL介导的凋亡。抗氧化剂如N-乙酰-L-半胱氨酸和谷胱甘肽逆转了TRAIL的凋亡诱导作用。水飞蓟宾进一步诱导内质网（ER）应激，如ER标记蛋白（如PERK、eIF 2 α和ATF-4）的增加所示，这些蛋白刺激CCAAT/增强子结合蛋白同源蛋白（CHOP）的表达。CHOP靶向siRNA消除了DR 5的诱导，并导致水飞蓟宾/TRAIL介导的细胞凋亡显著减少。我们还发现，水飞蓟宾/TRAIL诱导的细胞凋亡伴随着细胞内Ca 2+内流，这是由ER应激和Ca 2+螯合剂，乙二醇四乙酸（EGTA）刺激。Ca 2 +/钙调蛋白依赖性蛋白激酶（CaMK II）抑制剂K252 a阻断水飞蓟宾/TRAIL诱导的DR 5表达以及沿着TRAIL介导的凋亡。因此，我们发现ROS/ER应激诱导的CaMKII激活了Sp1，Sp1是DR 5表达的重要转录因子。我们的研究结果表明，水飞蓟宾增强TRAIL诱导的细胞凋亡，上调DR 5的表达，通过ROS-ER应激-CaMK Ⅱ-Sp1轴。
In this study, we addressed how silibinin enhances tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis in various cancer cells. Combined treatment with silibinin and TRAIL (silibinin/TRAIL) induced apoptosis accompanied by the activation of caspase-3, caspase-8, caspase-9, and Bax, and cytosolic accumulation of cytochrome c. Anti-apoptotic proteins such as Bcl-2, IAP-1, and IAP-2 were inhibited as well. Silibinin also triggered TRAIL-induced apoptosis in A549 cells through upregulation of death receptor 5 (DR5). Pretreatment with DR5/Fc chimeric protein and DR5-targeted small interfering RNA (siRNA) significantly blocked silibinin/TRAIL-mediated apoptosis in A549 cells. Furthermore, silibinin increased the production of reactive oxygen species (ROS), which led to the induction of TRAIL-mediated apoptosis through DR5 upregulation. Antioxidants such as N-acetyl-L-cysteine and glutathione reversed the apoptosis-inducing effects of TRAIL. Silibinin further induced endoplasmic reticulum (ER) stress as was indicated by the increase in ER marker proteins such as PERK, eIF2α, and ATF-4, which stimulate the expression of CCAAT/enhancer binding protein homologous protein (CHOP). CHOP-targeted siRNA eliminated the induction of DR5 and resulted in a significant decrease in silibinin/TRAIL-mediated apoptosis. We also found that silibinin/TRAIL-induced apoptosis was accompanied with intracellular influx of Ca2+, which was stimulated by ER stress and the Ca2+ chelator, ethylene glycol tetraacetic acid (EGTA). Ca2+/calmodulin-dependent protein kinase (CaMKII) inhibitor, K252a, blocked silibinin/TRAIL-induced DR5 expression along with TRAIL-mediated apoptosis. Accordingly, we showed that ROS/ER stress-induced CaMKII activated Sp1, which is an important transcription factor for DR5 expression. Our results showed that silibinin enhanced TRAIL-induced apoptosis by upregulating DR5 expression through the ROS-ER stress-CaMKII-Sp1 axis.