TRAIL-mediated apoptosis requires NF-κB inhibition and the mitochondrial permeability transition in human hepatoma cells
TRAIL-mediated apoptosis requires NF-κB inhibition and the mitochondrial permeability transition in human hepatoma cells
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DOI：
10.1053/jhep.2002.36942
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发表时间：
2002-12-01
期刊：
影响因子：
13.5
通讯作者：
Brenner, DA
中科院分区：
文献类型：
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作者：
Kim, YS;Schwabe, RF;Brenner, DA
肿瘤坏死因子（TNF）相关的凋亡诱导配体（TRAIL）可诱导多种恶性细胞凋亡。然而，包括人类肝癌在内的多种癌症对 TRAIL 具有耐药性。在这项研究中，我们分析了人肝癌细胞中 TRAIL 诱导的促凋亡和抗凋亡信号通路。核因子 kappa B (NF-kappaB) 被发现是 PLC/PRF/5、HepG2 和 Hep3B 细胞系中关键的 TRAIL 诱导的抗凋亡因子。 TRAIL 诱导的 NF-kappaB 激活先于 IkappaBalpha 激酶 (IKK) 激活和 IkappaBalpha 降解，并且依赖于 TRAF2、NF-kappaB 诱导激酶 (NIK)、IKK1 和 IKK2。因此，腺病毒显性失活 (dn) TRAF2、NIKdn、IKK1dn、IKK2dn 或 IkappaBsr 对 NF-kappaB 的抑制使 PLC/PRF/5 细胞对 rhTRAIL 敏感，导致 48 小时后 40% 至 50% 的细胞死亡，而单独使用 rhTRAIL 的细胞死亡率低于 10%。激动性抗 TRAIL 受体 1 和抗 TRAIL 受体 2 抗体或两者的组合在诱导细胞凋亡方面与 rhTRAIL 同等有效，表明在我们的研究条件下，诱饵受体不会导致对 TRAIL 的抵抗。 TRAIL 介导的细胞凋亡依赖于 FADD、caspase 8 和 3，FADDdn、CrmA 和药理学 caspase 抑制剂预防细胞凋亡的能力证明了这一点。共聚焦显微镜显示，rhTRAIL 加放线菌素 D 5 小时后，线粒体通透性转变 (MPT) 开始，随后细胞色素 c 释放。 MPT 对于 TRAIL 诱导的细胞凋亡至关重要，药理学 MPT 抑制剂能够完全保护 PLC/PRF/5 细胞，这证明了这一点。总之，NF-kappaB 通过 TRAIL 激活的 TRAF2-NIK-IKK 途径阻止 TRAIL 诱导的人肝癌细胞凋亡。 NF-kappaB 的抑制揭示了 TRAIL 诱导的细胞凋亡信号级联反应，涉及 FADD、caspase 8、MPT 和 caspase 3。
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) induces apoptosis in a wide range of malignant cells. However, several cancers, including human hepatoma, are resistant to TRAIL. In this study, we analyzed TRAIL-induced pro- and antiapoptotic signaling pathways in human hepatoma cells. Nuclear factor kappa B (NF-kappaB) was found to be a critical TRAIL-induced antiapoptotic factor in the PLC/PRF/5, HepG2, and Hep3B cell lines. TRAIL-induced NF-kappaB activation was preceded by IkappaBalpha kinase (IKK) activation and IkappaBalpha degradation and depended on TRAF2, NF-kappaB-inducing kinase (NIK), IKK1, and IKK2. Accordingly, inhibition of NF-kappaB by adenoviral dominant negative (dn) TRAF2, NIKdn, IKK1dn, IKK2dn, or IkappaBsr sensitized PLC/PRF/5 cells to rhTRAIL, resulting in 40% to 50% cell death after 48 hours as compared with < 10% with rhTRAIL alone. Agonistic anti-TRAIL receptor 1 and anti-TRAIL receptor 2 antibodies or combinations of both were equally efficient in inducing apoptosis as rhTRAIL, indicating that decoy receptors did not contribute to resistance toward TRAIL under the conditions of our study. TRAIL-mediated apoptosis depended on FADD, caspase 8 and 3 as demonstrated by the ability of FADDdn, CrmA, and pharmacologic caspase inhibitors to prevent apoptosis. Confocal microscopy showed the onset of the mitochondrial permeability transition (MPT) 5 hours after rhTRAIL plus actinomycin D, which was followed by cytochrome c release. The MPT was critical for TRAIL-induced apoptosis as demonstrated by the ability of pharmacologic MPT inhibitors to completely protect PLC/PRF/5 cells. In conclusion, NF-kappaB prevents TRAIL-induced apoptosis in human hepatoma through a TRAIL-activated TRAF2-NIK-IKK pathway. Inhibition of NF-kappaB unmasks a TRAIL-induced apoptotic signaling cascade that involves FADD, caspase 8, the MPT, and caspase 3.