Jiang Lei, Hao Jin-Li, Jin Mu-Lan, Zhang Yun-Gang, Wei Ping
Department of Pathology, Beijing Chao-Yang Hospital, Capital Medical University, Beijing, China E-mail :
Asian Pac J Cancer Prev. 2013;14(10):6115-20. doi: 10.7314/apjcp.2013.14.10.6115.
Some non-small cell lung cancer (NSCLC) tumor cells are insensitive to tumor necrosis factor- related apoptosis-inducing ligand (TRAIL) -based therapy. This study was conducted to examine the effect of embelin on the sensitivity of the A549 NSCLC cell line to TRAIL receptor2 (TRAILR2) monoclonal antibodies and to investigate the potential mechanisms.
A549 cells were treated with embelin, TRAILR2 mAb or a combination of both. Cell viability was measured using ATPlite assay and apoptosis rates were determined by flow cytometry with AnnexinV-FITC and propidium iodide staining, with the expression levels of proteins analyzed by Western blotting.
The cell survival rate of separate treatments with 100 ng/ml TRAILR2 antibody or 25 uM embelin were 81.5±1.57% and 61.7±2.84%, respectively. Their combined use markedly decreased cell viability in A549 cells to 28.1±1.97% (P<0.05). The general caspase inhibitor Z-VAD- FMK could inhibit the embelin-enhanced sensitivity of A549 cells to TRAILR2 mAb (75.97±3.17%)(P<0.05). Both flow cytometry and cell morphological analysis showed that embelin was able to increase TRAIL-induced apoptosis in A549 cells. Combined treatment with embelin and TRAILR2 mAb augmented the activation of initiator caspases and effector caspase. In addition, A549 cells showed increasing levels of TRAILR2 protein and decreasing levels of Bcl-2, survivin and c-FLIP following the treatment with embelin+TRAILR2 mAb.
Embelin could enhance TRAIL-induced apoptosis in A549 cells. The synergistic effect of the combination treatment might be due to modulation of multiple components in the TRAIL receptor-mediated apoptotic signaling pathway, including TRAILR2, XIAP, survivin, Bcl-2 and c-FLIP.
一些非小细胞肺癌（NSCLC）肿瘤细胞对基于肿瘤坏死因子相关凋亡诱导配体（TRAIL）的治疗不敏感。本研究旨在探讨 embelin 对 A549 非小细胞肺癌细胞系对 TRAIL 受体 2（TRAILR2）单克隆抗体敏感性的影响，并研究其潜在机制。
用 embelin、TRAILR2 单克隆抗体或两者联合处理 A549 细胞。使用 ATPlite 检测法测量细胞活力，通过 AnnexinV-FITC 和碘化丙啶染色的流式细胞术测定凋亡率，并用蛋白质印迹法分析蛋白质表达水平。
单独用 100 ng/ml TRAILR2 抗体或 25 μM embelin 处理的细胞存活率分别为 81.5±1.57%和 61.7±2.84%。它们联合使用显著降低了 A549 细胞的活力至 28.1±1.97%（P<0.05）。通用的半胱天冬酶抑制剂 Z-VAD-FMK 可抑制 embelin 增强的 A549 细胞对 TRAILR2 单克隆抗体的敏感性（75.97±3.17%）（P<0.05）。流式细胞术和细胞形态学分析均表明，embelin 能够增加 TRAIL 诱导的 A549 细胞凋亡。embelin 与 TRAILR2 单克隆抗体联合处理增强了起始半胱天冬酶和效应半胱天冬酶的激活。此外，用 embelin+TRAILR2 单克隆抗体处理后，A549 细胞中 TRAILR2 蛋白水平升高，Bcl-2、survivin 和 c-FLIP 水平降低。
Embelin 可增强 TRAIL 诱导的 A549 细胞凋亡。联合治疗的协同效应可能是由于调节了 TRAIL 受体介导的凋亡信号通路中的多个成分，包括 TRAILR2、XIAP、survivin、Bcl-2 和 c-FLIP。
