Downregulation of SNAIL sensitizes hepatocellular carcinoma cells to TRAIL-induced apoptosis by regulating the NF-κB pathway
Downregulation of SNAIL sensitizes hepatocellular carcinoma cells to TRAIL-induced apoptosis by regulating the NF-κB pathway
复制标题
SNAIL 的下调通过调节 NF-kappa B 通路使肝细胞癌细胞对 TRAIL 诱导的细胞凋亡敏感
DOI：
10.3892/or.2015.3743
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发表时间：
2015-03-01
期刊：
影响因子：
4.2
通讯作者：
Liang, Jun
中科院分区：
文献类型：
--
作者：
Wan, Zhaojun;Pan, Huazheng;Liang, Jun
肝细胞癌（HCC）是全球第六大常见癌症和第二大致死性癌症。有证据表明HCC细胞对TRAIL介导的凋亡具有抗性。在先前的研究中，我们证实沉默SNAIL下调HCC细胞的生长。此外，肝癌细胞对TRAIL的耐药机制与核因子-κ B（NF-κ B）的活化有关。因此，假设SNAIL的下调通过调节NF-κ B途径使HCC细胞对TRAIL诱导的凋亡敏感。本研究筛选出最有效的携带SNAIL shRNA的慢病毒载体，并构建了携带TRAIL的腺病毒载体。采用定量PCR和Western blotting检测SNAIL和TRAIL的表达。采用MTT法和Hoechst法检测肝癌细胞的存活率和凋亡情况。为了确定如何在沉默SNAIL后使HCC细胞对TRAIL诱导的细胞凋亡敏感，通过蛋白质印迹分析评估p53。Western blot检测Bcl-xL、cIAP 2、survivin和Raf-1蛋白表达，Hoechst法检测各基因表达抑制后HuH-7细胞凋亡程度，推测SNAIL下调可能是其诱导肝癌细胞凋亡的分子机制之一。沉默SNAIL通过增强所有HCC细胞中对TRAIL的敏感性而导致凋亡增加。此外，p53蛋白在HuH-7细胞中上调。Bcl-xL，cIAP 2，生存素和Raf-1的表达下调后沉默的SNAIL，而任何的蛋白质的下调有助于SNAIL抑制增强肝癌细胞对TRAIL诱导的凋亡的敏感性，除了cIAP 2。结果表明，沉默SNAIL可以通过上调p53蛋白和调节NF-κ B通路相关基因如Bcl-xL、Survivin和Raf-1来敏化TRAIL诱导的肝癌细胞凋亡。
Hepatocellular carcinoma (HCC) is the sixth most common cancer and the second most lethal cancer worldwide. Evidence has shown HCC cell resistance to TRAIL-mediated apoptosis. In a previous study, we verified that silencing SNAIL downregulated the growth of HCC cells. In addition, the mechanism of resistance to TRAIL in HCC cells was connected with the activation of nuclear factor-kappa B (NF-kappa B). Thus, it was hypothesized that the downregultaion of SNAIL sensitizes HCC cells to TRAIL-induced apoptosis by regulating the NF-kappa B pathway. In the present study, the most effective lentiviral vectors carrying shRNA against SNAIL were selected and adenoviral vectors harboring TRAIL were constructed. The expression of SNAIL and TRAIL was detected by quantitative PCR and western blotting. HCC cell viability and apoptosis were assessed using an MTT assay and the Hoechst test. To determine how to sensitize HCC cells to TRAIL-induced apoptosis after silencing SNAIL, p53 was assessed by western blot analysis. We also investigated the expression of Bcl-xL, cIAP2, survivin and Raf-1 protein using western blot analysis and the apoptotic degree of HuH-7 cells was detected using the Hoechst test following the suppression of each gene, which was a possible molecular mechanism to sensitive TRAIL-induced apoptosis through the downregulation of SNAIL in HCC cells. Silencing SNAIL resulted in increased apoptosis by enhancing sensitization to TRAIL in all the HCC cells. Additionally, p53 protein was upregulated in HuH-7 cells. Expression of Bcl-xL, cIAP2, survivin and Raf-1 was downregulated following silencing of SNAIL, while downregulation of any of the proteins contributed to SNAIL suppression enhancing HCC cell sensitivity to TRAIL-induced apoptosis, with the exception of cIAP2. The results demonstrated that silencing SNAIL can sensitize TRAIL-induced apoptosis in HCC cells by upregulating p53 protein and by regulating related genes of the NF-kappa B pathway such as Bcl-xL, survivin and Raf-1.