Bortezomib-induced apoptosis in cultured pancreatic cancer cells is associated with ceramide production
Bortezomib-induced apoptosis in cultured pancreatic cancer cells is associated with ceramide production
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DOI：
10.1007/s00280-013-2318-3
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发表时间：
2014-01-01
影响因子：
3
通讯作者：
Wu, Gao-jue
中科院分区：
文献类型：
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作者：
Gong, Lei;Yang, Bo;Wu, Gao-jue
蛋白酶体抑制剂硼替佐米 (PS-341) 对胰腺癌细胞表现出显着的功效。然而，其根本机制尚未完全了解。在这里，我们测试了神经酰胺的产生是否与硼替佐米的作用有关。使用了两种转化的胰腺癌细胞系（PANC-1和Mia）和原代胰腺癌细胞。通过MTT活力测定和台盼蓝染色分析细胞死亡。通过组蛋白 DNA-ELISA 测定和膜联蛋白 V FACS 分析细胞凋亡。蛋白质印迹用于测试信号蛋白的变化。还测定了硼替佐米处理后的细胞神经酰胺水平。在培养的胰腺癌细胞中，硼替佐米增加细胞神经酰胺的产生，促进细胞凋亡。神经酰胺从头合成酶抑制剂伏马菌素 B1 (F-B1) 抑制硼替佐米诱导的神经酰胺产生和细胞凋亡，而外源添加 C6-神经酰胺则促进硼替佐米诱导的胰腺癌细胞死亡。同时，葡萄糖神经酰胺合成酶抑制剂 1-苯基-2-癸酰氨基-3-吗啉代-1-丙醇 (PDMP) 以及鞘氨醇激酶 1 抑制剂（SKI-II 和 SKI-IV）促进硼替佐米诱导的神经酰胺产生和随后的细胞凋亡。此外，硼替佐米诱导的促凋亡 c-Jun N 末端激酶 (JNK) 激活也与神经酰胺的产生有关。在胰腺癌细胞中，硼替佐米的 JNK 激活被 F-B1 抑制，但被 SKI-II 和 PDMP 增强。最后，C6-神经酰胺、SKI-II 和 PDMP 显着增强了原代培养的胰腺癌细胞中硼替佐米诱导的细胞毒性。我们发现，硼替佐米诱导的细胞凋亡与原代和转化胰腺癌细胞中神经酰胺的产生有关。
The proteasome inhibitor bortezomib (PS-341) has displayed significant efficiency against pancreatic cancer cells. However, the underlying mechanisms are not fully understood. Here, we tested if ceramide production was involved in the bortezomib's effect.Two transformed pancreatic cancer cell lines (PANC-1 and Mia) and the primary pancreatic cancer cells were used. Cell death was analyzed by MTT viability assay and trypan blue staining. Cell apoptosis was analyzed by Histone DNA-ELISA assay and Annexin V FACS. Western blots were used to test signal protein changes. The cellular ceramide level after bortezomib treatment was also determined.In cultured pancreatic cancer cells, bortezomib increased cellular ceramide production to promote cell apoptosis. The ceramide de novo synthase inhibitor fumonisin B1 (F-B1) suppressed bortezomib-induced ceramide production and apoptosis, while exogenously added C6-ceramide facilitated bortezomib-induced pancreatic cancer cell death. Meanwhile, 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), the inhibitor of glucosylceramide synthetase as well as the sphingosine kinase 1 inhibitors (SKI-II and SKI-IV), facilitated bortezomib-induced ceramide production and subsequent cell apoptosis. Further, bortezomib-induced pro-apoptotic c-Jun N-terminal kinase (JNK) activation was also associated with ceramide production. JNK activation by bortezomib was suppressed by F-B1, but was enhanced by SKI-II and PDMP in pancreatic cancer cells. Finally, C6-ceramide, SKI-II, and PDMP dramatically enhanced bortezomib-induced cytotoxicity in primary cultured pancreatic cancer cells.We found that bortezomib-induced apoptosis was associated with ceramide production in primary and transformed pancreatic cancer cells.