The growth inhibition of hepatocellular and cholangiocellular carcinoma cells by gemcitabine and the roles of extracellular signal-regulated and checkpoint kinases
The growth inhibition of hepatocellular and cholangiocellular carcinoma cells by gemcitabine and the roles of extracellular signal-regulated and checkpoint kinases
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DOI：
10.3892/or_00000084
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发表时间：
2008-10-01
期刊：
影响因子：
4.2
通讯作者：
Murawaki, Yoshikazu
中科院分区：
文献类型：
--
作者：
Matsumoto, Kazuya;Nagahara, Takakazu;Murawaki, Yoshikazu
我们研究了嘧啶类似物吉西他滨对肝细胞癌（HCC）和胆管细胞癌（CCC）细胞的影响。分别采用WST-8法、流式细胞术、Hoechst 33258染色法和Western blotting法检测吉西他滨对肝癌细胞（HepG 2、Hep 3B、HLF和PLC/PRF/5）和CCC细胞（HuCCT-1）生长、细胞周期、核形态和信号分子活性的影响。我们发现，吉西他滨显着抑制肝癌和CCC细胞的生长在剂量和时间依赖性的方式。吉西他滨诱导细胞周期停滞在G1期，但未观察到Sub-G1级分，核形态学未表明诱导凋亡。吉西他滨分别在HCC和CCC细胞中诱导检查点激酶Chk 2和Chk 1的差异活化，吉西他滨在两种细胞类型中激活细胞外信号调节激酶（ERK）1/2。在用MEK抑制剂U 0126预处理细胞后，这些检查点激酶的激活被废除，并且细胞死亡被增强。这些结果表明，吉西他滨通过细胞周期阻滞抑制HCC和CCC细胞的生长，而不发生凋亡，ERK/Chk 1/2信号通路是吉西他滨耐药的部分原因。我们的研究结果揭示了吉西他滨治疗HCC和CCC患者，特别是与MEK抑制剂和Chk 1/2抑制剂联合使用时。
We examined the effects of gemcitabine, a pyrimidine analogue, on hepatocellular carcinoma (HCC) and cholangiocellular carcinoma (CCC) cells. After HCC cells (HepG2, Hep3B, HLF and PLC/PRF/5) and CCC cells (HuCCT-1) were treated with gemcitabine, cellular growth, cell cycle, nuclear morphology and activity of signaling molecules were evaluated by WST-8 assays, flow cytometry analysis, Hoechst 33258 staining and Western blotting, respectively. We found that gemcitabine significantly inhibited the growth of HCC and CCC cells in a dose- and time-dependent manner. Gemcitabine induced cell cycle arrest at the G1 phase, however, the Sub-G1 fraction was not observed and nuclear morphology did not indicate the induction of apoptosis. Gemcitabine induced differential activation of checkpoint kinases, Chk2 and Chk1, in HCC and CCC cells, respectively and gemcitabine activated extracellular signal-regulated kinase (ERK)1/2 in both cell types. After the cells were pretreated with a MEK inhibitor U0126, activations of these checkpoint kinases were abrogated and the cell death was enhanced. These results demonstrate that gemcitabine inhibited the growth of HCC and CCC cells by cell cycle arrest without apoptosis and that the ERK/Chk1/2 signaling pathway was in part responsible for the resistance to gemcitabine. Our findings shed light on treating patients with HCC and CCC by gemcitabine, especially when combined with a MEK inhibitor and Chk1/2 inhibitors.