抑制Shh信号通路增加胰腺癌细胞对紫杉醇敏感性的实验研究

Inhibition of Shh signaling pathway increases the sensitivity of pancreatic cancer cells to paclitaxel

目的研究抑制音猬因子(Shh)信号通路对胰腺癌细胞紫杉醇敏感性的影响。方法给予人胰腺癌SW1990细胞0、10 nmol/L、20 nmol/L、40 nmol/L、80 nmol/L、160 nmol/L的紫杉醇刺激,MTT测定细胞增殖情况,计算其半数抑制浓度约为50 nmol/L。用Western blot方法测定0、50 nmol/L紫杉醇处理后细胞中Shh信号通路蛋白Shh、锌指转录因子1(Gli1)、补丁蛋白1(Ptch 1)的表达水平。分别用紫杉醇、Shh信号通路抑制剂cyclopamine处理SW1990细胞,用紫杉醇和cyclopamine共同处理SW1990细胞,MTT测定增殖,克隆实验测定细胞克隆能力,流式细胞术测定细胞凋亡,Western blot测定细胞中剪切的含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved Caspase-3)、Bcl-2相关X蛋白(Bax)、Shh、Gli1、Ptch 1蛋白水平。结果 10 nmol/L、20 nmol/L、40 nmol/L、80 nmol/L、160 nmol/L的紫杉醇抑制SW1990细胞的增殖,50 nmol/L的紫杉醇可以抑制细胞中Shh、Gli1、Ptch 1蛋白表达。紫杉醇和cyclopamine单独处理后的细胞增殖能力和克隆形成能力降低,细胞凋亡增多,细胞中Cleaved Caspase-3、Bax蛋白水平升高。与紫杉醇和cyclopamine单独处理的细胞比较,紫杉醇和cyclopamine共同处理后的细胞增殖和克隆能力下降更多,细胞凋亡率更高,细胞中Cleaved Caspase-3、Bax蛋白水平更高,细胞中Shh、Gli1、Ptch 1蛋白水平更低。结论抑制Shh信号通路能够增加胰腺癌细胞对紫杉醇的敏感性。

Objective Study on the inhibitory effect of Shh signaling pathway on paclitaxel sensitivity in pancreatic cancer cells. Methods SW1990 cells were stimulated by 0 nmol/L, 10 nmol/L, 20 nmol/L, 40 nmol/L, 80 nmol/Land 160 nmol/L of paclitaxel, the proliferation of cells was measured by MTT, it was calculated that the median inhibitory concentration was about 50 nmol/L. The expression level of Shh signaling pathway protein Shh, Gli1 and Ptch 1 in 0 and 50 nmol/L paclitaxel treated cells was detected by Western blot. Paclitaxel and Shh signaling pathway inhibitor cyclopamine were used to treat cells respectively. Combined paclitaxel and cyclopamine co processed SW1990 cells, MTT to determine proliferation, cloning assay was used to determine the cell clone ability, cell apoptosis was measured by flow cytometry, Western blot was used to measure the levels of Cleaved Caspase-3, Bax, Shh, Gli1 and Ptch 1 protein in cells. Results 10 nmol/L, 20 nmol/L, 40 nmol/L, 80 nmol/L, 160 nmol/L paclitaxel inhibited the proliferation of SW1990 cells, paclitaxel from 50 nmol/L inhibited the expression of Shh, Gli1 and Ptch 1 protein in cells. Paclitaxel and cyclopamine alone reduced cell proliferation and colony forming ability, cell apoptosis increased, and the levels of Cleaved Caspase-3 and Bax protein increased, compared with paclitaxel and cyclopamine alone treated cells, paclitaxel combined with cyclopamine could decrease cell proliferation and clone ability more, the rate of cell apoptosis was higher, the level of Cleaved Caspase-3 and Bax protein was higher, the levels of Shh, Gli1 and Ptch 1 protein were lower in the cells. Conclusion Inhibition of Shh signaling pathway increases the sensitivity of pancreatic cancer to paclitaxel.