下调Wnt通路活性对非小细胞肺癌细胞株吉非替尼疗效的影响  

Effect of Wnt signaling suppression on gefitinib in non small cell lung cancer cell lines

作　　者：方霞[1] 顾盼[1] 周彩存[2] 任胜祥[2] 罗本芳 曾郁[1] 吴运瑾[1] 赵印敏 朱旭友[1] 易祥华[1] 

机构地区：[1]同济大学附属上海市同济医院病理科,200065 [2]同济大学附属上海市肺科医院肿瘤科 [3]特检科 [4]中心实验室

出　　处：《中华病理学杂志》2013年第7期455-459,共5页Chinese Journal of Pathology

基　　金：上海市科学技术委员会科研基金（034119868）;上海市科学技术委员会医学重点科研基金（09411951600）

摘　　要：目的探讨下调Wnt通路活性对吉非替尼抑制非小细胞肺癌细胞株增殖的影响及其潜在的作用机制。方法不同浓度吉非替尼处理PC9细胞株（吉非替尼敏感株）与PC9／AB2细胞株（吉非替尼耐药株），细胞计数检测试剂盒（CCK8）检测两种细胞株的增殖情况。Westernblot分别检测PC9与PC9／AB2细胞中Wnt信号通路相关蛋白的表达情况。双荧光素酶报告基因系统（TOPFlash）分别检测PC9和PC9／AB2细胞株Wnt通路的转录活性。采用β-catenin序列特异的siRNA抑制PC9／AB2中Wnt通路的转录活性后，用不同浓度吉非替尼处理，CCK8检测干预后细胞增殖情况。干扰后表皮生长因子受体及其下游通路的表达情况。结果不同浓度吉非替尼作用下，PC9／AB：细胞对吉非替尼的耐药性明显增高（P〈0．05）。PC9／AB2细胞Wnt通路相关蛋白的表达明显高于PC9，差异有统计学意义（t=24．590，P=0．000）。TOPFlash结果显示，PC9／AB2细胞株中Wnt通路的转录活性高于PC9细胞株，差异有统计学意义（t=4．983，P=0．008）。抑制Wnt通路后，与阴性对照组相比，干扰组细胞凋亡率增加，对药物敏感性增强（P〈0．05）。下调Wnt通路活性后p-ERK1／2的表达水平较阴性对照组明显降低，其他蛋白表达无明显差异。结论抑制wnt通路活性可部分逆转非小细胞肺癌细胞对吉非替尼的耐药性。Objective To explore the effect of Wnt signaling suppression on proliferation of non small cell lung cancer to gefitiuib, and its related mechanisms. Methods PC9 and PCg/AB2 cells of both gefitinib sensitive and resistant were treated with different concentrations of gefitinib, and the proliferation index was measured using CCK8 kit. The members of Wnt signaling pathway were detected by Western blot. Dual luciferase reportor gene assay （ TOP Flash） was used to document the transcriptional level of 13-catenitL [3-catenin siRNA was transfected into PC9/AB2 cells to suppress the Wnt signaling transcription, followed by treatment with different concentrations of gefitinib. Western blot was then used to detect the expression of EGFR and its downstream signaling after inhibit the expression of 13-eatenin. Results Treating with different concentrations of gefitinib, the resistance of PC9/AB2 cells to gefitinib was significantly increased （ P 〈 0. 05 ）. The members of Wnt signaling expressed at higher level in PC9/AB2 cells than in PC9 cells （ t = 24. 590, P = O. 000）. TOP Flash examination showed that the endogenous transcriptional activity of Wnt signaling was higher in PC9/AB2 cell than that in PC9 cell （ t = 4. 983, P = 0. 008 ）. Compared with the negative control group, apoptotic rate and sensitivity to gefitinib significantly increased in interfered group （P 〈 0. 05 ）. The expression of p-ERK1/2 significantly decreased after Wnt signaling suppression, although other proteins showed no significant alterations. Conclusion Suppressing the activity of Wnt signaling can partly reverse the celluar risistance to gefitinib in non small cell lung cancer.

关 键 词：癌 非小细胞肺 受体 表皮生长因子 蛋白酪氨酸激酶类 Β连环素 

分 类 号：R734.2[医药卫生—肿瘤]