沉默GOLPH3基因表达逆转人结肠癌细胞顺铂耐药的研究

doi:10.19401/j.cnki.1007-3639.2019.11.004

中国癌症杂志 ›› 2019, Vol. 29 ›› Issue (11): 862-868.doi: 10.19401/j.cnki.1007-3639.2019.11.004

沉默GOLPH3基因表达逆转人结肠癌细胞顺铂耐药的研究

王春晓，邱成志，洪钟时

福建医科大学附属第二医院普外科，福建泉州 362000

出版日期:2019-11-30 发布日期:2019-12-10
作者简介:邱成志 E-mail: qchengzhi@sohu.com
基金资助:
福建省医学创新项目（2017-CXB7）；福建医科大学校教授基金（JS14028）。

Reversal of cisplatin resistance in human colon cancer cells by silencing of GOLPH3 gene expression

WANG Chunxiao, QIU Chengzhi, HONG Zhongshi

Department of General Surgery, the Second Affiliated Hospital of Fujian Medical University, Quanzhou 362000, Fujian Province, China

Published:2019-11-30 Online:2019-12-10
Contact: QIU Chengzhi E-mail: qchengzhi@sohu.com

摘要/Abstract

摘要： 背景与目的：化疗是结肠癌的重要治疗方法之一，其方案常含有铂类药物，而化疗耐药会影响结肠癌疗效和预后，其发生机制与基因异常表达有关。高尔基磷酸化蛋白3（Golgi phosphoprotein 3，GOLPH3）是一个癌基因，在结肠癌组织中存在过表达，可促进结肠癌细胞的增殖，与预后不良相关。目前，GOLPH3基因的高表达与结肠癌对铂类耐药的相关性尚不明确。探讨沉默GOLPH3基因逆转人结肠癌HT29细胞对顺铂的化疗耐药效应和机制。方法：HT29细胞分为5组。① 对照组：人结肠癌HT29细胞；② 转染组：siRNA-GOLPH3转染HT29细胞；③ 实验组1：经顺铂处理的HT29细胞；④ 实验组2：经顺铂处理的siRNA-GOLPH3转染HT29细胞；⑤ 实验组3：经顺铂和细胞外调节蛋白激酶（extracellular signal-regulated protein kinases，ERK）1/2抑制剂PD98059处理的HT29细胞。四甲基偶氮唑蓝（methyl thiazolyl tetrazolium，MTT）法、平板克隆形成实验检测各组结肠癌HT29细胞增殖及克隆形成能力。蛋白质印迹法（Western blot）检测GOLPH3、P-糖蛋白（P-glycoprotein，P-gp）、ERK1/2和pERK1/2蛋白的表达。结果：经顺铂处理后，实验组1、实验组2的细胞在波长490 nm处的吸光度（D）值均显著低于对照组（P<0.05），实验组2的D₄₉₀值显著低于实验组1（P<0.001）；实验组1和实验组2的细胞集落数均显著低于对照组（P<0.01），实验组2的细胞集落数显著低于实验组1（P<0.001）。实验组1的P-gp、GOLPH3、pERK1/2蛋白表达量显著高于实验组2（P<0.01）；实验组3的P-gp蛋白表达量较实验组1显著降低（P<0.01）。结论：沉默GOLPH3基因可通过抑制丝裂原活化蛋白激酶/细胞外信号调节激酶（mitogen-activated protein kinase/extracellular signal-regulated kinase，MAPK/ERK）信号通路逆转HT29结肠癌细胞对顺铂化疗的耐药性。

关键词: 高尔基磷酸化蛋白3, 结肠癌细胞, 丝裂原活化蛋白激酶/细胞外信号调节激酶信号通路, 化疗耐药, 顺铂

Abstract: Background and purpose: Chemotherapy is an important treatment for colon cancer, which often consists of platinum drugs. However, chemotherapy resistance can affect efficacy of drug and prognosis of patient with colon cancer, and its mechanism is related to abnormal gene expression. Golgi phosphoprotein 3 (GOLPH3) is a new oncogene, which is overexpressed in colon cancer tissue. It can promote the proliferation of colon cancer cells, and is associated with poor prognosis. At present, the relationship between the GOLPH3 gene overexpression and the resistance to platinum drugs in colon cancer is still unknown. This study aimed to investigate the effect of GOLPH3 gene silencing on reversing cisplatin resistance and its mechanism in human colon cell HT29. Methods: Colon cancer cells were divided into five groups. ① Control group: HT29 human colon cancer cells; ② Transfection group: GOLPH3 gene silencing of HT29 cells by siRNA; ③ Experimental group 1: HT29 cells treated with cisplatin; ④ Experimental group 2: HT29 siRNA-GOLPH3 transfected cells treated with cisplatin; ⑤ Experimental group 3: HT29 cells treated with cisplatin and extracellular regulated protein kinases (ERK1/2) inhibitor PD98059. The proliferation and colony formation capability of HT29 cell were measured respectively by methyl thiazolyl tetrazolium (MTT) and plate colony formation assays. The expressions of GOLPH3、P-gp、ERK1/2 and pERK1/2 in HT29 cell were detected by Western blot. Results: Absorbance (D) at 490 nm in experimental group 1 and experimental group 2 was significantly lower than that in the control group (P<0.05), while the D₄₉₀ value in experimental group 2 was significantly decreased compared with experimental group 1 (P<0.01). The numbers of colony count in experimental group 1 and experimental group 2 were significantly lower than that in the control group (P<0.01), while the number of colony count in experimental group 2 was significantly decreased compared with experimental group 1 (P<0.001). Compared with experimental group 1, the expressions of GOLPH3, P-gp and pERK1/2 proteins in experimental group 2 were significantly decreased (P<0.01). The expression of P-gp protein in experimental group 3 was significantly lower than that in experimental group 1 (P<0.01). Conclusion: Silencing of GOLPH3 gene could reverse the resistance of human colon cancer HT29 cells to cisplatin by deactivating mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway.

Key words: Golgi phosphoprotein 3, Colon cancer cells, Mitogen-activated protein kinase/extracellular signal-regulated kinase signaling pathway, Chemoresistance, Cisplatin

王春晓, 邱成志, 洪钟时. 沉默GOLPH3基因表达逆转人结肠癌细胞顺铂耐药的研究[J]. 中国癌症杂志, 2019, 29(11): 862-868.

WANG Chunxiao, QIU Chengzhi, HONG Zhongshi. Reversal of cisplatin resistance in human colon cancer cells by silencing of GOLPH3 gene expression[J]. China Oncology, 2019, 29(11): 862-868.

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沉默GOLPH3基因表达逆转人结肠癌细胞顺铂耐药的研究

Reversal of cisplatin resistance in human colon cancer cells by silencing of GOLPH3 gene expression

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