EGFR表达与结肠癌侵袭转移的相关性及其调控作用

Correlation and Regulation between EGFR Expression and Invasion and Metastasis of Colon Cancer

目的探究表皮生长因子受体(epidermal growth factor receptor,EGFR)的表达与结肠癌侵袭转移的相关性及其调控作用。方法收集2017年3月~2019年3月于武汉大学中南医院接受手术治疗的120例结肠癌患者的癌组织及距离癌组织超过2.5 cm处的正常组织,以实时荧光定量聚合酶链式反应(qRT-PCR)检测EGFR mRNA在结肠癌及相应癌旁组织中的表达情况;免疫组化法检测EGFR的蛋白表达;将120例患者临床病理信息资料与EGFR的表达水平进行相关性分析。构建上调EGFR表达的病毒载体rAAV9-EGFR及阴性对照rAAV9-EGFR-NC,转染结肠癌细胞株HCT116;CCK-8及平板克隆实验检测过表达EGFR对HCT116细胞增殖能力的影响;Transwell和划痕实验检测过表达EGFR对HCT116细胞侵袭和迁移能力的影响;Western blot检测各组细胞中EGFR、整合素β6(anti-integrin beta 6,ITGB6)的表达。结果qRT-PCR结果显示,结肠癌组织中EGFR的mRNA表达水平显著高于癌旁正常组织(P<0.05);在收集的120例结肠癌组织中EGFR的表达阳性率为66.7%(80/120),而在癌旁组织中仅为8.3%(10/120),差异有统计学意义(P<0.05);结肠癌患者性别、年龄、肿瘤大小、组织分化程度以及T分期与EGFR的表达水平无显著相关(均P>0.05),而淋巴结转移、浸润深度、临床医学分期与EGFR的阳性表达有相关性(均P<0.01)。构建过表达EGFR的HCT116细胞株rAAV9-EGFR,与空白对照组和阴性对照组rAAV9-EGFR-NC相比,rAAV9-EGFR组细胞增殖、侵袭和迁移能力均增强,差异均具有统计学意义(均P<0.05);rAAV9-EGFR组细胞中EGFR、ITGB6的表达水平明显上升(均P<0.05)。结论EGFR在结肠癌中高表达,过表达EGFR影响结肠癌细胞的增殖、侵袭和迁移能力。

Objective To investigate correlation and regulation between epidermal growth factor receptor(EGFR)expression and invasion and metastasis of colorecal cancer.Methods From May 2017 to March 2019,totally 120 colon cancer patients receiving surgical treatment in our hospital were collected,and cancer tissues and normal tissues more than 2.5 cm away from the cancer tissues were harvested.Real-time quantitative reverse transcription polymerase chain reaction(qRT-PCR)was used to detect the expression of EGFR in colon cancer and corresponding paracancer tissues.Immunohistochemistry was used to detect the expression of EGFR.Correlation analysis was performed between the clinical pathological information of 120 patients and EGFR expression.The viral vector rAAV9-EGFR which up-regulated the protein expression and the negative control rAAV9-EGFR-NC were constructed,and then they were transfected into colon cancer HCT116 cells,and the blank control group was set up.Cell counting kit-8(CCK-8)and plate cloning assay were used to detect the effect of EGFR overexpression on the proliferation of HCT116 cells.Transwell and scratch assay were used to detect the effect of EGFR overexpression on the invasion and migration of HCT116 cells.The expression of EGFR and anti-integrin beta 6(ITGB6)in each group was detected by Western blotting.Results qRT-PCR results showed that the expression level of EGFR mRNA in colon cancer tissues was higher than that in the adjacent normal tissues,and the difference was statistically significant(P<0.05).In the collected 120 colon cancer tissues,the positive rate of EGFR expression was 66.7%(80/120),while it was only 8.3%(10/120)in the adjacent tissues,the difference was statistically significant(P<0.05).There was no significant correlation between gender,age,tumor size,tissue differentiation degree,T stage and EGFR overexpression(P>0.05),while there was statistical correlation between lymph node metastasis,depth of invasion,clinical medical stage and EGFR positive expression(P<0.01).After the transfected colon cancer cell line HCT116 was constructed,compared with the blank control group,the proliferation of the rAAV9-EGFR group increased significantly.The number of cell clones,the number of cells through matrigel increased significantly,and the invasion and migration ability of the cell enhanced,and the differences were statistically significant(all P<0.05).The expression levels of EGFR and ITGB6 in rAAV9-EGFR group increased significantly(P<0.05).Conclusion EGFR is highly expressed in colon cancer,and overexpression of EGFR affects the proliferation,invasion and migration of colon cancer cells.