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目的探讨内皮素3(endothelin 3,EDN3)基因对结直肠癌细胞的增殖、迁移和侵袭能力的影响及其相关机制。方法实时荧光定量PCR检测EDN3 mRNA在人结直肠癌组织及对应癌旁组织中的表达情况,半定量PCR检测EDN3及内皮素受体B(endothelin receptor B,EDNRB)mRNA在结直肠癌细胞(CaCO_2、LoVo、HT-29)中的表达情况。将包装过载体pCDH-CMV-MCS-EF1-Puro-EDN3的病毒感染至结直肠癌细胞LoVo,MTS、克隆形成法、划痕法、Transwell检测EDN3对细胞增殖、克隆、迁移和侵袭能力的改变。Western blot检测LoVo中p-ERK1/2和t-ERK1/2蛋白改变。结果 EDN3在人结直肠癌组织和细胞中表达降低。表达EDN3基因的重组载体成功感染至LoVo细胞。MTS和克隆实验结果显示,实验组和对照组细胞增殖和克隆形成差异无统计学意义(P>0.05),划痕实验和Transwell实验提示实验组细胞迁移和侵袭能力明显低于对照组(P<0.01)。Western blot检测结果显示磷酸化的ERK1/2蛋白表达下降(P<0.01)。结论 EDN3在结直肠癌组织和细胞中表达降低,过表达EDN3能抑制结直肠癌LoVo细胞的迁移和侵袭能力,其机制可能与ERK1/2信号通路有关。
Objective To investigate the effect of endothelin 3 (EDN3) gene on the proliferation, migration and invasion of colorectal cancer cells and its related mechanism. Methods The expression of EDN3 mRNA in human colorectal cancer tissues and corresponding paracancerous tissues was detected by real-time fluorescence quantitative PCR. The mRNA expression levels of EDN3 and EDNRB were detected by semi-quantitative PCR in colorectal cancer cells (CaCO 2 , LoVo, HT-29). The virus infected with pCDH-CMV-MCS-EF1-Puro-EDN3 was infected into colorectal cancer cells LoVo, MTS, clonogenic assay, scratch assay and Transwell assay to detect the effect of EDN3 on cell proliferation, cloning, migration and invasion . Western blot was used to detect the changes of p-ERK1 / 2 and t-ERK1 / 2 in LoVo. Results The expression of EDN3 decreased in human colorectal cancer tissues and cells. The recombinant vector expressing EDN3 gene was successfully infected into LoVo cells. The results of MTS and cloning showed that there was no significant difference in cell proliferation and colony formation between experimental group and control group (P> 0.05). The scratch test and Transwell assay indicated that the ability of cell migration and invasion in experimental group was significantly lower than that in control group (P < 0.01). Western blot results showed that phosphorylated ERK1 / 2 protein decreased (P <0.01). Conclusion The expression of EDN3 is decreased in colorectal cancer tissues and cells. Overexpression of EDN3 can inhibit the migration and invasion of colorectal cancer LoVo cells. The mechanism may be related to the ERK1 / 2 signaling pathway.