结直肠癌细胞同期放化疗后残余细胞侵袭转移相关生物学特性的变化及其机制

发布时间：2018-07-27 19:04

【摘要】：[目的]通过体外人结直肠癌细胞同期放化疗模型的建立,研究放化疗后残余癌细胞侵袭转移相关生物学行为的变化趋势,并利用基因芯片筛选与之相关的关键基因,探索调变放化疗后残余癌细胞侵袭转移能力变化的可能。本项目将为进一步揭示新辅助放化疗后残余结直肠癌生物学特性变化及其机制打下基础,为提高结直肠癌的综合治疗疗效提供新思路。[方法]1.建立体外同期放化疗模型:对HCT116和HT29细胞运用4Gy 6MVX-线照射和10 umol/L5-Fu药物同时处理,重复多次,得放化疗后残余细胞株(HCT116CR,HT29CR)。2.采用Transwell迁移实验和Transwell侵袭实验进行HCT116CR和HT29CR迁移能力和侵袭能力的测定。3.利用基因芯片检测同期放化疗残余细胞HCT116CR与其母细胞HCT116N长链非编码RNA (long non-coding RNA,1ncRNA)的差异表达水平,根据芯片结果和查阅文献,筛选关键基因。4.运用小干扰RNA(siRNA)干预放化疗后残余细胞HCT16中关键基因的表达或(和)功能,并运用qRT-PCR进行干扰效果的检测。5.再次利用Transwell迁移实验和Transwell侵袭实验检测siRNA干扰后残余细胞siRNA-HCT1 16CR细胞的迁移能力和侵袭能力。[结果]1. HCT116和HT29细胞经过4次同期放化疗处理后,成功建立体外同期放化疗残余细胞模型:HCT116CR 和 HT29CR。2. Transwell迁移实验和Transwell侵袭实验发现,放化疗后残余肠癌细胞迁移能力和侵袭能力较母细胞明显增强。3.基因芯片检测出与放化疗后残余肿瘤细胞生物学行为变化的1ncRNA表达谱,筛选出的三个关键分子:PVT1,LINC00152和MIR22HG。4. SiRNA技术干扰HCT116CR细胞中关键分子的表达后,siRNA-LINC00152组迁移能力、侵袭能力相比未干扰组明显减弱。[结论]1.同期放化疗后残余结直肠癌细胞侵袭转移能力增强,生物学行为发生恶化。2. LINC00152可能是调控放化疗后残余结直肠癌细胞生物学特性变化的关键的分子之一。
[Abstract]:[objective] to study the changes of biological behavior associated with invasion and metastasis of residual cancer cells after radiotherapy and chemotherapy in vitro, and to screen the key genes by gene chip. To explore the possibility of invasion and metastasis of residual cancer cells after adjuvant radiotherapy and chemotherapy. This project will lay a foundation for further revealing the biological characteristics and mechanism of residual colorectal cancer after neoadjuvant radiotherapy and chemotherapy, and provide a new idea for improving the curative effect of comprehensive therapy for colorectal cancer. [methods] 1. To establish the model of simultaneous radiotherapy and chemotherapy in vitro: HCT116 and HT29 cells were irradiated with 4Gy 6MVX- line and treated with 10 umol/L5-Fu drugs simultaneously and repeated for many times. The residual cell line (HCT116CR-HT29CR). 2 was obtained after radiotherapy and chemotherapy. The migration ability and invasion ability of HCT116CR and HT29CR were measured by Transwell migration test and Transwell invasion test. Gene chip was used to detect the differential expression level of HCT116CR and its mother cell HCT116N. According to the results of microarray and literature review, the key gene was screened. Small interfering RNA (siRNA) was used to interfere with the expression or / or function of key genes in HCT16 of residual cells after radiotherapy and chemotherapy, and qRT-PCR was used to detect the interference effect. Transwell migration assay and Transwell invasion assay were used to detect the migration and invasion ability of siRNA-HCT1 16CR cells after siRNA interference. [result] 1. After four times of simultaneous radiotherapy and chemotherapy, HCT116 and HT29 cells were successfully established in vitro concurrent radiotherapy and chemotherapy residual cell model: HCT116CR and HT29CR.2. Transwell migration assay and Transwell invasion assay showed that the migration and invasion ability of residual intestinal cancer cells after radiotherapy and chemotherapy was significantly higher than that of mother cells. The 1ncRNA expression profiles of biological behavior of residual tumor cells after radiotherapy and chemotherapy were detected by gene chip, and three key molecules: PVT1, LINC00152 and MIR22HG.4were selected. After SiRNA interfered with the expression of key molecules in HCT116CR cells, the migration ability of siRNA-LINC00152 group was significantly decreased than that of non-interference group. [conclusion] 1. After radiotherapy and chemotherapy, the invasion and metastasis of residual colorectal cancer cells increased, and biological behavior deteriorated. 2. 2. LINC00152 may be one of the key molecules to regulate the biological characteristics of residual colorectal cancer cells after radiotherapy and chemotherapy.
【学位授予单位】：昆明医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.34

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