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目的研究经典Wnt通路的异常活化与食管癌细胞放射抵抗性的关系,探讨食管癌放射抵抗相关基因及其作用机制。方法应用Illumine-6_V3人类全基因组RNA芯片筛选人类食管癌细胞株TE13、KYSE170(亲代株)及其对应的抵抗株TE13R、KYSE170R在6MV的X射线照射前及照射后24 h的基因表达差异。利用Illuminated Beadstudio Application软件对芯片数据进行通路分析,查找可能与放射抵抗相关的信号通路。应用逆转录-聚合酶链反应(RT-PCR)对相关通路基因进行验证。结果食管癌细胞株在照射前及照射后24 h,分别发现有共性表达差异基因460个和397个。经过通路分析发现其中4个差异特别显著的基因分布在Wnt信号通路上,即CTNNB1、LEF1、FRAT1、TCF3基因。RT-PCR结果证实在照射后24 h(以目标基因值/内参值表示表达量),CTNNB1(1.33±0.14)及LEF1(1.16±0.12)在放射抵抗细胞株TE-13R中的表达明显高于亲代株TE-13中CTNNB1(0.86±0.10)及LEF1(0.84±0.34)的表达(P均<0.05)。结论在食管癌放射抵抗形成过程中,经典Wnt通路异常活化可能发挥重要作用。
Objective To investigate the relationship between the abnormal activation of canonical Wnt pathway and esophageal cancer cells’ radioresistance, and to explore the mechanism of esophageal cancer radioresistance related genes. Methods Illumine-6_V3 human whole-genome RNA microarray was used to screen the gene expression differences between human esophageal cancer cell lines TE13 and KYSE170 (parent strain) and their corresponding resistant strains TE13R and KYSE170R before and after 6MV X-ray irradiation and 24 h irradiation. Illumination Beadstudio Application software was used to perform on-chip analysis of the chip data to find possible signal pathways associated with radioresistance. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to verify the related pathway genes. Results There were 460 genes and 397 genes differentially expressed in esophageal cancer cells before irradiation and 24 h after irradiation, respectively. Four of the most notable genes were found in the Wnt signaling pathway, namely CTNNB1, LEF1, FRAT1 and TCF3 genes. The results of RT-PCR confirmed that the expression of CTNNB1 (1.33 ± 0.14) and LEF1 (1.16 ± 0.12) in the radiation-resistant cell line TE-13R at 24 h after irradiation The expression of CTNNB1 (0.86 ± 0.10) and LEF1 (0.84 ± 0.34) in parent strain TE-13 (all P <0.05). Conclusion Abnormal activation of canonical Wnt pathway may play an important role in esophageal cancer radioresistance.