大鼠肝癌细胞照射后非实质细胞影响肝癌细胞基因的变化  

Transformational genes in rat hepatoma cells induced by irradiated nonparenchymal cells

作　　者：周乐源[1] 张勇[2] 姚苏芮 殷媛[3] 李杰[4] 黄朝晖[3] 

机构地区：[1]江南大学附属医院/无锡市第四人民医院肿瘤放疗科,无锡214062 [2]江南大学附属医院/无锡市第四人民医院肝外科,无锡214062 [3]江南大学附属医院/无锡市第四人民医院肿瘤研究所,无锡214062 [4]江南大学附属医院/无锡市第四人民医院介入科,无锡214062

出　　处：《中华实验外科杂志》2017年第7期1095-1098,共4页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金（81301920、81301784）;江苏省自然科学基金（BK20151108）;中国博士后基金（2016M601758）

摘　　要：目的探讨照射后微环境对残存的肝癌细胞转移能力影响及分子机制。方法分离大鼠肝脏非实质细胞（NPCs），贴壁后进行照射，然后收集培养上清液与照射过的大鼠肝癌细胞McA-RH7777-6 Gy共培养。经不同培液上清共培养McA-RH7777-6 Gy细胞得到RH 6 Gy（与未照射NPCs上清液共培养）和RH 6 Gy-R（与照射后NPCs上清液共培养）细胞株。采用基因芯片技术检测RH 6 Gy和RH 6 Gy-R细胞中基因的表达。通过生物信息学对显著变化的基因进行分析，并利用DAVID数据库对这些变化的基因进行功能富集分析（GO-analysis）和信号转导通路富集分析（KEGG-analysis，BIOCARTA-analysis）。进行细胞划痕实验，比较RH 6 Gy细胞和RH 6 Gy-R细胞的运动迁移能力。结果基因芯片检测结果显示，RH 6 Gy-R与RH 6 Gy细胞比较，周期蛋白依赖性激酶2（CDK2）、丝裂原活化蛋白激酶9（MAPK9）、细胞周期蛋白（Cyclin） A、Toll/白细胞介素-1受体相关蛋白（TIRAP）、3-磷酸肌醇依赖性蛋白激酶1/2（PDK1/2）、磷酸肌醇3激酶（PI3K）等15个肿瘤发生发展相关基因的表达都显著高于未照射上清共培养组，且这些表达差异显著的基因多数富集在肿瘤转移相关通路，如腺苷酸活化蛋白激酶（AMPK）信号通路、磷脂酰肌醇信号通路、Ras信号转导通路等。体外划痕实验显示，细胞划痕培养48 h后，RH 6 Gy细胞面积愈合率为（9.00±1.35）%，RH 6 Gy-R细胞面积愈合率为（41.30±6.32）%。结论经照射后的残存的肝癌表现出更强的转移能力，可能与其微环境中的NPCs相关，我们推测照射后的NPCs释放的某些细胞因子促进了肝癌残存细胞转移相关mRNA的表达，从而激活相关癌转移通路。Objective To investigate the influence of irradiated liver microenvironment on irradiated hepatoma metastatic potentials and its possible molecular mechanism.Methods Primary rats’ nonparenchymal cells （NPCs） were separated and cultured, then they were divided into irradiated （6-Gy X-ray） and non-irradiated groups. After irradiation, conditioned media from irradiated （SR） or non-irradiated （SnonR） cultures were collected and added to the hepatoma McA-RH7777 cell line cells which were given single exposure of 6 Gy X-ray. Then, hepatoma cells were continuously passaged （RH 6 Gy and RH 6 Gy-R）. The expression of mRNA of RH 6 Gy and RH 6 Gy-R cells was compared by gene microarray. Subsequent bioinformatic analysis of these target genes was performed by DAVID software （GO-analysis, Kyoto gene and genome encyclopedia （KEGG）-analysis and BIOCARTA-analysis）. In vitro wound scratch assay was conducted to compare the metastatic potentials of RH 6 Gy and RH 6 Gy-R cells.Results The gene microarray test showed that the expression of genes related to metastasis was up-regulated in RH 6 Gy-R cells, such as cyclin dependent kinase 2 （CDK2）, mitogen-activated protein kinase 9 （MAPK9）, Cyclin A, Toll/interleukin-1R domain-containing adaptor protein （TIRAP）, 3-phosphoinositide-dependent protein kinase 1/2 （PDK1/2） and phosphatidylinositol 3 kinase （PI3K）. These mRNAs were enriched in pathways involved in the development and metastasis of cancer, such as adenosine monophosphate-activated protein kinase （AMPK） signaling pathway, phosphatidylinositol signaling system, Ras signaling pathway.In vitro wound scratch assay revealed that the healing rate of RH 6 Gy and RH 6 Gy-R cells was （9.00±1.35）% and （41.30±6.32）% respectively.Conclusion Radiation can increase invasiveness and metastatic potentials of sublethally irradiated hepatoma cells, and co-culture with conditioned media from irradiated NPCs promotes these potentials. We speculated that some cytokines released by NPC

关 键 词：肝非实质细胞 肝癌细胞 转移 功能富集分析 信号转导通路富集分析 

分 类 号：R735.7[医药卫生—肿瘤]