95D细胞转移相关基因在原发灶、循环肿瘤细胞和转移灶中的差异表达

Differential expression of metastasis-associated genes in primary tumor,circulating tumor cells,and metastasis of human giant-cell lung cancer cell line 95D

目的利用人巨细胞肺癌高转移株(95D)裸鼠自发转移模型,分析CD9、RHOA、MYL12A、B2M、PTOV1、HSPA1B肿瘤转移相关基因在原发灶、循环血肿瘤细胞(circulating tumor cells,CTCs)和转移灶中的差异表达。方法根据前期实验结果并利用GoSufer软件进行GO(gene ontology)分析,筛选出可能与肿瘤转移相关的基因CD9、RHOA、MYL12A、B2M、PTOV1、HSPA1B。利用稳定表达绿色荧光蛋白(GFP)的人巨细胞肺癌细胞株95D/GFP,建立肺癌裸鼠自发转移模型,用流式细胞分选术(fluorescence-activated cell sorting,FACS)分别纯化原发灶、外周血和转移灶中的肿瘤细胞,利用Real-time PCR技术检测上述基因在上述肿瘤细胞中的相对表达量。结果 CD9、RHOA、MYL12A基因在CTCs中表达较少,其次是肝转移灶,在皮下原发灶中表达较多;B2M基因在皮下原发灶、CTCs和肝转移灶中依次降低;PTOV1基因在CTCs中高表达,其次是皮下原发灶,肝转移灶中表达最少;HSPA1B基因只在原发灶中表达。结论 CD9、RHOA、MYL12A、B2M、HSPA1B基因可能是潜在的肺癌转移抑制基因,而PTOV1可能促进肺癌微转移。

Objective To analyze the differential expression of metastasis-associated genes（CD9,RHOA,MYL12A,B2M,PTOV1,and HSPA1B） in primary tumor,circulating tumor cells（CTCs）,and metastasis in a xenograft nude mouse model of lung cancer with spontaneous metastasis,which is established with highly metastatic human giant-cell lung cancer cell line 95D.Methods Metastasis-associated genes CD9,RHOA,MYL12A,B2M,PTOV1,and HSPA1B were selected according to previous experimental data and gene ontology analysis by GoSufer software.The xenograft nude mouse model of lung cancer was established with 95D cells steadily expressing green fluorescent protein（GFP）.Fluorescence-activated cell sorting was used to purify the tumor cells in subcutaneous primary tumor,peripheral blood,and metastasis.Real-time PCR was used to detect relative expression levels of the above genes in the tumor cells.Results CD9,RHOA,and MYL12A had the lowest expression levels in CTCs,medium expression levels in liver metastasis,and the highest expression levels in primary tumor.PTOV1 had the highest expression level in CTCs,a medium expression level in subcutaneous primary tumor,and the lowest expression level in liver metastasis.B2M had the highest expression level in subcutaneous primary tumor,a medium expression level in CTCs,and the lowest expression level in liver metastasis.HSPA1B was detected only in subcutaneous primary tumor.Conclusion CD9,RHOA,MYL12A,B2M,and HSPA1B may be potential genes inhibiting lung cancer metastasis.Gene PTOV1 may contribute to the micrometastasis of lung cancer.