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目的:通过建立大肠癌细胞与血管内皮细胞间缝隙连接细胞间通讯(GJIC)模型,检测癌细胞与血管内皮细胞间GJIC以及缝隙连接蛋白Cx43表达的情况,分析其与肿瘤转移的关系。方法:采用细胞培养、免疫组化技术及激光漂白后荧光恢复技术,检测单独培养的内皮细胞间以及共培养的人大肠癌细胞系LoVo细胞、HT29细胞与内皮细胞之间GJIC的状况及Cx43的表达。结果:相邻接触的内皮细胞在激光漂白后出现荧光恢复现象,低转移能力的HT29细胞与内皮细胞共培养组荧光恢复明显减缓;高转移能力的LoVo细胞与内皮细胞共培养组荧光恢复更慢。Cx43免疫组化染色显示,单独培养的内皮细胞的细胞膜呈弥漫强阳性着色;HT29细胞与内皮细胞共培养组阳性细胞数量和着色强度明显低于单独内皮细胞培养组;LoVo细胞与内皮细胞共培养组的细胞几乎无Cx43阳性表达。结论:血管内皮细胞与大肠癌细胞粘附后,细胞间的GJIC减少,Cx43的表达降低,且高转移能力的LoVo细胞与血管内皮细胞间的GJIC减少尤其明显,Cx43的表达显著降低甚至缺失。表明肿瘤细胞与血管内皮细胞粘附后,其间的缝隙连接细胞间通讯将发生改变,从而影响恶性肿瘤的转移。
OBJECTIVE: To detect the expression of GJIC and connexin Cx43 between cancer cells and vascular endothelial cells by establishing gap junctional intercellular communication (GJIC) model between colorectal cancer cells and vascular endothelial cells, and to analyze their relationship with tumor metastasis. Methods: Cell culture, immunohistochemistry and fluorescence recovery after laser bleaching were used to detect the status of GJIC between endothelial cells and co-cultured human colorectal cancer cell line LoVo cells, HT29 cells and endothelial cells, and the relationship between Cx43 expression. Results: Fluorescence recovery was observed in adjacent contact endothelial cells after laser bleaching. Fluorescence recovery of HT29 cells with low metastatic potential and endothelial cells significantly slowed down. Fluorescence recovery of LoVo cells with high metastatic potential was slower than that of endothelial cells . Cx43 immunohistochemical staining showed that the cell membrane of cultured endothelial cells was diffusely strongly positive staining. The number of positive cells and the staining intensity of HT29 cells co-cultured with endothelial cells were significantly lower than those of endothelial cells alone. The LoVo cells were co-cultured with endothelial cells Groups of cells showed almost no Cx43 positive expression. CONCLUSION: GJIC decreased and the expression of Cx43 decreased after adhesion of vascular endothelial cells to colorectal cancer cells. In addition, the GJIC reduction between LoVo cells and vascular endothelial cells with high metastasis was especially obvious, and the expression of Cx43 was significantly decreased or even absent. It indicates that after the adhesion of tumor cells and vascular endothelial cells, the interstitial connections between the cells will change, which will affect the metastasis of malignant tumors.