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目的:探讨Rho蛋白GTP酶活化蛋白(Rho GTPase activation protein,RhoGAP)结构域在肝癌缺失基因1(frequentlydeleted in liver,DLC-1)抑制人结肠癌HT29细胞的增殖、侵袭中的作用。方法:脂质体转染DLC-1基因及RhoGAP结构域缺失的ΔDLC-1亚克隆至大肠癌HT29细胞株;应用MTT、平板克隆实验检测细胞增殖的改变;Transwell实验观察细胞侵袭迁移能力的改变;流式细胞术检测细胞周期。结果:转染成功后,全长DLC-1基因转染组(pcDNA3.1-DLC1-HT29)与野生型及空载组相比,细胞增殖能力下降,侵袭能力减弱,细胞周期G1期阻滞并诱导凋亡,而ΔDLC-1亚克隆转染组(pcDNA3.1-ΔDLC1-HT29)对细胞增殖、侵袭及细胞周期均无明显影响。结论:DLC-1基因可能是通过其内在的RhoGAP结构域抑制人结肠癌细胞HT29的增殖和侵袭。
Objective: To investigate the role of Rho GTPase activation protein (RhoGAP) activation domain in the proliferation and invasion of human colon cancer HT29 cells induced by DLC-1. METHODS: The DLC-1 gene and RhoGAP domain-deleted ΔDLC-1 were subcloned into HT29 cell line with lipofectamine 2000. The cell proliferation was detected by MTT assay and plate clone assay. The changes of cell invasion and migration were observed by Transwell assay Flow cytometry was used to detect cell cycle. Results: After the transfection, the full-length DLC-1 gene transfection group (pcDNA3.1-DLC1-HT29) had a decreased ability of cell proliferation, invasion and cell cycle G1 arrest And induced apoptosis. However, the expression of pcDNA3.1-ΔDLC1-HT29 in the sub-cloning group did not significantly affect cell proliferation, invasion and cell cycle. Conclusion: The DLC-1 gene may inhibit the proliferation and invasion of human colon cancer cell HT29 through its intrinsic RhoGAP domain.