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目的探讨RNA干扰乙酰肝素酶(HPSE)对人大肠癌细胞增殖、侵袭等生物学行为的影响。方法以人大肠癌细胞株HT-29为研究对象,联合应用生物信息学技术与高效siRNA设计原则进行siRNA分子设计,并通过体外转录与化学合成siRNA,瞬时转染HT-29大肠癌细胞,24、48、72 h后MTT法检测实验组与对照组细胞的增殖活性,RT-PCR检测各组细胞HPSE mRNA表达水平,免疫组化细胞染色检测各组HPSE蛋白表达水平,Transwell体外侵袭实验进行癌细胞体外侵袭力分析。结果实验组与对照组相比,细胞增殖活性明显受到抑制;HPSE mRNA及蛋白表达明显下降;细胞侵袭能力明显下降(均P<0.05)。阴性对照组与空白对照组之间无明显差异(P>0.05)。结论沉默大肠癌细胞HPSE基因,对体外培养的大肠癌细胞的增殖和侵袭有明显的抑制作用,其作用机制与HPSE基因和蛋白表达下调有关。
Objective To investigate the effects of heparanase (HPSE) on the biological behavior of human colorectal cancer cells such as proliferation, invasion and so on. Methods The human colorectal cancer cell line HT-29 was used as the research object. The combination of bioinformatics technology and high-efficiency siRNA design principle was used to design siRNA molecules. Transient transfection of HT-29 colorectal cancer cells by in vitro transcription and chemical synthesis of siRNA 24 , 48,72 h after the MTT assay of experimental group and control group cells proliferation activity, RT-PCR detection of each group HPSE mRNA expression levels, immunohistochemical staining of HPSE protein expression levels in each group, Transwell in vitro invasion experiments for cancer In Vitro Invasion Analysis of Cells. Results Compared with the control group, the cell proliferation activity was significantly inhibited in the experimental group and the expression of HPSE mRNA and protein was significantly decreased. The cell invasion ability was significantly decreased (all P <0.05). There was no significant difference between negative control group and blank control group (P> 0.05). Conclusion The silencing of HPSE gene in colorectal cancer cells significantly inhibits the proliferation and invasion of colorectal cancer cells in vitro, and its mechanism is related to the down-regulation of HPSE gene and protein expression.