摘要
目的以壳聚糖纳米粒作为人类表皮生长因子受体2小干扰RNA(HER-2siRNA)的载体转染人乳腺癌细胞SK-BR-3,观察HER-2沉默对其体外迁移能力的影响。方法采用离子凝胶法制备壳聚糖-HER-2siRNA纳米粒,原子力显微镜观察纳米粒的形态。分4组进行转染,分别加入转染试剂壳聚糖-HER-2siRNA纳米粒(实验组)、LipofectamineTM2000和HER-2siRNA(阳性对照组)、壳聚糖-阴性对照siRNA纳米粒(阴性对照组)和空白壳聚糖纳米粒(空白组)。一步法逆转录-聚合酶链反应(RT-PCR)和Westernblot鉴定HER-2在SK-BR-3细胞中的表达变化。Transwell小室法检测HER-2siRNA对细胞体外迁移能力的影响。统计分析采用单因素方差分析、K-WH秩和检验。结果成功制备了壳聚糖-siRNA纳米粒,呈球形,大小约100nm。实验组能有效抑制HER-2mRNA和蛋白的表达以及SK-BR-3细胞的体外迁移能力,与阴性对照组和空白组相比差异有显著的统计学意义(P均=0.000)。结论壳聚糖纳米粒介导的HER-2siRNA能有效抑制人乳腺癌细胞SK-BR-3中HER-2的表达及其体外迁移能力。
Objective To investigate the effect of chitosan nanoparticles loaded with small interference RNA （siRNA） of HER-2（ human epidermal growth factor receptor 2）on in vitro migratory activity of SK-BR-3 cells from a human breast carcinoma cell line. Methods Chitosan-siRNA nanoparticle targeting HER-2 mRNA was prepared by ionic gelation using tripolyphosphate, and the morphology of the nanopartieles was observed with atomic force microscope. The transfeeted SK-BR-3 cells were divided into 4 groups： the experimental group, which was transfected with ehitosan-HER-2 siRNA nanoparticle, the positive control group, which was transfected with HER-2 siRNA by LipofectamineTM 2000, the negative control group, which was transfected with chitosan-negative control siRNA nanopartiele, and the blank group, which was transfected with blank chitosan- nanopartiele. The expression levels of HER-2 mRNA and protein in SK-BR-3 cells were assessed by RT-PCR （reverse transcription polymerase chain ） and Western blot respectively. Transwell Chamber assay was performed to detect the effect of ehitosan nanoparticles-mediated HER-2 siRNA on the migratory capacity of SK- BR-3 cells. Statistical analysis was performed using one-way analysis of variance or K-W H test. Results The chitosan-siRNA nanoparticles were successfully prepared. The nanoparticles were spherical in shape and well distributed, and the mean diameter was about 100 nm. The expressions of HER-2 mRNA and the protein level were significantly decreased and the migratory ability of SK-BR-3 breast cancer ceils in vitro was also obviously decreased in the experimental group compared with the negative control group and the blank group （P=0.000）. Conclusion HER-2 siRNA transfectd by chitosan nanoparticles can effectively inhibit the expression of HER-2 in SK-BR-3 breast cancer cells and the migration of SK-BR-3 cells in vitro.
出处
《中华乳腺病杂志（电子版）》
CAS
2011年第5期36-40,共5页
Chinese Journal of Breast Disease(Electronic Edition)
基金
广东省科技厅科研基金资助项目(2008B030301033)
