厉周 1黄宗海 1姚航 2俞金龙 1李强 1崔大祥3
作者信息
- 1. 南方医科大学附属珠江医院普通外科,广东省,广州市,510282
- 2. 镇江市第三人民医院普通外科,江苏省镇江市,212003
- 3. 上海交通大学微纳米科学技术研究院生物纳米工程研究室,上海市,200030
- 折叠
摘要
Abstract
BACKGROUND: Conventional viral and non-viral vectors exhibit obvious drawbacks in gene therapy, It is promising to acquire better, safer gene transduction and to enhance the efficacy and safety of gene therapy using nanometer materials as antisense oligonucleotide vector.OBJECTIVE: To investigate the feasibility of survivin antisense oligonucleotide (survivin-asODN) transfecting colorectal cancer cells SW620 mediated by polyamidoamine dendrimer (PAMAM) and the effects on the expression of survivin and the apoptosis of colorectal cancer cells SW620.DESIGN, TIME AND SETTIND: An in vitro experiment regarding tumor gene therapy was performed in the Shanghai Research Institute of Micro/Nanometer Science & Technology of Shanghai Jiao Tong University and Central Laboratory of Zhujiang Hospital of Southern Medical University between August 2007 and May 2008.MATERIALS: Human colorectal cancer cells SW620 were purchased from Shanghai Research institute of Cell Biology, Chinese Academy of Sciences. PAMAM dendrimer was a kind gift from professor Cui Da-xiang, Nanometer Engineering Group, Micronamometer Institute, Shanghai Jiao Tong University. LipofectamineTM 2000 was purchased from Invitrogen, USA. Survivin-asODN was ordered from Shanghai Bioengineering Company, China.METHODS: PAMAM antisense gene complex was prepared by 300 μg/L survivin-asODN and 4.06 μg/L PAMAM. Similarly, cationic liposome antisense gene complex was prepared as control. The shape of the complex was observed by transmission electron microscope, the particle size was determined by laser-scattering particle size analyzer and the zeta potential was measured by zeta potential analyzer. The encapsulation efficiency, DNA loading level, and release velocity in vitro were determined using ultraviolet spectrephotometer and centrifugation technique. Both of the above gene transfection complexes were used to transfect colorectal cancer cells and the transfection efficiency of survivin-asODN in SW620 cells was measured. Survivin protein expression was checked by Western blotting. For two groups of cells, apoptosis was assessed by flow cytometry.MAIN OUTCOME MEASURES: Particle size, zeta potential, DNA loading level, encapsulation efficiency, DNA release rate of cationic liposome-survivin-asODN complex and PAMAM-survivin-asODN complex, as well as survivin expression rate and apoptosis rate after tansfection.RESULTS: The particle size of PAMAM-survMn-asODN complex was smaller (P < 0.01), but the zeta potential was greater (P < 0.05), compared with liposome-survivin-asODN. There was no significant difference between PAMAM and liposome groups in terms of DNA loading rate and transfection efficiency. DNA release lasted for 14 days for PAMAM, but only 5 days for liposome. Transfection efficiency of PAMAM-survivin-asODN in the colorectal cancer cells was significantly higher compared with liposome-survivin-asODN (P < 0.05). After colorectal cancer cell transfection, survivin protein expression was lower, but apoptosis rate was higher, in the PAMAM-survivin-asODN complex than in the liposome-survivin-asODN complex (both P < 0.05).CONCLUSION: PAMAM facilitates delivery of survivin-asODN into colorectal cancer cells SW620. As a result, survivin protein expression was decreased, and apoptosis rate was increased.关键词
聚酰胺树形分子高聚合物/反义寡核苷酸/结直肠癌/Survivin/凋亡分类
医药卫生
