Fe_3O_4磁性纳米颗粒/CD/5-FC系统对U251细胞化疗作用

Chemotherapy effect of Fe_3O_4 magnetic nanoparticles/CD/5-FU gene therapy system on U251 cells

目的:建立以Fe3O4磁性纳米颗粒(Fe3O4,MNP)/胞嘧啶脱氨酶(CD)/5-氟胞嘧啶(5-FC)系统为基础的神经系统胶质瘤治疗方法.方法:利用高温分解铁有机物法以2-吡咯烷酮和乙酰丙酮铁为原料制备出Fe3O4 MNP,用γ-氨丙基三乙氧基硅烷对磁性材料进行表面修饰.以Fe3O4 MNP为载体将CD基因转染U251胶质瘤细胞,检测Fe3O4 MNP对质粒pCMVCD的结合与保护能力.通过RT-PCR,Western Blot法和免疫荧光等方法检测CD基因在U251细胞的表达.噻唑蓝(MTT)法检测5-FC化疗后U251-CD细胞生长曲线的变化.结果:制备出粒径为(10±2)nm的Fe3O4 MNP;使用Fe3O4 MNP作为载体将CD基因成功转染U251细胞;脂质体转染组转染率为(39.23±12.12)%,而示Fe3O4 MNP转染组转染率为(67.35±11.19)%,2组相比较差异显著(P<0.01).Fe3O4 MNP作为载体转染的U251-CD细胞CD基因稳定表达,并呈时间相关性增强表达模式;U251-CD细胞加入5-FC后能显著抑制U251细胞生长.结论:Fe3O4 MNP可作为胶质瘤基因治疗的理想载体;Fe3O4 MNP/CD/5-FC系统可作为脑胶质瘤辅助治疗手段之一.

AIM： To evaluate the feasibility of using Fe3O4 magnetic nanoparticles（ Fe3O4 MNP） as cytosine deaminase gene carrier for glioma gene therapy in vitro. METHODS ： Fe3O4 MNP were prepared by thermal decomposition of Fe （acac）3 in 2- pyrrolidone and 3-aminopropyltriethoxy-silane （ APITS ） was used to modify the surface of the nanoparticles. Transfection was determined by delivering CD gene to gliobastoma cell line U251 using Fe3O4 MNP as gene vector. The mRNA and protein expressions of intraeellular CD gene were tested by RT-PCR, Western blotting and hnmunofluorescent staining, respectively. Methyl thiazolyl tetrazolium（MTT） method was used to detect the proliferation of U251 cells in the presence of 5-FC. RESULTS： The diameter of Fe3O4 MNP was（ 10 ±2） nm. Transfection efficiency was （67. 35 ±11. 19）% in Fe3O4 MNP group, higher than （39.23± 12.12） % in liposome group（P 〈0.01 ）. The results of RT-PCR, Western blotting and immunofluorescent staining revealed that the intracellular CD gene levels continuously increased in a time-dependent manner after transfection of U251 cells with Fe3O4 MNP-pCMVCD complex. CONCLUSION： Fe3O4 MNP can be used as one of the ideal gene carriers for CD gene delivery and offers a basis for gene delivery in vivo. Fe3O4 MNP/ CD/5-FC system can serve as brain glioma adjunctive therapy.