摘要
目的采用婴儿双歧杆菌为载体,探讨尿嘧啶磷酸核糖转移酶基因(UPRT)对胞嘧啶脱氨酶/5-氟胞嘧啶(CD/5-FC)自杀基因系统抗瘤作用的增强效应。方法构建原核表达质粒pGEX-UPRT,以电穿孔法将该质粒转化入婴儿双歧杆菌,筛选阳性重组菌并鉴定。采用M TT法检测表达的U PRT是否可以与CD产生抗瘤协同作用,并观察细胞形态学改变。结果重组婴儿双歧杆菌可以正确表达UPRT。体外M TT检测显示CD+UPRT组细胞存活率低于对照组(P<0.01),且可使B 16-F 10鼠黑色素瘤细胞对5-FC的杀伤敏感性(IC50=0.015μm o l/mL)提高,是CD组(IC50=0.127μm o l/mL)的8.5倍。形态学观察CD+UPRT组肿瘤细胞显示出明显的损伤性改变,细胞生长受到明显抑制,而UPRT组和CD组变化明显不如前者。结论婴儿双歧杆菌联合转导UPRT基因可明显增强CD/5-FC自杀基因系统对鼠黑色素瘤细胞B 16-F 10的杀伤作用。
Objective This study was conducted by using Bifidobacterium Infantis as a delivery system to transport suicide gene CD and UPRT to tumors in an attempt to assess the UPRT-enhanced antitumor effect of CD/5-FC. Methods The recombinant plasmid pGEX-UPRT was constructed and transfered into Bifidobacterium Infantis by electroporation and then identified. The synergistic antitumor effect of coexpression CD and UPRT was determined by MTT method. And the morphologic changes of B16-F10 cells were observed. Results Recombinant Bifidobacterium Infantis could express UPRT correctly. The suvival rate of cells administrated CD＋ UPRT and 5-FC was significantly lower than that of control （P〈0. 01）, and the 5-FC sensitivity （IC50=0. 015 μmol/mL） exhibited a 8.5-fold increase when compared with that of cells administrated CD alone （IC50= 0. 127 μmol/mL）. The cells treated with CD＋UPRT were remarkably damaged morphologically, and the growth of cells was significantly inhibited as compared with that of other groups. Conclusion Recombinant Bifidobacterium Infantis with UPRT gene can significantly enhance the killing effect of CD/5-FC suicide gene system on melanoma B16-F10 cells of mice.
出处
《四川大学学报（医学版）》
CAS
CSCD
北大核心
2007年第1期27-30,63,共5页
Journal of Sichuan University(Medical Sciences)
基金
四川省应用基础研究基金(04JY029-020-2)资助
关键词
婴儿双歧杆菌
尿嘧啶磷酸核糖转移酶
胞嘧啶脱氨酶
基因治疗
Bifidobacterium Infantis Uracil phosphoribosyltransferase （UPRT） Cytosine deaminase （CD） Gene therapy
