中文摘要：

目的探讨叶酸／聚酰胺-胺（FA／PAMAM）介导的微小RNA-7（miR-7）基因治疗体内胶质瘤的实际效果。方法以络合物FA／PAMAM为基因载体转染miR-7至人脑胶质瘤细胞系U251，荧光下观察转染效率，逆转录定量PCR（qRT-PCR）检测miR-7水平。制备去胸腺小鼠颅内U251胶质瘤模型，于成瘤后第3天在肿瘤原位实施络合物移植，M砒动态观察荷瘤小鼠颅内肿瘤体积及水肿变化，记录小鼠生存期。荷瘤小鼠瘤组织切片行原位凋亡检测，免疫组织化学法检测细胞增殖活性抗原（PCNA）、基质金属蛋白酶2（MMP-2）和MMP-9的表达，Westernblot法检测表皮生长因子受体（ECFR）和丝氨酸／苏氨酸蛋白激酶2（AKT-2）的蛋白表达量。结果与脂质体转染比较，FA／PAMAM对miR-7具有更高的基因转染效率。M砌显示，FA/PAMAh量／miR-7组的肿瘤体积增长速度明显低于脂质体／mia-7组，小鼠生存期延长。对照组的细胞凋亡率为（5．3±0．9）％，脂质体／miR-7组的细胞凋亡率为（11．4±2．4）％，FA／PAMAM／miR-7组的细胞凋亡率为（17．7±3．7）％，FA／PAMAM／miR-7组的细胞凋亡率明显增加。免疫组化结果显示，PCNA、MMP-2和MMP-9在对照组中的阳性率分别为（57．3±7．4）％、（45．4±6．9）％和（55．1±7．3）％，在脂质体／miR-7组中分别为（49．3±5．9）％、（31．7±7．1）％和（39．4±6．4）％，在FA／PAMAM／miR-7组中分别为（34．6±5．4）％、（24．5s4．1）％和（25．4S5．1）％。EC,FR和AKT-2蛋白在对照组中的相对表达强度分别为1．09±0．12和0．62±0．10，在脂质体/miR-7组中分别为0．63±0．11和0．43s0．07，在FA／PMM／面R-7组中分别为0．47s0．09和O．31S0．04。FA／PAMAM／miR-7组中PCNA、MMP-2、MMP-9、EGFR和AKT-2蛋白的表达水平均有不同程度下降（均P〈0．05）。结论FA／PAMAM具有较脂质体更高的基因转染效率，药物作用时间?

英文摘要：

Objective To explore if folic acid/polyamide-amine （FA/PAMAM） enhances the therapeutic effect of miR-Tgene therapy for glioma in vivo. Methods The miR-7 gene was transfected into U251 glioma cells by FA/PAMAM. The efficiency of gene tranafection was assessed by fluorescence microscopy. The miR-7 level was detect by quantitative RT-PCR. Intracranial glioma models were established in thymectomized mice, and FA/PAMAM nanoparticles were transplanted into the tumors in situ 3 days later. The animal survival was recorded and the gross tumor volume and degree of edema were observed by MRI. Apoptesis in the glioma cells and expression of proliferating cell nuclear antigen（PCNA）, matrix metalloproteinases 2 and 9 （MMP-2 and MMP-9 ） were assessed by immunohistochemistry, and EGFR and AKT-2 protein expression was detected by Western blot assay. Resulls Compared with the lipesomes, the FA/PAMAM nanoparticles were more efficient to transfer miR-7 gene into U251 glioma cells, MRI showed that the tumor growth was much slower in the FA/PAMAM/miR-7 group, and the animal survival time was longer. The apoptosis rate was （ 5.3± 0.9） % in the control group, （ 11.4 ±2.4 ） % in the lipesome/miR-7 group, and （17.7 ±3.7）% in the FA/PAMAM/miR-7 group. The immunohi assay showed that the levels of PCNA, MMP-2 and MMP-9 protein in the FA/PAMAM/miR-7 group were （34.6 ± 5.4 ） %, （ 24.5 ±4. 1 ） %, （ 25.4±5.1 ） %, respectively, significantly lower than those in the liposome/miR-7group（49.3±5.9）%, （31.7 ±7.1）% and （39.4 ＋6.4）%, respectively, and those in the control group （57.3 ± 7.4） %, （45.4± 6.9 ） % and （ 55.1± 7.3 ） %, respectively （ all P 〈 0.05 ）. The expressions of EGFR and AKT-2 proteins were 1.09 ＋ 0.12 and 0.62± 0.10 in the control group, 0.63± 0.11 and 0.43 ± 0.07 in the liposome/miR-7 group, and significantly deceased （0.47 ± 0.09 and 0.31 ± 0.04, respectively） in the FA/PAMAM/miR-7 group （ all P 〈 0.05）. Conclusion Compared with the liposo