FA-MNP-MMP-9-ASODN复合物构建及其分子靶向研究

FA-MNP-MMP-9-ASODN complex construction and the study of its FA molecular targeting ability

目的:构建叶酸(FA)分子靶向载基质金属蛋白酶9(MMP-9)反义寡核苷酸(ASODN)磁性纳米复合物(FA-MNP-MMP-9-ASODN),研究该复合物对叶酸受体(FR)阳性鼻咽癌的靶向能力及基因转染效率。方法:采用课题组前期制备的FA靶向磁性纳米载体,通过醛基与氨基缩合反应与MMP-9-ASODN偶联,制备FA-MNP-MMP-9-ASODN,采用琼脂糖凝胶电泳分析寡核苷酸与载体的偶联。通过分析荷HNE-1及CNE-2瘤裸鼠的MRI,和铁染色、透射电镜结果研究该2种细胞及瘤块吞噬纳米复合物情况以及观察细胞内FITC分析该载体的基因转染。结果:电泳结果显示ASODN已成功连接在FA靶向磁性纳米载体上。HNE-1细胞能有效摄取该纳米复合物,细胞内见较多FITC,而CNE-2细胞不能摄取该纳米复合物,细胞内未见FITC。HNE-1瘤块也能有效地吞噬该纳米复合物。结论:成功构建FA-MNP-MMP-9-ASODN纳米复合物,具有良好的FA分子靶向性及基因转染效率。

Objective： To construct FA targeted magnetic nanocomplex（FA-MNP-MMP-9-ASODN） loading matrix metalloproteinase 9（MMP-9）antisense oligonucleotide（ASODN） and evaluate its targeting capacity and efficiency of gene transfection to folate receptor（FR） positive NPC.Method： FA-MNP-MMP-9-ASODN was constructed by MMP-9-ASODN coupling with FA-MNP prepared by our research team through the aldehyde-ammonia condensation reaction.To analyze the feasibility of ASODN coupling with nanovector by agarose gel electrophoresis.Two kinds of HNE-1 and CNE-2 cells and implanted tumors phagocytosis of FA-MNP-MMP-9-ASODN were observed by MRI on tumor-bearing nude mice,iron staining and TEM.To analyze gene transfection of the vector by observing FITC in the cell.Result： The electrophoresis results revealed ASODN successfully coupling with FA-MNP.HNE-1 cell can effectively ingest the nanocomposite,with more FITC in the cell,but CNE-2 cell had not uptake for the nanocomposite,with no FITC in the cell.By comparing with CNE-2 tumor,HNE-1 tumor also can efficiently swallow the nanocomposite.Conclusion： FA-MNP-MMP-9-ASODN nanocomplex is constructed successfully with good FA targeting ability and gene transfection.