脂质体介导S100A4基因反义寡核苷酸对神经母细胞瘤细胞的作用效率及其稳定性的研究

Liposome-mediated transfection efficacy of S100A4 antisense oligodeoxynucleotide and its stability in neuroblastoma cells

摘要:

目的：S100A4基因在神经母细胞瘤早期转移中发挥重要作用。该研究以脂质体介导S100A4基因反义寡核苷酸转染体外培养的神经母细胞瘤细胞,观察其作用效率及稳定性。方法：以脂质体包裹5’端带有FAM荧光标记的S100A4基因反义寡核苷酸转染体外培养的神经母细胞瘤细胞,用无脂质体包裹的反义寡核苷酸为对照,在激光共聚焦显微镜下观察荧光强度变化、作用效率及存留时间。结果：经脂质体介导的S100A4基因反义寡核苷酸可以高效进入神经母细胞瘤细胞,荧光强度转染后8 h达最高值173,24 h后仍高达135,作用效率达68%,稳定表达72 h以上;无脂质体包裹的反义寡核苷酸荧光强度在转染后6 h达最高值90,作用效率35%,稳定表达仅12 h。结论：脂质体介导的S100A4基因反义寡核苷酸作用于神经母细胞瘤细胞效率高、时间持久,值得在神经母细胞瘤基因治疗中进一步探讨。

Abstract:

OBJECTIVE: S100A4 gene plays an important role in neuroblastoma cell invasion and metastasis. This paper aimed to evaluate the liposome-mediated transfection efficacy of S100A4 antisense oligodeoxynucleotide (AS-ODN) and its stability in neuroblastoma cells. METHODS: The fluorescence (FAM) labeled S100A4 AS-ODN was transfected with Lipofectamine~(TM) 2000 into human neuroblastoma cells (LA-N-5). The transfection efficacy, stability and persistence time were observed by laser scanning confocal microscopy, and were compared with those of naked AS-ODN without Lipofectamine~(TM) 2000. RESULTS: The transfection efficacy of the S100A4 AS-ODN mediated by Lipofectamine~(TM) 2000 in neuroblastoma cells was high.The maximal signal intensity of intracellular fluorescence was 173 at 8 hrs after transfection and remained as high as 135 at 24 hrs. The maximal efficacy of transfection was 68%, and the stable expression lasted for more than 72 hrs. The transfection efficacy of the naked AS-ODN without Lipofectamine~(TM) 2000 was 35%. The maximal signal intensity of intracellular fluorescence was 90 and occurred at 6 hrs after transfection. The stable expression lasted for only 12 hrs. CONCLUSIONS: The S100A4 AS-ODN can be effectively transfected with Lipofectamine~(TM) 2000 into neuroblastoma cells, and be stably expressed,which may be useful in gene therapy for neuroblastoma