纳米羟基磷灰石可介导转染人肺癌A549细胞

Transfection of A549 human lung cancer cells is mediated by nano-hydroxyapatite

目的探讨纳米羟基磷灰石（nHAP）介导带有增强型绿色荧光蛋白（EGFP）基因的pGenesil-1质粒转染人肺癌A549细胞的可行性。方法用均相沉淀法制备nHAP;透射电镜观察纳米粒结构;经超声分散及碳酸钠预处理后,在pH值7.4环境下应用多聚赖氨酸（PLL）修饰nHAP;实验分为nHAP-PLL组、脂质体组及对照组,分别配制转染复合物并转染A549细胞;荧光显微镜观察EGFP的表达;流式细胞仪检测pGenesil-1的转染率。结果透射电镜下nHAP呈针状颗粒,粒径较均匀,约（15-20）nm×（60-80）nm,分散程度良好;nHAP-PLL组和脂质体组均见EGFP表达阳性的A549细胞,对照组未见EGFP表达阳性的A549细胞;nHAP-PLL组pGenesil-1转染A549细胞的转染率为（31.8±1.9）%,与脂质体组的转染率（33.4±3.7）%无差异,对照组未见转染阳性的A549细胞。结论nHAP经PLL表面修饰后可介导人肺癌A549细胞基因转染。

Objective To evaluate whether the expressing vector pGenesil-1 encoding enhanced green fluorescent protein （EGFP） could be transfeeted into A549 human lung cancer cells mediated by nano-hydroxyapatite （nHAP） and to determine the transfeetion efficiency. Methods nHAP was synthesized by the homogeneous precipitation method. The structure of the nanopartieles was observed under transmission electron microscope. The nHAP was prepared using ultrasonieation and Na2CO3 and was modified with poly-L-lysine （PLL） at pH 7.4. The transfection of pGenesil-1 into A549 was divided into three groups as follows ： nHAP-PLL group （ n = 8 ） mediated by nanohydroxyapatite modified with poly-L-lysine （nHAP-PLL）, liposome group （n = 8） mediated by Lipofeetamine and control group （ n = 8 ）. After 48 hours transfection, the expression of EGFP was observed with fluorescent microscope. Flow eytometry was used to detect the transfection rate of each group. Results Under transmission electron microscope, the synthesized product presented needle-like and well dispersed particles with evenly distributed sizes of （15 -20 nm） × （60 -80 nm）. Under fluorescent microscope, the expression of EGFP was detected in A549 cells of nHAP-PLL group and liposome group while no expression of EGFP was seen in A549 cells of control group at 48 hours after transfection. Flow cytometry analyses showed that, the transfection rate of A549 cells was （31.8 ± 1.9） % of nHAP-PLL group and （33.4 ± 3.7） % of liposome group. Compared with the control group, there was significant difference respectively （P 〈0.01 ）. Conclusion The nHAP modified with poly-L-lysine can medi- ate the transfection of plasmid into A549 human lung cancer cells.