Transfection of HEK cells via DNA-loaded PLGA and P(FASA) nanospheres.
Transfection of HEK cells via DNA-loaded PLGA and P(FASA) nanospheres.
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通过负载 DNA 的 PLGA 和 P(FASA) 纳米球转染 HEK 细胞。
DOI：
10.1080/1061186021000038373
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发表时间：
2002
影响因子：
4.5
通讯作者：
Mathiowitz,E
中科院分区：
文献类型：
--
作者：
Sandor,M;Mehta,S;Harris,J;Thanos,C;Weston,P;Marshall,J;Mathiowitz,E
使用各种载体用GFP基因转染HEK细胞：裸DNA、脂质体以及PLGA和P（FASA）质粒负载的纳米球。所有的方法进行了评估，单独和使用氯喹，溶酶体酶抑制剂。使用荧光标准曲线在孵育后的不同时间测定并比较转染效率。无论是单独的裸DNA还是裸DNA和氯喹都不能使细胞凋亡。脂质体与氯喹和单独的脂质体之间没有明显差异，脂质体转染细胞的效率持续增加长达2周。虽然单独用聚合物纳米球转染是不可行的，但加入氯喹允许从细胞内的纳米球释放的DNA逃避内体降解并转染细胞。通过纳米球的转染效率随时间的增加是指数增加的，直到1周，与对于脂质体的推注型施用所观察到的恒定速率相比，表明纳米球通过受控释放机制将DNA递送至细胞。此外，通过纳米球递送至细胞的有效剂量是脂质体的约25%，表明通过PLGA和P（FASA）纳米球的转染实际上可能更有效。
HEK cells were transfected with the GFP gene using various vectors: naked DNA, lipofectamine, and both PLGA and P(FASA) plasmid-loaded nanospheres. All methods were assessed alone and with the use of chloroquine, a lysosomal enzyme inhibitor. Transfection efficiencies were determined and compared at various times post-incubation using a fluorescence standard curve. Neither naked DNA alone nor naked DNA and chloroquine were capable of transfecting cells. No differences were evident between lipofectamine with chloroquine and lipofectamine alone which transfected cells with a constant increase in efficiency up to 2 weeks. While transfection was not feasible with polymeric nanospheres alone, the addition of chloroquine allowed DNA released from nanospheres within cells to escape endosomal degradation and transfect the cells. The increase in transfection efficiency via nanospheres over time was exponential up to 1 week, as compared to the constant rate seen for the bolus-type administration of lipofectamine, indicating that nanospheres delivered DNA to the cells by a controlled release mechanism. Additionally, the effective dose delivered to cells via nanospheres was approximately 25% that of lipofectamine, indicating that transfection via PLGA and P(FASA) nanospheres might actually be more efficient.