Comparison of the Gene Transfer Efficiency of mRNA/GL67 and pDNA/GL67 Complexes in Respiratory Cells
Comparison of the Gene Transfer Efficiency of mRNA/GL67 and pDNA/GL67 Complexes in Respiratory Cells
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DOI：
10.1021/mp200604h
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发表时间：
2012-08-01
影响因子：
4.9
通讯作者：
Sanders, Niek N.
中科院分区：
文献类型：
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作者：
Andries, Oliwia;De Filette, Marina;Sanders, Niek N.
配制并表征mRNA与GL 67：DOPE：DMPE-PEGS 000（GL 67）脂质体之间的复合物。随后，在呼吸细胞中比较各自以其最佳比例产生的mRNA/GL 67复合物和pDNA/GL 67复合物的体外和体内表达特性。用mRNA/GL 67复合物转染A549细胞比用pDNA/GL 67复合物转染后的表达快得多。mRNA和pDNA转染后GFP阳性细胞的百分比分别在8和24 h后达到峰值。在这些时间点，mRNA转染后GFP阳性细胞的百分比比pDNA转染后高两倍。此外，mRNA/GL 67复合物的功效与细胞周期无关。这与pDNA/GL 67复合物形成鲜明对比，后者在非分裂细胞中仅引起弱表达。这证实了核屏障是pDNA的关键障碍，而不是mRNA的关键障碍。最后，将编码荧光素酶的mRNA/GL 67和pDNA/GL 67复合物鼻内施用至小鼠的肺。mRNA/GL 67复合物在鼠肺中没有引起可测量的荧光素酶表达。相反，在接受pDNA/GL 67复合物的小鼠的肺中存在可检测的生物发光信号。我们发现，mRNA/GL 67复合物在生物液体中具有较低的稳定性。因此，这可能是其体内性能较低的原因。
Complexes between mRNA and GL67:DOPE:DMPE-PEGS000 (GL67) liposomes were formulated and characterized. Subsequently, the in vitro and in vivo expression characteristics of mRNA/GL67 complexes and pDNA/GL67 complexes, each produced at their optimal ratio, were compared in respiratory cells. Transfection of A549 cells with mRNA/GL67 complexes resulted in a much faster expression than after transfection with pDNA/GL67 complexes. The percentage of GFP-positive cells after mRNA and pDNA transfection peaked after 8 and 24 h, respectively. At these time points the percentage of GFP-positive cells was two times higher after mRNA transfection than after pDNA transfection. Furthermore, the efficacy of mRNA/GL67 complexes was independent of the cell cycle. This was in sharp contrast with pDNA/GL67 complexes that caused only a weak expression in nondividing cells. This confirms that the nuclear barrier is a crucial obstacle for pDNA but not for mRNA. Finally, mRNA/GL67 and pDNA/GL67 complexes encoding luciferase were administered intranasally to the lungs of mice. The mRNA/GL67 complexes did not give rise to a measurable luciferase expression in the murine lungs. In contrast, a detectable bioluminescent signal was present in the lungs of mice that received the pDNA/GL67 complexes. We showed that mRNA/GL67 complexes have a lower stability in biological fluids. Consequently, this may be an explanation for their lower performance in vivo.