Department of Drug Delivery (DDEL), Helmholtz-Institute for Pharmaceutical Research Saarland (HIPS), Helmholtz Center for Infection Research (HZI), Campus E8.1, 66123, Saarbrücken, Germany.
Department of Pharmacy, Saarland University, 66123, Saarbrücken, Germany.
J Nanobiotechnology. 2018 Sep 19;16(1):72. doi: 10.1186/s12951-018-0401-y.
Messenger RNA (mRNA) has gained remarkable attention as an alternative to DNA-based therapies in biomedical research. A variety of biodegradable nanoparticles (NPs) has been developed including lipid-based and polymer-based systems for mRNA delivery. However, both systems still lack in achieving an efficient transfection rate and a detailed understanding of the mRNA transgene expression kinetics. Therefore, quantitative analysis of the time-dependent translation behavior would provide a better understanding of mRNA's transient nature and further aid the enhancement of appropriate carriers with the perspective to generate future precision nanomedicines with quick response to treat various diseases.
A lipid-polymer hybrid system complexed with mRNA was evaluated regarding its efficiency to transfect dendritic cells (DCs) by simultaneous live cell video imaging of both particle uptake and reporter gene expression. We prepared and optimized NPs consisting of poly (lactid-co-glycolid) (PLGA) coated with the cationic lipid 1, 2-di-O-octadecenyl-3-trimethylammonium propane abbreviated as LPNs. An earlier developed polymer-based delivery system (chitosan-PLGA NPs) served for comparison. Both NPs types were complexed with mRNA-mCherry at various ratios. While cellular uptake and toxicity of either NPs was comparable, LPNs showed a significantly higher transfection efficiency of ~ 80% while chitosan-PLGA NPs revealed only ~ 5%. Further kinetic analysis elicited a start of protein translation after 1 h, with a maximum after 4 h and drop of transgene expression after 48 h post-transfection, in agreement with the transient nature of mRNA.
Charge-mediated complexation of mRNA to NPs enables efficient and fast cellular delivery and subsequent protein translation. While cellular uptake of both NP types was comparable, mRNA transgene expression was superior to polymer-based NPs when delivered by lipid-polymer NPs.
信使 RNA（mRNA）作为生物医学研究中基于 DNA 疗法的替代物引起了广泛关注。已经开发出了多种可生物降解的纳米颗粒（NPs），包括基于脂质和聚合物的系统，用于 mRNA 的递送。然而，这两种系统在实现高效转染率和对 mRNA 转基因表达动力学的详细了解方面仍存在不足。因此，对时间依赖性翻译行为进行定量分析将有助于更好地了解 mRNA 的瞬时性质，并进一步有助于增强合适载体，以期生成具有快速响应能力的未来精准纳米药物，以治疗各种疾病。
通过同时对颗粒摄取和报告基因表达进行活细胞视频成像，评估了与 mRNA 复合的脂质-聚合物混合系统转染树突状细胞（DC）的效率。我们制备并优化了由阳离子脂质 1,2-二油酰基-3-三甲铵丙烷（简称 LPN）包被的聚（乳酸-共-羟基乙酸）（PLGA）组成的 NPs。一种较早开发的聚合物基递送系统（壳聚糖-PLGA NPs）用于比较。两种 NPs 类型均与 mRNA-mCherry 以不同比例复合。虽然两种 NPs 的细胞摄取和毒性相当，但 LPN 的转染效率显著更高（80%），而壳聚糖-PLGA NPs 仅为（5%）。进一步的动力学分析表明，在转染后 1 小时开始翻译蛋白质，在 4 小时后达到最大值，在 48 小时后转基因表达下降，与 mRNA 的瞬时性质一致。
通过带电荷的 NPs 与 mRNA 之间的复合，能够实现高效快速的细胞递送和随后的蛋白质翻译。虽然两种 NP 类型的细胞摄取相当，但当通过脂质-聚合物 NPs 递送时，mRNA 转基因表达优于聚合物基 NPs。
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