Neuroscience Unit, University Hospital Center of Quebec-Laval University, 2705 Laurier Boulevard, Quebec City, Quebec G1V 4G2, Canada.
Endocrinology and Nephrology Unit, University Hospital Center of Quebec-Laval University, 2705 Laurier Boulevard, Quebec City, Quebec G1V 4G2, Canada.
J Leukoc Biol. 2024 May 29;115(6):1165-1176. doi: 10.1093/jleuko/qiae059.
The effective delivery of synthetic RNA into mononuclear phagocytes is a prerequisite for experimental research and therapeutic development. However, traditional methods are highly ineffective and toxic for these cells. Here, we aimed to optimize a transfection protocol for primary bone marrow-derived phagocytes, specifically dendritic cells and macrophages, using lipid nanoparticles generated by microfluidics. Our results show that a lipid mixture similar to that used in Moderna's COVID-19 messenger RNA vaccine outperforms the others tested. Improved messenger RNA transfection can be achieved by replacing uridine with methylpseudouridine but not methoxyuridine, which interferes with transfection. The addition of diphenyleneiodonium or apocynin can enhance transfection in a cell type-dependent manner without adverse effects, while apolipoprotein E provides no added value. These optimized transfection conditions can also be used for microRNA agonists and antagonists. In sum, this study offers a straightforward, highly efficient, reproducible, and nontoxic protocol to deliver RNA into different primary mononuclear phagocytes in culture.
将合成 RNA 有效递送至单核吞噬细胞是实验研究和治疗开发的前提。然而，传统方法对这些细胞的效率和毒性都很高。在这里，我们旨在使用微流控技术生成的脂质纳米颗粒优化用于原代骨髓来源的吞噬细胞（特别是树突状细胞和巨噬细胞）的转染方案。我们的结果表明，类似于 Moderna 的 COVID-19 信使 RNA 疫苗中使用的脂质混合物的转染效果优于其他测试的脂质混合物。通过用甲基假尿嘧啶核苷替代尿嘧啶而不是干扰转染的甲氧尿嘧啶核苷，可以实现信使 RNA 的转染效果改善。二苯基碘鎓或阿朴肉桂醇可以以细胞类型依赖性的方式增强转染而没有不良影响，而载脂蛋白 E 则没有提供额外的价值。这些优化的转染条件也可用于 microRNA 激动剂和拮抗剂。总之，本研究提供了一种简单、高效、可重复且无毒的方案，可将 RNA 递送至培养中的不同原代单核吞噬细胞。
