College of Pharmacy, State Key Laboratory of Medicinal Chemical Biology, Nankai University Tianjin 300071, China.
NSF Nanoscale Science and Engineering Center for Affordable Nanoengineering of Polymeric Biomedical Devices (NSEC-CANPBD) The Ohio State University Columbus, Ohio 43210, USA.
Pharm Res. 2012 Jun;29(6):1627-1636. doi: 10.1007/s11095-012-0676-x. Epub 2012 Jan 25.
Delivery of siRNA into cells remains a critical challenge. Our lab has shown a novel polyamidoamine (PAMAM) dendrimer with modified pentaerythritol derivative core (PD dendrimer) to exhibit high plasmid DNA transfection efficiency and low cytotoxicity. Here, we evaluate PD dendrimer as a siRNA carrier.
Agarose gel electrophoresis and AFM were used to confirm formation of generation 5 (G5)-PD dendrimer/siRNA nanoparticles (NPs). G5 PD dendrimer/anti-luciferase siRNA NPs were used to transfect SK Hep-1 cells with stable luciferase expression. Effects of various endocytic pathway inhibitors on uptake of G5 PD dendrimer/siRNA NPs in SK Hep-1 cells were also investigated.
Agarose gel electrophoresis indicated that G5 PD dendrimer and siRNA formed NPs at weight ratios >0.5:1. G5 PD dendrimer showed effective luciferase gene silencing when weight ratio was 3.0:1 and above. Treatment with endocytosis inhibitors showed that clathrin-mediated endocytosis was the main endocytic pathway by which G5-PD dendrimer/siRNA NPs enter the cell.
These results show that the novel G5 PD dendrimer has high siRNA delivery activity and is promising as a delivery agent for its therapeutic application.
将 siRNA 递送到细胞中仍然是一个关键的挑战。我们的实验室已经证明，一种具有改良季戊四醇衍生物核心的新型聚酰胺胺（PAMAM）树枝状聚合物（PD 树枝状聚合物）具有很高的质粒 DNA 转染效率和低细胞毒性。在这里，我们评估 PD 树枝状聚合物作为 siRNA 载体。
琼脂糖凝胶电泳和 AFM 用于确认第 5 代（G5）-PD 树枝状聚合物/ siRNA 纳米颗粒（NPs）的形成。使用 G5 PD 树枝状聚合物/抗荧光素酶 siRNA NPs 转染具有稳定荧光素酶表达的 SK Hep-1 细胞。还研究了各种内吞途径抑制剂对 SK Hep-1 细胞中 G5 PD 树枝状聚合物/ siRNA NPs 摄取的影响。
琼脂糖凝胶电泳表明，G5 PD 树枝状聚合物和 siRNA 在重量比> 0.5：1 时形成 NPs。当重量比为 3.0：1 及以上时，G5 PD 树枝状聚合物显示出有效的荧光素酶基因沉默作用。内吞作用抑制剂的处理表明，网格蛋白介导的内吞作用是 G5-PD 树枝状聚合物/ siRNA NPs 进入细胞的主要内吞途径。
这些结果表明，新型 G5 PD 树枝状聚合物具有很高的 siRNA 递送活性，有望成为其治疗应用的递送剂。
