Department of Biomedical Engineering, University of Rochester , Rochester, New York 14627, United States.
Center for Musculoskeletal Research, University of Rochester Medical Center , Rochester, New York 14627, United States.
ACS Nano. 2018 Jan 23;12(1):187-197. doi: 10.1021/acsnano.7b05528. Epub 2017 Dec 15.
Nanoparticles (NPs) interact with complex protein milieus in biological fluids, and these interactions have profound effects on NP physicochemical properties and function. Surprisingly, most studies neglect the impact of these interactions, especially with respect to NP-mediated siRNA delivery. Here, the effects of serum on colloidal stability and siRNA delivery of a pH-responsive micellar NP delivery system were characterized. Results show cationic NP-siRNA complexes aggregate in ≥2% serum in buffer, but are stable in serum-free media. Furthermore, nonaggregated NP-siRNA delivered in serum-free media result in 4-fold greater siRNA uptake in vitro, compared to aggregated NP-siRNA. Interestingly, pH-responsive membrane lysis behavior, which is required for endosomal escape, and NP-siRNA dissociation, necessary for gene knockdown, are significantly reduced in serum. Consistent with these data, nonaggregated NP-siRNA in serum-free conditions result in highly efficient gene silencing, even at doses as low as 5 nM siRNA. NP-siRNA diameter was measured at albumin and IgG levels mimicking biological fluids. Neither albumin nor IgG alone induces NP-siRNA aggregation, implicating other serum proteins in NP colloidal instability. Finally, as a proof-of-principle that stability is maintained in established in vivo models, transmission electron microscopy reveals NP-siRNA are taken up by ductal epithelial cells in a nonaggregated state when injected retroductally into mouse salivary glands in vivo. Overall, this study shows serum-induced NP-siRNA aggregation significantly diminishes efficiency of siRNA delivery by reducing uptake, pH-responsive membrane lysis activity, and NP-siRNA dissociation. Moreover, these results highlight the importance of local NP-mediated drug delivery and are broadly applicable to other drug delivery systems.
纳米粒子（NPs）与生物流体中的复杂蛋白质环境相互作用，这些相互作用对 NP 的物理化学性质和功能有深远的影响。令人惊讶的是，大多数研究都忽略了这些相互作用的影响，尤其是在 NP 介导的 siRNA 递送上。在这里，研究了血清对 pH 响应胶束 NP 递药系统胶体稳定性和 siRNA 递药的影响。结果表明，带正电荷的 NP-siRNA 复合物在缓冲液中的 2%以上血清中聚集，但在无血清培养基中稳定。此外，在无血清培养基中递药的未聚集 NP-siRNA 与聚集的 NP-siRNA 相比，在体外的 siRNA 摄取量增加了 4 倍。有趣的是，对于内涵体逃逸所必需的 pH 响应膜裂解行为以及基因敲低所必需的 NP-siRNA 解离，在血清中显著降低。与这些数据一致的是，在无血清条件下的未聚集 NP-siRNA 导致高效的基因沉默，即使在低至 5 nM siRNA 的剂量下也是如此。在模拟生物流体的白蛋白和 IgG 水平下测量了 NP-siRNA 的直径。单独的白蛋白或 IgG 都不会引起 NP-siRNA 聚集，表明其他血清蛋白在 NP 胶体不稳定性中起作用。最后，作为在既定体内模型中保持稳定性的原理验证，当将 NP-siRNA 通过逆行注射到小鼠唾液腺中时，透射电子显微镜显示 NP-siRNA 以未聚集的状态被摄取到导管上皮细胞中。总的来说，这项研究表明，血清诱导的 NP-siRNA 聚集通过降低摄取、pH 响应膜裂解活性和 NP-siRNA 解离来显著降低 siRNA 递药效率。此外，这些结果强调了局部 NP 介导的药物递药的重要性，并且广泛适用于其他药物递药系统。
