Department of Molecular Medicine, Sapienza University of Rome, Viale Regina Elena 291, 00161, Rome, Italy.
Nanoscale. 2019 Feb 7;11(6):2733-2741. doi: 10.1039/c8nr09245a.
Graphene oxide (GO) is a single-atomic-layered material made of a sheet of oxidized carbon atoms arranged in a honeycomb structure. Thanks to the notable physical and chemical properties of GO, GO-based nanomaterials have applications in many fields of research, including gene delivery. It has been reported that pristine GO can absorb single-stranded DNA and RNA through π-π stacking, which cannot be used as a gene carrier because it is hard to load double-stranded DNA (dsDNA). To tackle this issue, this work was aimed at developing a hybrid nanoparticle (NP) system made of GO coated with cationic lipids (hereafter referred to as GOCL) with suitable physical-chemical properties for gene delivery applications. To this end, nanosized GO flakes (nGO) were coated with the cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) by microfluidic mixing. Comprehensive characterization of GOCL NPs was performed by a combination of dynamic light scattering (DLS), micro-electrophoresis and atom force microscopy (AFM). Our results show that GOCL NPs exhibit adequate size (<150 nm) and surface charge (ξ = +15 mV) for gene delivery purposes. Complexes made of GOCL NPs and plasmid DNA (pDNA) were used to transfect human cervical cancer cells (HeLa) and human embryonic kidney (HEK-293) cells. Pristine nGO and DOTAP cationic liposomes were used as a reference. GOCL NPs exhibited a similar TE but a much higher cell viability compared with DOTAP cationic liposomes. Confocal fluorescence microscopy provided a reasonable explanation for the superior performance of GOCL/DNA complexes showing that they are much more numerous, regular in size and homogeneously distributed than DOTAP/DNA complexes, thus splitting their gene payload over the entire cell population. Because of the imperative demand for efficient and safe nanocarriers, this study will contribute to the development of novel surface-functionalized GO-based hybrid gene vectors.
氧化石墨烯（GO）是由单层碳原子排列在蜂窝状结构中的氧化碳原子组成的单层材料。由于 GO 的显著物理和化学性质，基于 GO 的纳米材料在许多研究领域都有应用，包括基因传递。据报道，原始 GO 可以通过π-π 堆积吸收单链 DNA 和 RNA，由于难以负载双链 DNA（dsDNA），因此不能用作基因载体。为了解决这个问题，本工作旨在开发一种由带正电荷的脂质（以下简称 GOCL）涂覆的氧化石墨烯（GO）制成的混合纳米粒子（NP）系统，该系统具有适合基因传递应用的物理化学性质。为此，通过微流混合将纳米 GO 薄片（nGO）涂覆上阳离子脂质 1,2-二油酰基-3-三甲胺丙烷（DOTAP）。通过动态光散射（DLS）、微电泳和原子力显微镜（AFM）的组合对 GOCL NPs 进行了综合表征。我们的结果表明，GOCL NPs 具有适合基因传递的适当大小（<150nm）和表面电荷（ξ=+15mV）。GOCL NPs 与质粒 DNA（pDNA）形成的复合物被用于转染人宫颈癌（HeLa）和人胚肾（HEK-293）细胞。将原始 nGO 和 DOTAP 阳离子脂质体用作参考。与 DOTAP 阳离子脂质体相比，GOCL NPs 表现出相似的转染效率（TE），但细胞活力更高。共焦荧光显微镜为 GOCL/DNA 复合物的优异性能提供了合理的解释，表明它们比 DOTAP/DNA 复合物更多、尺寸更规则且均匀分布，从而将其基因有效负载分配到整个细胞群体中。由于对高效和安全纳米载体的迫切需求，本研究将有助于开发新型表面功能化的基于 GO 的混合基因载体。
