Torres-Vanegas Julian Daniel, Cifuentes Javier, Puentes Paola Ruiz, Quezada Valentina, Garcia-Brand Andres J, Cruz Juan C, Reyes Luis H
Department of Chemical and Food Engineering, Grupo de Diseño de Productos y Procesos (GDPP), Universidad de Los Andes, Bogotá, Colombia.
Department of Biomedical Engineering, Universidad de Los Andes, Bogotá, Colombia.
Front Chem. 2022 Sep 15;10:974218. doi: 10.3389/fchem.2022.974218. eCollection 2022.
Cell-penetrating agents based on functionalized nanoplatforms have emerged as a promising approach for developing more efficient and multifunctional delivery vehicles for treating various complex diseases that require reaching different intracellular compartments. Our previous work has shown that achieving full cellular coverage and high endosomal escape rates is possible by interfacing magnetite nanoparticles with potent translocating peptides such as Buforin II (BUF-II). In this work, we extended such an approach to two graphene oxide (GO)-based nanoplatforms functionalized with different surface chemistries to which the peptide molecules were successfully conjugated. The developed nanobioconjugates were characterized via spectroscopic (FTIR, Raman), thermogravimetric, and microscopic (SEM, TEM, and AFM) techniques. Moreover, biocompatibility was assessed via standardized hemocompatibility and cytotoxicity assays in two cell lines. Finally, cell internalization and coverage and endosomal escape abilities were estimated with the aid of confocal microscopy analysis of colocalization of the nanobioconjugates with Lysotracker Green. Our findings showed coverage values that approached 100% for both cell lines, high biocompatibility, and endosomal escape levels ranging from 30 to 45% and 12-24% for Vero and THP-1 cell lines. This work provides the first routes toward developing the next-generation, carbon-based, cell-penetrating nanovehicles to deliver therapeutic agents. Further studies will be focused on elucidating the intracellular trafficking pathways of the nanobioconjugates to reach different cellular compartments.
基于功能化纳米平台的细胞穿透剂已成为一种有前景的方法，用于开发更高效、多功能的递送载体，以治疗各种需要到达不同细胞内区室的复杂疾病。我们之前的工作表明，通过将磁铁矿纳米颗粒与强效转运肽（如蟾蜍灵II，BUF-II）连接，可以实现完全的细胞覆盖和高内涵体逃逸率。在这项工作中，我们将这种方法扩展到两种基于氧化石墨烯（GO）的具有不同表面化学性质的纳米平台，肽分子成功地与这些平台缀合。通过光谱（傅里叶变换红外光谱、拉曼光谱）、热重分析和显微镜（扫描电子显微镜、透射电子显微镜和原子力显微镜）技术对所开发的纳米生物缀合物进行了表征。此外，通过在两种细胞系中进行标准化的血液相容性和细胞毒性测定来评估生物相容性。最后，借助共聚焦显微镜分析纳米生物缀合物与溶酶体示踪剂绿色的共定位，估计细胞内化、覆盖和内涵体逃逸能力。我们的研究结果表明，两种细胞系的覆盖值均接近100%，具有高生物相容性，Vero细胞系和THP-1细胞系的内涵体逃逸水平分别为30%至45%和12%至24%。这项工作为开发下一代基于碳的细胞穿透纳米载体以递送治疗剂提供了第一条途径。进一步的研究将集中于阐明纳米生物缀合物到达不同细胞区室的细胞内运输途径。
