Cifuentes Javier, Muñoz-Camargo Carolina, Cruz Juan C
Department of Biomedical Engineering, School of Engineering, Universidad de Los Andes, Carrera 1 No. 18A-12, Bogotá 111711, Colombia.
Nanomaterials (Basel). 2022 Aug 19;12(16):2857. doi: 10.3390/nano12162857.
The development of novel regenerative technologies based on the implementation of natural extracellular matrix (ECM), or individual components of ECM combined with multifunctional nanomaterials such as graphene oxide and reduced graphene oxide, has demonstrated remarkable results in wound healing and tissue engineering. However, the synthesis of these nanocomposites involves great challenges related to maintaining the biocompatibility with a simultaneous improvement in their functionalities. Based on that, in this research we developed novel nanoengineered ECM-scaffolds formed by mixing small intestinal submucosa (SIS) with graphene oxide (GO)/reduced graphene oxide (rGO) to improve electrical conductivity while maintaining remarkable biocompatibility. For this, decellularized SIS was combined with GO to form the scaffold precursor for subsequent lyophilization, chemically crosslinking and in situ reduction. The obtained GO and rGO were characterized via Raman spectroscopy, Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), X-ray diffraction (XRD), electrical conductivity testing and atomic force microscopy (AFM). The results confirm the suitable synthesis of GO, the effective reduction to rGO and the significant increase in the electrical conductivity (more than four orders of magnitude higher than bare GO). In addition, the graphene oxide/reduced graphene oxide-SIS scaffolds were characterized via Raman spectroscopy, FTIR, TGA, SEM, porosity assay (higher than 97.5% in all cases) and protein secondary structural analysis. Moreover, the biocompatibility of scaffolds was studied by standardized assays of hemolysis activity (less than 0.5%), platelet activation and deposition, and cell viability in Vero, HaCat and HFF-1 cells (higher than 90% for all evaluated cell lines on the different scaffolds). The obtained results confirm the remarkable biocompatibility, as supported by high hemocompatibility, low cytotoxicity and no negative impact on platelet activation and deposition. Finally, structural characteristics such as pore size and interconnectivity as well as superior cell attachment abilities also corroborated the potential of the developed nanoengineered ECM-scaffolds as a multifunctional nanoplatform for application in regenerative medicine and tissue engineering.
基于天然细胞外基质（ECM）或ECM的单个成分与多功能纳米材料（如氧化石墨烯和还原氧化石墨烯）相结合而开发的新型再生技术，在伤口愈合和组织工程方面已取得显著成果。然而，这些纳米复合材料的合成面临着巨大挑战，即要在保持生物相容性的同时提高其功能。基于此，在本研究中，我们通过将小肠黏膜下层（SIS）与氧化石墨烯（GO）/还原氧化石墨烯（rGO）混合，开发了新型纳米工程化ECM支架，以提高导电性并同时保持显著的生物相容性。为此，将脱细胞的SIS与GO结合形成支架前体，随后进行冻干、化学交联和原位还原。通过拉曼光谱、傅里叶变换红外光谱（FTIR）、热重分析（TGA）、X射线衍射（XRD）、电导率测试和原子力显微镜（AFM）对所得的GO和rGO进行了表征。结果证实了GO的合适合成、有效还原为rGO以及电导率的显著增加（比裸GO高四个数量级以上）。此外，通过拉曼光谱、FTIR、TGA、扫描电子显微镜（SEM）、孔隙率测定（所有情况下均高于97.5%）和蛋白质二级结构分析对氧化石墨烯/还原氧化石墨烯-SIS支架进行了表征。此外，通过溶血活性（小于0.5%）、血小板活化和沉积以及Vero、HaCat和HFF-1细胞中的细胞活力的标准化测定来研究支架的生物相容性（在不同支架上，所有评估的细胞系的细胞活力均高于90%）。所得结果证实了其显著的生物相容性，高血液相容性、低细胞毒性以及对血小板活化和沉积无负面影响均支持了这一点。最后，孔径和互连性等结构特征以及优异的细胞附着能力也证实了所开发的纳米工程化ECM支架作为多功能纳米平台在再生医学和组织工程中的应用潜力。
