Development of Man-rGO for Targeted Eradication of Macrophage Ablation.This study was aimed to develop and evaluate a smart nanosystem that targeted photothermal ablation of inflammatory macrophages in atherosclerotic plaque.  Mannosylated-reduced graphene oxide (Man-rGO) was synthesized using three step procedures: (1) preparation of ox-GOs, (2) microwave-assisted synthesis of PEI-rGOs, and (3) mannosylation of PEI-rGO using reductive amination reaction (Man-rGOs).  The ζ-potential of Man-rGO that signifies electrophoretic mobility of the charged surface was examined using Zetasizer Nano ZS.  The effects of Man-rGO on the cell viability was evaluated using LDH assay and AlamarBlue assay.  The targeting efficacy of Man-rGO was assessed using the cellular uptake rate by M2-polarized (i.e., which is induced by IL-4) macrophage.  The effects of NOMela loaded in Man-rGO on the enhancement of phagocytosis were evaluated by examining the phagocytic clearance rate of zymosan-FITC particles.  The mannose functionalization (Man-rGO) of PEI-rGO produced a greater number of amide linkages formed by reductive amination reaction between PEI-rGO and mannose.  The ζ-potential of PEI-rGO was +30.6 ± 3.3 mV, whereas that of Man-rGO was down to +13.1 ± 3.8 mV upon interaction with mannose mainly due to the conjugation of mannose on the PEI-rGO surface.  Near-infrared (NIR) irradiation increased the temperature of Man-rGO solution to around 45 °C, suggesting that Man-rGO is more potent than ox-GO or rGO in photothermal ablation activity triggered by NIR laser irradiation (808 nm).  The fluorescent microscopy images of cells after 1 h incubation demonstrated that Man-rGO were mainly accumulated at the subcellular level where the mannose receptors were overexpressed.  The cell viability of macrophages significantly decreased upon exposure to Man-rGO irradiated with NIR, but no changes were observed from that of mast cells (for mast cells, 98.3 ± 0.3%; for macrophages, 67.8 ± 1.3%, p < 0.01), indicating that Man-rGO achieved enhanced targetability toward mannose receptor mediated cellular uptake.  It was concluded that the enhanced photothermal ablation activity of Man-rGO triggered by NIR laser irradiation was mediated through their targetability toward overexpressed mannose receptor, a marker of M2-phenotype of macrophage.  The results of this study supported that Man-rGO can serve as an efficient platform for the targeted therapy against atherosclerosis.
Man-rGO