Preparation,Cellular Uptake Of Graphene-based Nanocomposites And Their Effects On Cell Functions

Graphene and its derivatives have superior properties,such as extraordinary mechanical strength,a large surface area and good chemical stability.Graphene-based nanocomposites have great potential for application in the area of energy storage,environmental governance and biomedicine,etc.However,the biosafety and biocompatibility of these nanocomposites are still not fully addressed.Therefore,their influences on cell viability and cell functions are of paramount importance for the development and application of new-type graphene-based nanocomposites.In this study,three kinds of graphene-based nanocomposits were synthesized,and their interactions with cells were studied.1)rGO/TNT nanocomposites were successfully synthesized through a modified hydrothermal treatment method.Then,the interactions between the rGO/TNT nanocomposites and Raw264.7 mouse monocyte-macrophage cells were further investigated.The results show that the rGO/TNT nanocomposites could be internalized by Raw264.7 cells and mainly gathered inside the cytoplasm.No rGO/TNT nanocomposites were observed in the nucleus.Moreover,the rGO/TNT nanocomposites exhibited low cytotoxicity toward Raw264.7 cells at a lower dose,though they may exhibit cytotoxicity to some extent at very high concentrations.In addition,the uptake of the nanocomposites influenced the cell cytoskeleton organization,while the cell adhesion and migration abilities were also impaired.2)NGO with an average size of 168.3 nm was successfully synthesized by a modified Hummers'method and further treated with an ultrasonic cell disruptor.NGO-PEI and NGO-PEG complexes with different zeta potentials of 47.2 mV and-43.0 mV,respectively,were successfully synthesized through amide linkages between the COOH groups of GO and the NH₂ groups of PEI and PEG.In addition,the complexes both showed good stability in DMEM containing 10%FBS solution.Then,the interactions between NGO-PEI and NGO-PEG complexes and Raw264.7mouse monocyte-macrophage cells were investigated.The NGO-PEI and NGO-PEG complexes could both be internalized by Raw264.7 cells.Moreover,the NGO-PEI complex not only gathered in endosomes but also in the cytoplasm.However,the NGO-PEG complex gathered in endosomes only.Compared with the NGO-PEG complex,the NGO-PEI complex showed higher intracellular delivery efficiency in Raw264.7 cells.The MTT tests showed that both NGO-PEI and NGO-PEG exhibited very low cytotoxicity towards Raw264.7 cells.And NGO-PEG complex would cause a stronger immune response in Raw264.7 cells,while NGO-PEI complex show no obvious stimulus response.3)NGO loaded?GA/PAH?₄ ployelectrolyte microcapsules were successfully synthesized by a modified template and layer-by-layer?LBL?self-assembly method.The morphology and structure of the resulting NGO@?GA/PAH?₄ ployelectrolyte microcapsules were verified by scanning electron microscopy?SEM?,the microcapsules were spherical with hollow structure with an average size of 2?m,and were apt to collapse under dry condition.The loaded GO sheets in the?GA/PAH?₄microcapsules were observed in transmission electron microscope?TEM?.In addition,the drug entrapment efficiency?EE?and drug loading capacity?LC?of DOX in NGO@?GA/PAH?₄ and single?GA/PAH?₄ were tested and calculated.Compared with single?GA/PAH?₄,NGO@?GA/PAH?₄ showed a higher EE of 93%and LC of46%.To test the drug release performance,NGO@?GA/PAH?₄ showed a pH-dependent and sustained manner under physiological condition.Then,the interactions between NGO@?GA/PAH?₄ and Hela cells were investigated.The NGO@?GA/PAH?₄ could easily be internalized by Hela cells,indicating the loaded of NGO inside the microcapsules would not change the surface properties.The MTT tests showed that single NGO@?GA/PAH?₄ exhibited very low cytotoxicity towards Hela cells when at a low concentration.Further studies on the cytotoxicity of DOX loaded NGO@?GA/PAH?₄ microcapsules towards Hela cells were evaluated.Improved efficacy of DOX loaded NGO@?GA/PAH?₄ microcapsules was observed compared to DOX loaded?GA/PAH?₄ group.Together,these results indicated that the NGO@?GA/PAH?₄ ployelectrolyte microcapsules,with a higher LC and pH-dependent drug release manner,exhibited the potential to be used in effective drug and gene delivery.