Dynamic interactions and intracellular fate of label-free, thin graphene oxide sheets within mammalian cells: role of lateral sheet size

Abstract

Graphene oxide (GO) holds great potential for biomedical applications, however fundamental understanding of the way it interacts with biological systems is still lacking even though it is a prerequisite for successful clinical translation. In this study, we exploited intrinsic fluorescent properties of GO to establish the relationship between lateral dimensions of the material, its cellular uptake mechanism and intracellular fate. Label-free GO with distinct lateral dimensions, small (s-GO) and ultra-small (us-GO), was synthesized and thoroughly characterised both in water and in biologically relevant cell culture medium. Interactions of the material with a range of non-phagocytic mammalian cell lines (BEAS-2B, NIH/3T3, HaCaT, 293T) were studied using a combination of complementary analytical techniques (confocal microscopy, flow cytometry and TEM). The uptake mechanism was interrogated using a range of pharmaceutical inhibitors for main endocytic pathways (ethyl-isopropyl amiloride, monodansylcadaverine, chlorpromazine, genistein, cytochalasin D, latrunculin A, dynasore and sodium azide), and validated using negatively charged polystyrene beads with different diameters (0.1 and 1 μm). Regardless of lateral dimension, both types of GO were found to interact with the plasma membrane and to be efficiently taken up by a panel of cell lines in a timeand dose-dependent manner. s-GO was internalised mainly via macropinocytosis while us-GO used mainly clathrin-and caveolae-mediated endocytosis. Lastly, we show that both s-GO and us-GO terminate in lysosomal compartments for up to 48 h. Our results aim to offer significant insight into the mechanism of interaction of GO with non-phagocytic cell lines that can be exploited for the design of biomedically applicable 2D transport systems.

Stable, concentrated, biocompatible, and defect-free graphene dispersions with positive charge

“…S11 and Table S3 †). Although DLS characterisation is valid only for spherical nanoparticles, it can be used to detect changes in size and polydispersibility of graphene dispersions, 53,66,67 in combination with other characterisation techniques. Table S3 † shows that all graphene dispersions have monodisperse size distribution upon incubation in complete cell culture medium for 24 h. This is confirmed by the lack of sedimentation/precipitation of material upon incu-bation in the cell culture medium (Fig.…”

Defect-free graphene enhances enzyme delivery to fibroblasts derived from patients with lysosomal storage disorders

“…The four types of GBMs that we investigated showed excellent biocompatibility in HPF, MPS VI and Pompe cells in concentrations of up to 100 g/mL (see Figures 2 and S1 -4). In fact, the GBMs employed in our study repeatedly show exceptional biocompatibility in many cell types [12][13][14] . In contrast, many studies reported concentration-dependent cytotoxicity of GBM in primary fibroblast, with the material being toxic at concentrations up to 50 g/mL [34][35][36][37] .…”

“…Gr materials (Gr-TMA 3 and Gr-PS1) were synthesized using liquid phase exfoliation in water, as already reported 13,25 . GO materials (s-GO and us-GO) were synthesized using the modified Hummer's method, as already reported 12,27 .…”

“…In our case, two stabilisers were used: pyrene bearing a linked trimethylammonium functionality (TMA 3 ) and 1-pyrenesulfonic acid sodium salt (PS1), giving rise to cationic and anionic graphene dispersions (Gr-TMA 3 and Gr-PS1) 8 13,25,26 , respectively. Defective graphene, in the form of GO, was produced by utilising the modified Hummer's method 12,27 . The size distribution of the flakes was selected by changing the sonication conditions, as reported in previous work 12,27 , giving rise to two samples: small GO (s-GO) and ultrasmall GO (us-GO).…”

“…Defective graphene, in the form of GO, was produced by utilising the modified Hummer's method 12,27 . The size distribution of the flakes was selected by changing the sonication conditions, as reported in previous work 12,27 , giving rise to two samples: small GO (s-GO) and ultrasmall GO (us-GO).…”