Research ArticleAPPLIED SCIENCES AND ENGINEERING

Transport of a graphene nanosheet sandwiched inside cell membranes

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Science Advances  07 Jun 2019:
Vol. 5, no. 6, eaaw3192
DOI: 10.1126/sciadv.aaw3192
  • Fig. 1 Experimental evidence of the sandwiched graphene membrane superstructure and the following cell responses.

    (A) Cartoon illustrating the superstructure integrating GO inside the cell membrane. (B) Cyro-TEM images (left) and tomography views (right) of the lipid vesicles in the presence of GO. Scale bars, 20 nm. The sandwiched graphene-membrane superstructure was formed via three possible processes: primary contact, perpendicular/slant insertion, and final settlement. The lipid vesicle in the presence of a GO nanosheet (as indicated by the arrows) shows higher mass density. (C) Super-resolution confocal images showing the GO sandwiched in the membrane of different cells (macrophage Mφ and nonphagocyte Nonφ). The green fluorescence indicates the phospholipid bilayer membranes bound by the lipid probe [3,3′-dioctadecyloxacarbocyanine perchlorate (DiO)], while the false-colored red spots indicate the GO signal. Yellow spots indicate the sandwiched GO in the membrane. Scale bars, 1 μm. (D) TEM images exhibiting the GO-sandwiched superstructure. The GO is marked by the yellow triangle. Scale bar, 100 nm. (E) AFM images and the roughness index of the cell membrane with embedded GOs. (F) The Young’s modulus indicating the rigidity change of cell surface upon GO interaction. (G) FRAP analysis showing the fluidity of the membrane lipids. Diff Co and the t1/2 represent the diffusion coefficient and half-time for the fluorescence recovery of membrane after photobleaching, respectively.

  • Fig. 2 Transition of diffusion patterns of the sandwiched GO from Brownian to Lévy and even directional dynamics.

    (A to C) Representative trajectories tracked for 4 × 104 τ, when the interaction parameter between GO and lipid tails, χGT, is (A) 1.43, (B) 7.15, and (C) 14.3. Colors denote the time lapse of each trajectory. In (B), arrows and dashed ovals indicate alternating persistent segments and jiggling periods, respectively, which are quantitatively identified by wavelet analysis shown in the inset. Inset of (C): The averaged position of GO in the z axis as a function of time, where z = 0 represents the middle of the membrane. (D) Time-averaged mean square displacement 〈Δr2(t)〉 as a function of lag time on log-log scales for persistent (red) and jiggling (blue) segments of the trajectory at χGT = 7.15. (E) The probability distribution of step length, l, on log-log scales, showing exponential statistics at χGT = 1.43 but power-law statistics with slope −2 at χGT = 7.15 and −1 at χGT = 14.3. (F) Statistic distributions of angles between neighboring persistent segments for the trajectories at χGT = 7.15 (top) and 14.3 (bottom). (G) The plot of the exponent defining the power-law distribution of step length, μ, against χGT for the systems exhibiting Lévy dynamics. The step-length distribution from about 50 sets of simulated trajectories at each χGT is summarized. The color bar encodes the value of μ.

  • Fig. 3 Combined simulations and experiments demonstrate the sandwiched GO–induced pore in the leaflets of cell membranes.

    (A) Probability distribution of the number of GO beads contacting solvent, N, during a simulation period of 4 × 104 τ, p(N), at a series of χGT encoded by the color bar. (B to E) p(N) at various χGT: (B) 1.43, (C) 7.15, (D) 10.01, and (E) 14.3. N indicates the size of the pore in the leaflet of the membrane, as schemed by the insets. (F) The time-averaged N in the whole period, N¯, as a function of χGT. The red circle in the plot marks the start point from which N¯ is above 0 and pore occurs. The insets are two snapshots showing the typical GO-sandwiched structures without (left) and with (right) a pore, where the dashed squares mark the contour of GO and the red outline displays the pore. (G) QCM data (frequency, Δf; dissipation, ΔD) during the interaction between GO and the lipid bilayer, as schematically illustrated by the right diagram.

  • Fig. 4 Analytical model of the sandwiched GO–induced pore.

    (A) Schematic representation of the detailed structure and parameters considered in the analytical model of the sandwiched GO–induced pore. (B) The energy cost of pore formation, ER, as a function of pore radius at different Ka, i.e., the interaction energy density of lipid tails with GO. The solid and hollow circles at each plot mark the local maximum and the local minimum of ER, respectively. The dashed cyan and purple curves give the analytical solutions of both of these types of points, which meet at the transition point with Ka = 25 kBT/nm2. (C) Plot of GO–lipid tail interaction energy, Ea, versus the number of GO beads contacting solvent N, where χGT = 10.01. Averaged from 5 million time steps (raw data obtained from simulation results and shown as gray circles), the yellow crosses have the linear fitting indicated in the figure. (D) The ratio between the most probable pore (pore at the energy minimum) area and GO area as a function of Ka. The green points and red curve represent the results from the simulation and analytical model, respectively. Top inset: The maximum of GO beads contacting solvent, Nmax, remains almost unchanged with increasing χGT. Bottom inset: The plot of Ka used in the analytical model versus χGT used in the simulation, approximating a linear fitting as indicated by the dashed red line.

  • Fig. 5 Correlation between diffusive dynamics and membrane-pore states elucidates the mechanisms of Lévy walk and directional motion of sandwiched GO.

    (A) The area ratio between pore and GO and the diffusion exponent, α, as a function of Ka. The red and blue curves depict the cases of the most probable pore area and the averaged pore area, respectively. The shaded regions divide four regimes of the membrane-pore states, where the colors correspond to those adopted in Fig. 3 (B to E). (B) The energy barrier in the ER-R curve, ΔE (Fig. 4B), as a function of Ka. The dashed blue line plots the thermal energy, 1 kBT. Inset: Time-averaged mean square displacement 〈Δr2(t)〉 as a function of lag time on log-log scales for various Ka, where the slopes give α. (C and D) Successive stages of a sandwiched GO exhibiting Lévy walk at χGT = 10.01 (Ka = 25.29 kBT/nm2) (C) and directional motion at χGT = 14.3 (Ka = 34.74 kBT/nm2) (D). The dashed squares mark the contour of GO, the red outlines display the pore induced by the sandwiched GO, and the arrows denote the moving direction of the GO. The times of the simulation snapshots are (from left to right) 22,160 τ, 22,240 τ, 22,480 τ, 22,800 τ, and 23,040 τ for (C) and 22,320 τ, 22,360 τ, 22,400 τ, 22,440 τ, and 22,480 τ for (D).

  • Fig. 6 The feasibility of delivering membrane-specific drugs via the sandwiched superstructure.

    (A) Relative cell viability of VTB with the assistant of GO or liposome in breast cancer cells MCF-7. Control, ctrl. (B) Corresponding live (green)/dead (red) images at a dose of 5 μg/ml. (C) Analysis of total drug entry amount and relative drug distribution in the cell membrane. (D) The initial and final simulation configurations of the drug beads undergoing the intramembrane delivery from a sandwiched GO. Left: The GO carrying drug beads is moved away from the membrane interior for the exhibition of its initial configuration, and the dashed square marks the contour of the sandwiched GO. (E) The initial and final configurations of the drug beads undergoing the intracellular delivery. In (D) and (E), the right panel with transparent membranes displays the final configuration, where the first drug bead is captured by the transmembrane receptor, as featured by the circle. (F) The capturing time as a function of the drug-bead number for the intramembrane and intracellular deliveries.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/5/6/eaaw3192/DC1

    Section S1. Details of simulation methods, models, and data analysis

    Section S2. Details of analytical models

    Fig. S1. Characterizations of the prepared 2D material of GO.

    Fig. S2. Cryo-TEM images of the blank liposomes, the formation process of the sandwiched GO superstructure, and the sandwiched structure at different detection angles.

    Fig. S3. Tomography views of the 3D map for the GO-membrane superstructure and the blank liposome vesicles.

    Fig. S4. A series of TEM images of the GO–cell membrane interaction and the cells after exposure to different dimensional materials.

    Fig. S5. The interaction between GO and the cells.

    Fig. S6. Molecular models for the individual entities used in the simulations.

    Fig. S7. Translocation pathways of GO across the lipid membrane toward the sandwiched GO structure.

    Fig. S8. Translocation pathways of GO, with the model representing outcomes from standard oxidization, across the lipid membrane.

    Fig. S9. The displacement probability distributions and the translational diffusion coefficients of the GO sandwiched inside the membrane.

    Fig. S10. A schematic diagram illustrating the definition of the turning angle between the neighboring persistent segments.

    Fig. S11. Diffusive properties of GO with χGT = 7.15.

    Fig. S12. Transition of diffusion patterns of the sandwiched GO from Brownian to Lévy and even directional dynamics with a membrane size of 40 × 40 rc2.

    Fig. S13. Diffusive dynamics and membrane-pore states of a circular GO.

    Fig. S14. Simulation results demonstrate various membrane-pore states and the mechanism of pore formation.

    Fig. S15. Representative snapshots from simulations feature the sandwiched GO–induced pores in the single leaflet of cell membranes.

    Fig. S16. The energy of the sandwiched GO–induced pore as a function of the radius of the pore R at KaKa0 (Ka0 ≈ 25 kBT/nm2).

    Fig. S17. Correlation between the analytical model and simulation results.

    Fig. S18. Diffusive dynamics of lipids varies from Fickian to superdiffusive.

    Fig. S19. Sandwiched GO–induced pores in the single leaflet of the cell membrane for the GO model representing outcomes from standard oxidization processes.

    Fig. S20. The efficacy of the GO-sandwiched structure on drug delivery.

    Fig. S21. Diffusive dynamics of a representative drug bead captured by the transmembrane receptor.

    Fig. S22. Probability distribution of the capturing time for the drug beads released from the sandwiched GO and the center of the intracellular region.

    Movie S1. Detailed translocation pathway of GO across the lipid membrane toward the sandwiched GO structure at χGT = 15.73.

    Movie S2. Detailed translocation pathway of the GO model, representing outcomes from standard oxidization processes with the oxidation degree ρO = 0.3, across the lipid membrane toward the sandwiched GO structure.

    Movie S3. Detailed diffusive dynamics of a sandwiched GO exhibiting Brownian motion at χGT = 1.43.

    Movie S4. Detailed diffusive dynamics of a sandwiched GO exhibiting Lévy walk at χGT = 10.01.

    Movie S5. Detailed diffusive dynamics of a sandwiched GO exhibiting directional motion at χGT = 14.3.

    References (5559)

  • Supplementary Materials

    The PDF file includes:

    • Section S1. Details of simulation methods, models, and data analysis
    • Section S2. Details of analytical models
    • Fig. S1. Characterizations of the prepared 2D material of GO.
    • Fig. S2. Cryo-TEM images of the blank liposomes, the formation process of the sandwiched GO superstructure, and the sandwiched structure at different detection angles.
    • Fig. S3. Tomography views of the 3D map for the GO-membrane superstructure and the blank liposome vesicles.
    • Fig. S4. A series of TEM images of the GO–cell membrane interaction and the cells after exposure to different dimensional materials.
    • Fig. S5. The interaction between GO and the cells.
    • Fig. S6. Molecular models for the individual entities used in the simulations.
    • Fig. S7. Translocation pathways of GO across the lipid membrane toward the sandwiched GO structure.
    • Fig. S8. Translocation pathways of GO, with the model representing outcomes from standard oxidization, across the lipid membrane.
    • Fig. S9. The displacement probability distributions and the translational diffusion coefficients of the GO sandwiched inside the membrane.
    • Fig. S10. A schematic diagram illustrating the definition of the turning angle between the neighboring persistent segments.
    • Fig. S11. Diffusive properties of GO with χGT = 7.15.
    • Fig. S12. Transition of diffusion patterns of the sandwiched GO from Brownian to Lévy and even directional dynamics with a membrane size of 40×40 rc2.
    • Fig. S13. Diffusive dynamics and membrane-pore states of a circular GO.
    • Fig. S14. Simulation results demonstrate various membrane-pore states and the mechanism of pore formation.
    • Fig. S15. Representative snapshots from simulations feature the sandwiched GO–induced pores in the single leaflet of cell membranes.
    • Fig. S16. The energy of the sandwiched GO–induced pore as a function of the radius of the pore R at KaKa0 (Ka0 ≈ 25 kBT/nm2).
    • Fig. S17. Correlation between the analytical model and simulation results.
    • Fig. S18. Diffusive dynamics of lipids varies from Fickian to superdiffusive.
    • Fig. S19. Sandwiched GO–induced pores in the single leaflet of the cell membrane for the GO model representing outcomes from standard oxidization processes.
    • Fig. S20. The efficacy of the GO-sandwiched structure on drug delivery.
    • Fig. S21. Diffusive dynamics of a representative drug bead captured by the transmembrane receptor.
    • Fig. S22. Probability distribution of the capturing time for the drug beads released from the sandwiched GO and the center of the intracellular region.
    • References (5559)

    Download PDF

    Other Supplementary Material for this manuscript includes the following:

    • Movie S1 (.mov format). Detailed translocation pathway of GO across the lipid membrane toward the sandwiched GO structure at χGT = 15.73.
    • Movie S2 (.mov format). Detailed translocation pathway of the GO model, representing outcomes from standard oxidization processes with the oxidation degree ρO = 0.3, across the lipid membrane toward the sandwiched GO structure.
    • Movie S3 (.mov format). Detailed diffusive dynamics of a sandwiched GO exhibiting Brownian motion at χGT = 1.43.
    • Movie S4 (.mov format). Detailed diffusive dynamics of a sandwiched GO exhibiting Lévy walk at χGT = 10.01.
    • Movie S5 (.mov format). Detailed diffusive dynamics of a sandwiched GO exhibiting directional motion at χGT = 14.3.

    Files in this Data Supplement:

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