Biomimetic anisotropic polymeric nanoparticles coated with red blood cell membranes for enhanced circulation and toxin removal

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Science Advances  15 Apr 2020:
Vol. 6, no. 16, eaay9035
DOI: 10.1126/sciadv.aay9035
  • Fig. 1 Schematic of anisotropic nanoparticle fabrication and RBC membrane coating.

    (A) Spherical PLGA nanoparticles (NPs) were synthesized and cast into a thin plastic film of 10% polyvinyl alcohol (PVA) and 2% glycerol. Particles were then stretched under heat in one or two dimensions (2D) to generate prolate or oblate ellipsoidal particles, respectively. (B) RBCs underwent hypotonic lysis and were then sonicated to generate sub–200 nm vesicles. RBC-derived vesicles were then coated onto PLGA nanoparticles of all shapes under sonication.

  • Fig. 2 Nanoparticle characterization.

    (A) Uncoated and RBC-coated spherical, prolate ellipsoidal, and oblate ellipsoidal nanoparticles were imaged by TEM, and coated particles were found to have a halo around the particle core that was not present on uncoated particles. Scale bars. 200 nm. (B) Dynamic light scattering (DLS) analysis of coated versus uncoated spherical nanoparticles indicates an increase in diameter of 17.2 nm for coated nanoparticles. (C) Zeta potential measurements of uncoated and coated spherical nanoparticles compared to RBC vesicles. (D) RBC membrane–coated spherical, prolate ellipsoidal, and oblate ellipsoidal particles and uncoated spherical particles were stained with fluorescent anti-CD47. Data are shown as means ± SEM (n = 4 replicates). **P < 0.01 and ***P < 0.001. Student’s t test was used to assess the difference in size between coated and uncoated particles, and one-way analysis of variance (ANOVA) with post hoc Dunnett’s test was used to compare anti-CD47 staining to the uncoated control. a.u., arbitrary units.

  • Fig. 3 In vitro macrophage uptake of nanoparticles.

    (A) Fluorescent nanoparticles were incubated with RAW 264.7 macrophages for 4 hours, and uptake was analyzed by flow cytometry. Relative uptake, as measured by geometric mean fluorescence normalized to untreated cells, was significantly reduced as a result of anisotropy and membrane coating. (B) Macrophage uptake of nanoparticles (pink) was visualized by confocal imaging and found to be reduced for anisotropic coated particles. Macrophages were stained for nuclei (blue) and actin (green). Scale bars, 20 μm. Data are shown as means ± SEM (n = 4 replicates). Statistics were performed by a two-way ANOVA with Bonferroni’s posttests (*P < 0.05 and ***P < 0.001).

  • Fig. 4 In vivo clearance and biodistribution of nanoparticles.

    (A) Blood elimination of nanoparticles following intravenous administration as assessed by fluorescence readings of the blood sample (dots) and fit to a single exponential decay model (lines). (B) Particle bloodstream half-life was derived from the exponential fit of blood decay curves and was increased for RBC membrane–coated particles and prolate ellipsoidal particles. (C) Mice were euthanized after 24 hours, and organs were dissected out and imaged. Data are shown as means ± SEM (n = 3 mice per group). Statistics were performed by a one-way ANOVA with post hoc Tukey’s test (*P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001).

  • Fig. 5 Anisotropic RBC membrane–coated nanoparticles as detoxification treatment.

    (A) Schematic of mechanism of RBC coated nanoparticles (NPs) as detoxification treatment. RBC NPs neutralize alpha toxin by binding toxin that would otherwise bind the body’s RBCs and cause lysis. (B) In vitro evaluation of hemolytic toxin absorption by RBC-coated nanoparticles. The anisotropic particles were able to absorb significantly more alpha toxin as evidenced by reduction in relative lysis. Data are shown as means ± SEM (n = 4 replicates), and one-way ANOVA with post hoc Tukey’s test was used to compare across groups. (C) Survival following intravenous alpha toxin administration followed by nanoparticle administration (n = 6 mice per group). Mice receiving prolate ellipsoidal RBC-coated nanoparticles had a significant long-term survival benefit compared to spherical coated nanoparticles (P = 0.0481), and both anisotropic particle groups had a significant survival benefit over uncoated particles (P = 0.0105 for coated prolate ellipsoidal, P = 0.0169 for coated oblate ellipsoidal) by log rank (Mantel-Cox) test. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.

Supplementary Materials

  • Supplementary Materials

    Biomimetic anisotropic polymeric nanoparticles coated with red blood cell membranes for enhanced circulation and toxin removal

    Elana Ben-Akiva, Randall A. Meyer, Hongzhe Yu, Jonathan T. Smith, Drew M. Pardoll, Jordan J. Green

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