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Microfabricated Particles for Engineered Drug Therapies: Elucidation into the Mechanisms of Cellular Internalization of PRINT Particles

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Abstract

Purpose

To investigate the cellular internalization pathways of shape- and size-specific particles as a function of zeta potential in different cell types.

Methods

A top-down particle fabrication technique called PRINT was utilized to fabricate monodisperse 1 μm cylindrical particles. Cellular internalization of these PRINT particles was monitored using confocal microscopy, flow cytometry, and transmission electron microscopy. The endocytic pathway used by 1 μm cationic PRINT particles was evaluated using different inhibitory strategies. Cytotoxicity assays were used to determine the toxicity of both cationic and anionic PRINT particles in multiple cell types.

Results

Particle internalization was confirmed using confocal microscopy, flow cytometry and transmission electron microscopy. The mechanism of internalization of positively charged PRINT particles was found to be predominantly clathrin-mediated endocytosis and macropinocytosis with very few particles utilizing a caveolae-mediated endocytic pathway. The exposed charge on the surface of the particles had a significant effect on the rate of endocytosis in all cell types tested, except for the macrophage cells. No significant cytotoxicity was observed for all PRINT particles used in the present study.

Conclusions

Cylindrical 1 μm PRINT particles were readily internalized into HeLa, NIH 3T3, OVCAR-3, MCF-7, and RAW 264.7 cells. Particles with a positive zeta potential exhibited an enhanced rate of endocytosis compared to negatively charged particles with identical sizes and shapes. It was found that PRINT particles with a positive zeta potential were endocytosed into HeLa cells using predominantely clathrin-mediated and macropinocytotic pathways.

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Acknowledgements

This work was supported in part by the STC Program of the NSF (CHE-9876674), NIH PPG P01-GM059299-07, NIH 5-654-CA119343-02 (the Carolina Center of Cancer Nanotechnology Excellence), the William R. Kenan Professorship of the University of North Carolina at Chapel Hill, and Liquidia Technologies (SRA 04-0013).

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Authors and Affiliations

  1. Department of Chemistry and Carolina Center of Cancer Nanotechnology Excellence, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, 27599, USA

    Stephanie E. A. Gratton, Mary E. Napier, Patricia A. Ropp, Shaomin Tian & Joseph M. DeSimone

  2. Department of Chemical Engineering, North Carolina State University, Raleigh, North Carolina, 27695, USA

    Joseph M. DeSimone

  3. Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, 27599, USA

    Joseph M. DeSimone

  4. Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, 27599, USA

    Joseph M. DeSimone

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  1. Stephanie E. A. Gratton
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  2. Mary E. Napier
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Correspondence to Joseph M. DeSimone.

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Gratton, S.E.A., Napier, M.E., Ropp, P.A. et al. Microfabricated Particles for Engineered Drug Therapies: Elucidation into the Mechanisms of Cellular Internalization of PRINT Particles. Pharm Res 25, 2845–2852 (2008). https://doi.org/10.1007/s11095-008-9654-8

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  • DOI: https://doi.org/10.1007/s11095-008-9654-8

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