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Microfluidic-Generated Immunomodulatory Nanoparticles and Formulation-Dependent Effects on Lipopolysaccharide-Induced Macrophage Inflammation

  • Research Article
  • Theme: Rising Stars in Drug Delivery and Novel Carriers
  • Published:
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Abstract

Nanoparticles (NPs) have emerged as a highly useful and clinically translatable drug delivery platform for vast therapeutic payloads. Through the precise tuning of their physicochemical properties, NPs can be engineered to exhibit controlled drug release properties, enhanced circulation times, improved cellular uptake and targeting, and reduced toxicity profiles. Conventional bulk methods for the production of polymeric NPs suffer from the ability to control their size and polydispersity, batch-to-batch variability, significant preparation times, and low recovery. Here, we describe the development and optimization of a high-throughput microfluidic method to produce cargo-less immunomodulatory nanoparticles (iNPs) and their formulation-dependent anti-inflammatory properties for the modulation of lipopolysaccharide (LPS)-induced macrophage responses. Using poly(lactic acid) (PLA) as the core-forming polymer, a rapid and tunable microfluidic hydrodynamic flow-focusing method was developed and optimized to systematically evaluate the role of polymer and surfactant concentration, surfactant chemistry, and flow rate ratio (FRR) on the formation of iNPs. A set of iNPs with 6 different surface chemistries and 2 FRRs was then prepared to evaluate their inherent anti-inflammatory effects using bone marrow-derived macrophages stimulated with the Toll-like receptor 4 agonist, LPS. Finally, a lyophilization study was performed using various cryoprotectants and combinations to identify preferable conditions for iNP storage. Overall, we demonstrate a highly controlled and reproducible method for the formulation of iNPs using microfluidics and their formulation-dependent inherent anti-inflammatory immunomodulatory properties, which represents a potentially promising strategy for the management of inflammation.

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Funding

Research reported in this publication was supported by Startup funds provided by the University of Maryland, Baltimore (UMB), and the National Institute of General Medical Sciences of the National Institutes of Health under Award Number R35GM142752. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

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Authors and Affiliations

  1. Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, 20 N. Pine Street, MD, 21201, Baltimore, USA

    Nhu Truong, Sheneil K. Black, Brianna L. Scotland & Ryan M. Pearson

  2. Department of Microbiology and Immunology, University of Maryland School of Medicine, 685 W. Baltimore Street, MD, 21201, Baltimore, USA

    Jacob Shaw & Ryan M. Pearson

  3. Marlene and Stewart Greenebaum Comprehensive Cancer Center, University of Maryland School of Medicine, 22 S. Greene Street, MD, 21201, Baltimore, USA

    Ryan M. Pearson

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  1. Nhu Truong
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Contributions

NT and RMP contributed to the design of experiments, analysis of data, and compilation of the manuscript. JS and NT contributed to optimization and development of the microfluidic system. SKB and JS executed experiments related to in vitro testing of nanoparticles and cryoprotectant studies. BS operated the SEM and collected images of the nanoparticles. All authors discussed and analyzed the results and contributed to the final version of the manuscript.

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Correspondence to Ryan M. Pearson.

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Guest Editors: Aliasger Salem, Juliane Nguyen and Kristy Ainslie.

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Supplementary Information

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12248_2021_645_MOESM1_ESM.tiff

Figure S1. Six iNP formulations utilizing one of the six various surfactants on the iNP exterior. Chemical structures of (1) poly(ethylene-alt-maleic acid) E60/E400, (2) poly(vinyl alcohol) (PVA), (3) poly(acrylic acid) (PAA), (4) poly(glutamic acid) (PGA), and (5) hyaluronic acid (HA) utilized for iNP synthesis. Supplementary file1 (TIFF 190 kb)

Figure S2. Representative SEM image of iNP-PVA1 prepared using microfluidics. Supplementary file2 (TIFF 325 kb)

12248_2021_645_MOESM3_ESM.tiff

Figure S3: Comparison of two different nanoparticle synthesis methods: emulsification and microfluidics. Both nanoparticles were synthesized using E400 and PLA and measured for particle size (nm) and PDI. (A) A statistical F test to compare variance was performed, where particle size measurements for emulsification (n=18, SD=144.4) and microfluidics (n=19, SD=51.99) were statistically different (p<0.0001). (B) A statistical F-test to compare variance was performed, where PDI measurements for emulsification (n=18, SD=0.07031) and microfluidics (n=19, SD=0.0414) were statistically different (p<0.0315). Supplementary file3 (TIFF 4880 kb)

12248_2021_645_MOESM4_ESM.tiff

Figure S4. Correlation between particle size and zeta potential of iNPs on pro-inflammatory cytokine secretions. Data was obtained from experiments presented in Figure 5. Supplementary file4 (TIFF 103 kb)

12248_2021_645_MOESM5_ESM.tiff

Figure S5. Correlation between FRR of iNPs on pro-inflammatory cytokine secretions. Data was obtained from experiments presented in Figure 5. Supplementary file5 (TIFF 46 kb)

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Truong, N., Black, S.K., Shaw, J. et al. Microfluidic-Generated Immunomodulatory Nanoparticles and Formulation-Dependent Effects on Lipopolysaccharide-Induced Macrophage Inflammation. AAPS J 24, 6 (2022). https://doi.org/10.1208/s12248-021-00645-2

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