Live Cell Imaging by Förster Resonance Energy Transfer Fluorescence to Study Trafficking of PLGA Nanoparticles and the Release of a Loaded Peptide in Dendritic Cells

• Main Authors: Mengshan Liu (Author), Chun Yin Jerry Lau (Author), Irene Trillo Cabello (Author), Johan Garssen (Author), Linette E. M. Willemsen (Author), Wim E. Hennink (Author), Cornelus F. van Nostrum (Author)
• Format: Book
• Published: MDPI AG, 2023-05-01T00:00:00Z.
• Subjects: article
• Online Access: Connect to this object online.

Summary

Our previous study demonstrated that a selected β-lactoglobulin-derived peptide (BLG-Pep) loaded in poly(lactic-<i>co</i>-glycolic acid) (PLGA) nanoparticles protected mice against cow's milk allergy development. However, the mechanism(s) responsible for the interaction of the peptide-loaded PLGA nanoparticles with dendritic cells (DCs) and their intracellular fate was/were elusive. Förster resonance energy transfer (FRET), a distance-dependent non-radioactive energy transfer process mediated from a donor to an acceptor fluorochrome, was used to investigate these processes. The ratio of the donor (Cyanine-3)-conjugated peptide and acceptor (Cyanine-5) labeled PLGA nanocarrier was fine-tuned for optimal (87%) FRET efficiency. The colloidal stability and FRET emission of prepared NPs were maintained upon 144 h incubation in PBS buffer and 6 h incubation in biorelevant simulated gastric fluid at 37 °C. A total of 73% of Pep-Cy3 NP was internalized by DCs as quantified using flow cytometry and confirmed using confocal fluorescence microscopy. By real-time monitoring of the change in the FRET signal of the internalized peptide-loaded nanoparticles, we observed prolonged retention (for 96 h) of the nanoparticles-encapsulated peptide as compared to 24 h retention of the free peptide in the DCs. The prolonged retention and intracellular antigen release of the BLG-Pep loaded in PLGA nanoparticles in murine DCs might facilitate antigen-specific tolerance induction.

Item Description

10.3390/ph16060818
1424-8247