Design and Functional Characterization of HIV-1 Envelope Protein-Coupled T Helper Liposomes
Abstract
:1. Introduction
2. Materials and Methods
2.1. Animals
2.2. Env Trimer Purification
2.3. Nanoparticle Preparation
2.3.1. First-Generation Liposomes
2.3.2. Uncoupled Anionic T Helper Liposomes
2.3.3. Second-Generation Anionic T Helper Liposomes
2.3.4. Peptide Quantification via HPLC
2.3.5. T Helper VLPs
2.4. Isolation of Murine Primary Cells
2.4.1. B Cells
2.4.2. T Cells
2.4.3. Dendritic Cells (DCs)
2.4.4. Lymphocytes
2.5. Analysis of B Cell Activation and Proliferation
2.6. T Cell Proliferation by T Cell/Dendritic Cell Co-Culture
2.7. Ex Vivo DC Transfer
2.8. In Vitro Intrastructural Help
2.9. Antibody Staining of Liposomes
2.9.1. Immunogold EM Imaging of T Helper Liposomes
2.9.2. Liposomal Surface FACS
2.10. Statistical Analysis
3. Results and Discussion
3.1. First-Generation T Helper Liposomes Produced by Passive Peptide Entrapment
3.2. Functional Characterization of Uncoupled, Anionic Liposomes with Improved Peptide Encapsulation
3.3. Production of Second-Generation T Helper Liposomes by Covalent Coupling of Native-Like Env Trimers
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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First-generation T helper liposomes | ||
---|---|---|
| 1/~//OT2 | Uncoupled liposomes with zeta potential in a neutral range. OT2 peptides (black) were encapsulated by passive inclusion during hydration of the lipid film. |
| 1/~/JRFL// | Empty liposomes with zeta potential in a neutral range. JRFL NFL trimers (blue) were coupled to the liposomal surface by interaction of C-terminal His-tags with Ni-NTA lipids. |
| 1/~/JRFL/OT2 | T helper liposomes with zeta potential in a neutral range. OT2 peptides (black) were encapsulated by passive inclusion during hydration of the lipid film. JRFL NFL trimers (blue) were coupled to the liposomal surface by interaction of C-terminal His-tags with Ni-NTA lipids. |
Second-generation T helper liposomes | ||
| 2/-//OT2 | Uncoupled liposomes with an anionic zeta potential. OT2 peptides (black) were quantitatively encapsulated by an electrostatically driven approach. |
| 2/-/SUFO/OT2 | T helper liposomes with an anionic zeta potential. OT2 peptides (black) were encapsulated by an electrostatically driven approach. conSOSL.UFO.664 trimers (red) were covalently coupled to the liposomal surface by His-tag / Ni-NTA interaction followed by EDC / Sulfo-NHS crosslinking. |
Lentiviral T helper VLPs | ||
| SUFO-OT2-VLP | T helper VLPs that display conSOSL.UFO.750 trimers (SUFO.750, red) and 293T producer cell-derived proteins (black) on the surface and encapsulate both HIV-1 capsid proteins (grey) and OT2 peptides (black). |
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Damm, D.; Suleiman, E.; Theobald, H.; Wagner, J.T.; Batzoni, M.; Ahlfeld, B.; Walkenfort, B.; Albrecht, J.-C.; Ingale, J.; Yang, L.; et al. Design and Functional Characterization of HIV-1 Envelope Protein-Coupled T Helper Liposomes. Pharmaceutics 2022, 14, 1385. https://doi.org/10.3390/pharmaceutics14071385
Damm D, Suleiman E, Theobald H, Wagner JT, Batzoni M, Ahlfeld B, Walkenfort B, Albrecht J-C, Ingale J, Yang L, et al. Design and Functional Characterization of HIV-1 Envelope Protein-Coupled T Helper Liposomes. Pharmaceutics. 2022; 14(7):1385. https://doi.org/10.3390/pharmaceutics14071385
Chicago/Turabian StyleDamm, Dominik, Ehsan Suleiman, Hannah Theobald, Jannik T. Wagner, Mirjam Batzoni, Bianca Ahlfeld (née Kohlhauser), Bernd Walkenfort, Jens-Christian Albrecht, Jidnyasa Ingale, Lifei Yang, and et al. 2022. "Design and Functional Characterization of HIV-1 Envelope Protein-Coupled T Helper Liposomes" Pharmaceutics 14, no. 7: 1385. https://doi.org/10.3390/pharmaceutics14071385