This project, led by Philippe Frachet, Pascale Tacnet, and Quentin Depeyre (PhD student at UGA) in collaboration with other PIXEL members, aims to study specific molecular complexes involved in the interaction between a macrophage and its potential targets, such as apoptotic and cancerous cells.
We are particularly interested in the CD47-SIRPα immune checkpoint, which plays a key role in regulating phagocytosis. Our research focuses on the influence of membrane cholesterol in this process.
To achieve this, we use :
• The development of recombinant photoconvertible fluorescent probes for imaging membrane cholesterol,
• Confocal and super-resolution microscopy (dSTORM, PALM, and sptPALM),
• A dynamic approach to analyze, at the nanometric scale, the organization of contacts between macrophages and their targets during phagocytosis (phagocytic synapse).
This study builds on our previous research on apoptotic cell clearance (efferocytosis) and benefits from a close collaboration with Arnaud Millet’s team (Mechanobiology, Immunity, and Cancer at IAB-Grenoble).
Ultimately, one of the objectives is to propose new therapeutic approaches for cancer treatment targeting CD47-SIRPα.
Main methods : Eukaryotic cell culture, phagocytosis assay, flow cytometry, fluorescence microscopy. Recombinant photoconvertible fluorescent proteins engineering.
Highlights : (A) Single Particle Tracking (SPT) of CD47 at the surface of an apoptotic HeLa cell. CD47 was detected using an anti-CD47-AF647 antibody and imaged with a TIRF PALM/STORM microscope under low laser power. The trajectories were reconstructed, and the Mean Jump Distances (MJD) are color-coded (Imaging Platform of IBS). (B) Dual STORM detection of CD47 and soluble SIRPα binding at the surface of an apoptotic cell. The results show that SIRPα does not bind to apoptotic bodies, which are characterized by low cholesterol content. The corresponding bright-field (BF) image is shown on the right. Scale bar : 10 µm. Data from Dufour et al., Communication Biology, 2023.
References
Nanoscale imaging of CD47 informs how plasma membrane modifications shape apoptotic cell recognition. Samy Dufour, Pascale Tacnet-Delorme, Jean-Philippe Kleman, Oleksandr Glushonkov, Nicole Thielens, Dominique Bourgeois & Philippe Frachet Communication Biology (2023)6:207 | https://doi.org/10.1038/s42003-023-04558-y | www.nature.com/commsbio
Calreticulin Release at an Early Stage of Death Modulates the Clearance by Macrophages of Apoptotic Cells. Osman, R. ; Tacnet-Delorme, P. ; Kleman, J.-P. ; Millet, A. ; Frachet, P. Frontiers in Immunology 2017, 8. doi.10.3389/fimmu.2017.01034. Open
Molecular and cellular interactions of scavenger receptor SR-F1 with complement C1q provide insights into its role in the clearance of apoptotic cells Wicker-Planquart, C. ; Dufour, S. ; Tacnet-Delorme, P. ; Bally, I. ; Delneste, Y. ; Frachet, P. ; Housset, D. ; Thielens, N. M. Front. Immunol. 2020, 11. doi. 10.3389/fimmu.2020.00544. Open
Proteinase 3 Interferes With C1q-Mediated Clearance of Apoptotic Cells. Tacnet-Delorme P ; Gabillet J ; Chatfield S ; Thieblemont N ; Frachet P* ; Witko-Sarsat V. Front Immunol. 2018 April DOI : 10.3389/FIMMU.2018.00818. Open
Relative Contribution of C1q and Apoptotic Cell-Surface Calreticulin to Macrophage Phagocytosis. Verneret M, Tacnet-Delorme P, Osman R, Awad R, Grichine A, Kleman JP, Frachet P*. J Innate Immun. 2014 Feb 15
Role of C1q in efferocytosis and self-tolerance : links with autoimmunity. Book title : Autoimmunity - Pathogenesis of Autoimmune Diseases, Clinical Aspects and Therapy of Specific Autoimmune Diseases (ISBN 978-953-51-4188-4) edited by : Dr. Katerina Chatzidionysiou. Philippe Frachet*, Pascale Tacnet-Delorme, Christine Gaboriaud and Nicole Thielens. Intech 2015 DOI : 10.5772/60519