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Detection of the interactions of tumour derived extracellular vesicles with immune cells is dependent on EVā€labelling methods

1 dƩc. 2023Journal of Extracellular Vesicles

DOI : 10.1002/jev2.12384

Auteurs

Luisa Loconte, Davinia Arguedas, Rojbin El, Alix Zhou, Anna Chipont, Lea Guyonnet, Coralie Guerin, Ester Piovesana, JosĆ© Luis VĆ”zquezā€Ibar, Alain Joliot, Clotilde ThĆ©ry, Lorena MartĆ­nā€Jaular

RƩsumƩ

Abstract

Cellā€cell communication within the complex tumour microenvironment is critical to cancer progression. Tumorā€derived extracellular vesicles (TDā€EVs) are key players in this process. They can interact with immune cells and modulate their activity, either suppressing or activating the immune system. Deciphering the interactions between TDā€EVs and immune cells is essential to understand immune modulation by cancer cells. Fluorescent labelling of TDā€EVs is a method of choice to study such interaction. This work aims to determine the impact of EV labelling methods on the detection by imaging flow cytometry and multicolour spectral flow cytometry of EV interaction and capture by the different immune cell types within human Peripheral Blood Mononuclear Cells (PBMCs). EVs released by the tripleā€negative breast carcinoma cell line MDAā€MBā€231 were labelled either with the lipophilic dye MemGlowā€488 (MGā€488), Carboxyfluorescein diacetate, succinimidyl ester (CFDAā€SE) or through ectopic expression of a MyrPalmā€superFolderGFP reporter (mpā€sfGFP), which incorporates into EVs during their biogenesis. Our results show that these labelling strategies, although analysed with the same techniques, led to diverging results. While MGā€488ā€labelled EVs incorporate in all cell types, CFSEā€labelled EVs are restricted to a minor subset of cells and mpā€sfGFPā€labelled EVs are mainly detected in CD14+ monocytes which are the main uptakers of EVs and other particles, regardless of the labelling method. Furthermore, our results show that the method used for EV labelling influences the detection of the different types of EV interactions with the recipient cells. Specifically, MGā€488, CFSE and mpā€sfGFP result in observation suggesting, respectively, transient EVā€PM interaction that results in dye transfer, EV content delivery, and capture of intact EVs. Consequently, the type of EV labelling method has to be considered as they can provide complementary information on various types of EVā€cell interaction and EV fate.