Characterization of exosomes derived from a murine T-cell lymphoma

HERSCHLIK, LETICIA; GRAVISACO MARÍA JOSÉ; DI SCIULLO MARÍA PAULA; VENDRELL A; WALDNER CLAUDIA; MONGINI CLAUDIA

Buenos Aires

Congreso; First French-Argentine Immunology Congress; 2010

Exosomes are 60 to 100 nm lipid bilayer vesicles with a density of 1.13 to 1.19 g/ml comprising an enclosed compartment topologically equivalent to the cytoplasm, and with the extracellular domains of transmembrane proteins at their surface. Exosomes originate in the late endosomal compartment by inward budding of the limiting endosomal membrane, thereby generating intracellular multivesicular bodies released in the extracellular medium upon constitutive or induced fusion of MVBs and plasma membranes. LBC cell line is a murineT-cell lymphoma expressing MHC I, CD8, CD24, CD16 and TCR. The aim of this work was to characterize exosomes derived from the T-cell lymphoma LBC to be used as a cell free immunogen. Supernatants of LBC cells cultured until a density of 1,50 to 2,00 E+06/ml were collected and sequentially centrifuged at 300 g then at 1200 g . and finally exosomes were then pelleted at 100,000 g and washed once in PBS. Traditional cultures of LBC cells gave an average yield of 0.78 ug±0.14ug of protein (measured by Lowry assay) per 1,0 E+06 LBC cells. Exosome preparations, analyzed by electron microscopy, displayed the characteristic exosomal cup-shaped morphology. Beads coated with exosomes were stained with conjugated monoclonal antibodies, and analyzed by flow cytometry. Exosomes derived from LBC lymphoma abundant in MHC Class I, the heat stable protein CD24 and the heat shock protein Hsp 90. Other immunologically important molecules detected included CD8 but there was no detectable Hsp 60, and Hsp70, proteins present intracytoplasmatically in LBC cells. Our results demonstrated the presence of proteins with immunological relevance on exosomes derived from LBC cells. Expression of MHC I, the heat stable antigen CD24 or the heat shock protein Hsp90, molecules involved in antigen presentation and ligand-receptor interactions can lead to the activation or modulation of various immune responses and therefore might be useful in developing cancer immunotherapies.