ISOLATION OF TUMOR EXOSOMES WITH IMUNOREGULATORY PROPERTIES FROM THE ASCITIC FLUID OF TUMOR BEARING MICE

MENAY F; COCOZZA F; DE TORO J; VENDRELL A; WALDNER C; MONGINI C

Mar del Plata

Congreso; Reunión conjunta de la Sociedad Argentina de Inmunología Clínica (SAIC) y la Sociedad Argentina de Inmunología; 2014

SAIC-SAI

Exosomes (Exo) are micro vesicles of 60-100 nm secreted by most cells. Tumor derived Exo may have either immuno stimulating or immunosuppressant properties. The aim of this work was to isolate and define the immuno-modulating characteristics of Exo derived from the ascitic fluid obtained from T-cell lymphoma bearing mice. BALB/c mice were inoculated ip. with 1,00E+06 LBC cells and after 20 ± 2 days ascites were collected by ventral puncture. Exo were isolated by a serial of differential centrifugation, ultrafiltration and ultracentrifugation at 100,000xg. The microvescicles were characterized by their floating density in a sucrose gradient and the presence of marker proteins Alix, Tsg, CD63, Hsp70 determined by Western blotting and flow cytometry. To study the immune properties in vivo, BALB/c mice were immunized twice with 20 μg /mouse with an interval of 7 days and 14 dpv were challenged with 1,00E+06 LBC. To assess the specificity of the immune response generated, mice that rejected LBC tumor, were re-challenged with 2,00E+05 cells of the breast adenocarcinoma LM3. Fifty percent of the mice immunized with Exo did not develop tumors, while the tumor incidence in control mice (non-immunized and challenged with the LBC cells) was 100% (n = 8). All LBC tumor free mice that were re-challenged with tumor cells developed the LM3 breast tumor. The humoral immune response induced was studied in sera obtained from immunized mice and compared with those from naïve mice, by dot and Western blot. All mice studied had high titre of serum antibodies against both Exo and LBC cells. Antibodies recognized  proteins of 45 and 50 kDa present in the Exo as determined by Western blot. We conclude that Exo are present in the ascitic  fluid of LBC tumor bearing mice and have immune-stimulating properties. The immune response elicited in hosts was specific  as it was sufficient to prevent tumor development but failed to prevent the progression of a non-related tumor.