Characterization of extracellular vesicles released by triiodothyronine-stimulated mice dendritic cells (DCS): rol in paracrine dc activation

FLORENCIA SOLER; ANA CAROLINA DONADIO; ROCÍO DEL CARMEN BRAVO MIANA; ANA LUCÍA DE PAUL; CLAUDIA PELLIZAS; DANA MARÍA NEGRETTI BORGA; GIUSIANO LUCILA; MARÍA DEL MAR MONTESINOS

Mar del Plata

Congreso; LXIII Reunión Anual de la Sociedad Argentina de Investigación Clínica (SAIC); 2018

Sociedad Argentina de Investigación Clínica (SAIC)

We provided evidence of triiodothyronine (T3) stimulation of mice dendritic cells (DCs), driving pro-inflammatory and cytotoxic responses, exploited in an antitumor DC-based vaccination protocol. Extracellular Vesicles (EVs) exhibit a crucial role in cellular communication and EVs secreted by DCs (EVs-DCs) may be involved in the amplification of the immune response. Our aim was to characterize the populations of EVs-DCs and assess their role in paracrine DC communication after T3 exposition. Immature bone marrow DCs (iDC) were obtained from WT C57BL/6 mice and stimulated (or not) with T3 (5nM-18h, DCs-T3). Secreted EVs-DCs from iDCs and DCs-T3 (EVs-DCs-T3) were isolated by differential ultracentrifugation at 2,000g (2K, large EVs); 10,000g (10k, Microvesicles); and100,000g (100K, small EVs: sEVs). Morphological analysis of EVs was conducted by Transmission Electron Microscopy (TEM) and dynamic light scattering (DLS), and molecular characterization of EVs by western blot analysis (CD63 and CD81). A functional assay evaluated the syngeneic DC profile induced by treatment with EVs-DCs-T3 (markers of DC maturation: MHCII, CD86, and CD40, Flow Cytometry-FACS; and IL-12, FACS and ELISA). Statistical analysis: ANOVA-SNK. Data obtained from TEM analysis of the different fractions from both, iDCs and DCs-T3, showed the presence of secreted EVs. Size profile analysis revealed a significant higher frequency of 150-600 nm for 2K and 10K, whereas 100k exhibited more than90% (p