Extracellular vesicles released by triiodothyronine-stimulated mice dendritic cells spread the immune signature

NEGRETTI BORGA, DM; BLANCO A; DONADIO A; SOLER MF; DE PAUL AL; PELLIZAS CG; BRAVO MIANA R; MONTESINOS MM

Chicago

Congreso; 89th Annual Metting of the American Thyroid Association (ATA); 2019

American Thyroid Association

We reported that triiodothyronine (T3) triggers Th1 and Th17 adaptiveimmune profiles through mice dendritic cell (DC) activation, drivingpro-inflammatory and cytotoxic responses that were exploited in anantitumor DC-based vaccination protocol. Extracellular Vesicles(EVs) exhibit a crucial role in cellular communication and EVs secretedby DCs (EV-DC) may be involved in the amplification of theimmune response.Immature bone marrow DCs (iDC) obtained fromC57BL/6 mice were stimulated (or not, control) with T3 (5nM-18h).Secreted EV-DC from control and T3-stimulated DCs (EV-DC-T3)were isolated by differential ultracentrifugation (2,000g: 2K; 10,000g:10K and 100,000g: 100K). Morphological analysis was conducted byTransmission Electron Microscopy (TEM) and Dynamic Light Scattering(DLS), and specific molecular marker analysis by western blot(CD9 and CD81). A functional assay evaluated the syngeneic DCphenotypic and functional profile induced by EV-DC fromcontrol andT3-treated DCs (CD11c, MHCII, CD86 and IL-12 by FACS). Statisticalanalysis: Dunn?s Multiple Comparisons and Mann Whitneytests.TEM and DLS analysis allowed the morphological characterizationof EV-DC (control or T3-stimulated conditions) and revealedthat they did not exhibited differences between the experimentalgroups. A higher frequency of EV-DC >150nm for 2K and 10K-EVswas registered, whereas 100K exhibited more than 90% of EV-DCsized 30-150 nm. The EV-DC from both groups expressed CD9 andCD81. All EV-DC-T3 fractions increased the phenotypic maturationmarkers (MHCII, CD86) in recipient iDC (vs control). Besides, 10Kand 100K from both control and EV-DC-T3 activated syngeneic DCs(IL-12 production). The same effect was induced by 2K EV-DC-T3,but not by control EV-DC. The study allowed the characterization ofEV-DC-T3 and demonstrated their ability to induce syngeneic iDCactivation, spreading the T3-induced effects at the initiation of adaptiveimmunity. Therefore, these vesicles may be involved in theadaptive T response induced by T3 exposure to DCs. Further ongoingstudies will enlighten this issue, providing valuable tools for manipulatingthe immunogenic potential of DCs.