Involvement of the sphingolipid intracelular signaling pathway in the effects of T3 on dendritic cell activation

GIUSIANO L; ALAMINO VA; FOZZATTI L; SOLER MF; GIGENA N; MONTESINOS MM; PELLIZAS CG

Rio de Janeiro

Congreso; XVI Congress of the Latin American Thyroid Society; 2017

Latin American Thyroid Society (LATS)

Introduction: We reported that mice DCs, the main antigen presenting cells, express thyroid hormone receptor β1 with apreferential cytoplasmic localization and that physiological levels of T3 promote their maturation and their ability to directadaptive responses towards a Th1-type profile. Mechanistically, T3 effects involved activation of Akt independently of PI3K.Besides, sphingolipids revealed as key elements in signal transduction cascades, regulate events related to death and survival ofcells. The major bioactive sphingolipids include: sphingosine, sphingosine-1-phosphate (S1P), ceramide and ceramide-1-phosphate.Noteworthy, there is evidence that they are involved in the non-classical Akt activation. Objectives: We aim to furtherdisclose the molecular mechanism underlying the effects of T3 on DC functioning, in particular in the non-classical Akt activation.For this purpose, we characterized the intracellular pathway mediated by sphingolipids in DCs and initiated the studiesof its involvement in T3-mediated DC activation. Methods: Mice immature DCs were cultured for 18h with T3 (5 nM) in thepresence of chemical inhibitors of the sphingolipids pathway (GW4869: neutral sphingomyelinase, nSMAse and SKI: sphingosinekinases, SphK). DC viability was analyzed by Annexin V/7-AAD assay (flow cytometry). Intracellular and secreted IL-12production was assayed by flow cytometry and ELISA, respectively (IL-12 is a sensitive marker of T3 action on DCs). ThemRNAs expression coding the enzymes nSMAse, acid SMAse, SphK and ceramide kinase was evaluated through RT-PCR. Statistics:ANOVA/SNK test. Results: In this study we demonstrated that DCs express mRNA for the enzymes evaluated whichare essential for the balance between intracellular levels of interconvertible sphingolipids. Furthermore, exposure of maturingDCs to both inhibitors significantly suppressed their ability to produce IL-12 in response to T3. To note, neither GW4869 norSKI induced any significant effect on DC viability. Conclusion: Our results suggest that S1P is involved in T3 effect on DCmaturation in agreement with our previous report indicating an increase of Akt activation in T3-stimulated DC. These findingsprovide evidence for active involvement of the sphingolipid signaling pathway in the effects registered by T3 treatment to DCs.Considering the therapeutic impact we reported for T3-treated DCs, these results provide molecular tools to manipulate theimmunogenic potential of DCs.