Sphingosine kinase 1 is involved in triiodothyronine effects in murine dendritic cells and the driven adaptive immunity

NEGRETTI- BORGA DANA MARÍA; BLANCO ANTONELLA; PIRES TEIXEIRA MARIANA; ALAMINO VANINA ALEJANDRA; PUENTES ELIDA NAHIR; SOLER MARÍA FLORENCIA; DONADIO ANA CAROLINA; CHRISTOPHER J. CLARKE; MONTESINOS MARÍA DEL MAR; YUSUF A. HANNUN; PELLIZAS CLAUDIA GABRIELA

Curitiba

Congreso; XIX Latin American Thyroid Society Congress; 2023

Latin American Thyroid Society

Triiodothyronine (T3) is the biologically active thyroid hormone (TH), essential for homeostatic control. Dendritic cells (DCs) arehighly phagocytic, specialized antigen presenting cells. They orchestrate the adaptive immune response through T-cell activation,leading to the production of pro-inflammatory cytokines. DCs additionally control immunity through the generation of regulatory Tcells. Our research group reported that mice DCs express TH receptor β1 (TRβ1) and that physiological levels of T3 promotes DCs’maturation, survival, and ability to direct pro-inflammatory responses with Th1, Th17 and cytotoxic profiles, restraining tolerogenicsignals. These findings were successfully exploited in T3-activated DC-based antitumor vaccines against melanoma and colon carcinomain mice. T3 effects in DCs are mainly triggered by non-genomic mechanisms involving TRβ1, Akt and NF-kB. In turn, Sphingolipidsand their synthetic enzymes have been involved in inflammation, aging and cancer. Sphingosine-1-phosphate (S1P) is produced bySphingosine Kinase 1 (SK1) and 2. Although this pathway is involved in many pro-inflammatory conditions, little is known aboutits role in innate immune cells. Moreover, the putative role of this pathway in the reported pro-inflammatory effects induced byT3 in DCs, is of high interest. The aim of this study was to evaluate the role of SK1 in T3-stimulated DCs, and the driven adaptiveimmunity. DCs differentiated from bone marrow precursors from Wild-Type (WT) and SK1 Knockout (KO) C57BL/6 mice werestimulated with T3 (10 nM, T3-DC) for 18h. Immature DCs (iDC) from WT mice were incubated with PF-543 (SK1 inhibitor, 100nM), and 30 min later with the T3 stimuli (PF-T3-DC). After 30 min, p-Akt and total Akt were analyzed by Western Blot. Allogenicsplenocytes isolated from BALB/c mice were co-cultured with T3-DC or PF-T3-DC (exposed to T3 for 18h), for 3 days. Viability andproliferation were evaluated by FACS. Cytokines were measured by FACS and ELISA. Statistical significance of differences betweenmeans was determined by Two-way ANOVA/Tukey test, and paired t test (p < 0.05, statistically significant). Results showed thatintracellular IL-12 production was increased in T3-DC from SK1-KO vs. WT mice(p < 0.0001). Accordingly, IL-12 secretion washigher in PF-T3-DC vs. T3-DC (p < 0.005). Of note, DC viability was not modified by PF-543. In turn, SK1 inhibition reduced p-Aktin T3-DC (p < 0.005). Splenocytes proliferation, as well as IFN-γ and IL-17 production and secretion, markers of pro-inflammatoryadaptive responses, were modified in the co-culture with PF-T3-DC (vs. T3-DC, p < 0.05). Our results revealed for the first time thatthe Sphingolipid intracellular pathway is involved in T3-DC activation. The immunomodulation exerted by SK1 on T3-DC and thedriven adaptive response provide the first insights into a novel role of Sphingolipids in the immune-endocrine crosstalk, which will beintimately unveiled by further research.