HIGH SODIUM CONCENTRATION MODULATES DENDRITIC CELLS IMMUNE FUNCTIONS

Abstract

Objective: Excessive Na+ intake is associated with the development of hypertension and increases skin sodium concentration [Na+] (up to 200 mM). The transfer of dendritic cells (DCs) from hypertensive to normotensive leads to the development of hypertension in animals. DCs modulate both the innate and adaptive immune response. However, the effect of high [Na+] on DCs remains unclear in humans. Design and method: DCs derived from human blood monocytes, differentiated for 6 days in RPMI 10% fetal calf serum (FCS) supplemented with GM-CSF (66 ng/ml) and IL4 (25 ng/ml) then stimulated during 48 h with LPS (2.5 or 50 ng/ml) at different [Na+] (140 vs 200 mM). We studied DCs morphological changes by confocal microscopy, analyzed cell viability, expression of CD25, CD83, CD86 and CD80 costimulatory markers, FITC-DEXTRAN endocytosis, reactive oxygen species (ROS) production and CCR7 chemokine receptor expression by flow cytometry. We studied DCs CCL19-driven chemotaxis using transwell migration assay with 8 μm pore size. We measured secreted cytokines (IL-12p70, IL-6, IL-23, IL-10, TGF-β) using ELISA. We also looked for MAP Kinase activity and SGK1 expression by western blot. Results: At high [Na+] of 200 mM as compared to normal [Na+] 140 mM, we found that DCs viability was maintained (over 84.5 ± 5%). DCs morphology changed towards a more elongated aspect. CD25, CD83 CD80 and CD86 expression significantly decreased (−67.7 ± 32.3%, −60.4 ± 14.5%, −25 ± 20.8%, −13.6 ± 10.3%, respectively, p < 0.0001). There were also less ROS production (−36.1 ± 12.3, P < 0.005) and a persistence of endocytosis capacity (+108.3 ± 44.6%, P < 0.005). CCR7 expression (−59.3 ± 15.3 %, P < 0.0001) and CCL19−driven chemotaxis (−49.7 ± 25.6%, P < 0.0005) index significantly decreased. Cytokines measurement showed a reduced secretion of IL−12p70, IL−6 and IL−23 (−78.5 ± 9%, −66.6 ± 9.2%,−90.4 ± 7.12%, respectively, p < 0.0001) and an increase of IL−10 and TGF−β (+111 ± 51%, +191.1 ± 110.4, respectively, P < 0.005). At high [Na+] reduced phosphorylation of p38 (−43.13 ± 25%, P < 0.03) leads to a higher ERK ½ protein phosphorylation (+30.572 ± 11.6%, P < 0.0078) and greater expression of SGK1 protein (+35.078 ± 11.5%, P < 0.015). Conclusions: High [Na+] concentration downregulates pro-inflammatory human DCs immune response to LPS and inhibits their migration towards lymph nodes, through MAP Kinase signaling pathway and SGK1-related mechanisms. However, the implication of these changes in the development of hypertension remains unknown.