Effects of 1,25(OH)2D3 in immune response regulation of systemic lupus erithematosus (SLE) patient with hypovitamin D

Abstract

Vitamin D deficiency has been associated with pathogenesis of autoimmune diseases including SLE; however, there were still lack of data about the effects of administration of vitamin D in immune regulation in SLE patients. The aim of this study was to investigate the effects of calcitriol/1,25(OH)2D3 on dendritic cells maturation and Th17 and Treg cells activation in SLE patients with hypovitamin D. The monocytes and lymphocytes of five SLE patients with hypovitamin D were divided into 4 groups, P0 (0 nM/control), P1 (1 nM), P2 (10 nM), and P3 (100 nM) as cultured samples. Flowcytometry analysis was used to evaluate dendritic cell maturation (the percentage of CD40, CD86, and HLA-DR expression) and the amount of Th17 and Treg cells (the percentage of Th17 and Treg cells). Cytokines production of IL-12, IL-17A, and TGF-β measured by ELISA. This study showed significant differences in CD40, CD86, HLA-DR expressions, and Th17 percentage in 10 nM of 1,25(OH)2D3 compared to that of control. For cytokines secretion, there was also significant difference between IL-12p70 and IL-17A levels in 10 nM of 1,25(OH)2D3 compared to that of control. The 1,25(OH)2D3 increased Treg cells and TGF-β level but not significant. Our study concluded that 1,25(OH)2D3 inhibited dendritic cells maturation and Th17 cells activation in SLE patients. The 1,25(OH)2D3 increased Treg cells but not significant.

Keywords: 25(OH)2D3]; Calcitriol [1; Th17 cells; Treg cells; dendritic cell; systemic lupus erythematosus.

Figures

Figure 1

Expression of CD40, CD86, dan HLA-DR of DC inhibited by 1,25(OH)2D3. Immature DC taken from monocytes culture supplemented with GM-CSF dan IL-4 for 6 days. On day 7, 1x10-6 cells/well were added with 1,25(OH)2D3 with different doses: 0, 1, 10, 100 nM, then cultured in medium with TNF-α for 4 days. On day 10, cells were harvested and stained with monoclonalantibodies PerCp/Cy5.5 anti-human CD40, FITC anti-human CD86, dan PE anti-human HLA-DR prior to flow cytometry (FACScalibur) analysis. A. Expression of CD40 percentage with different doses of 1,25(OH)2D3. B. Expression of CD86 percentage with different doses of 1,25(OH)2D3. C. Expression of HLA-DR percentage with different doses of 1,25(OH)2D3.

Figure 2

Expression of CD40, CD86, and HLA-DR in DC. There were reduced mean of CD40, CD86, and HLA-DR among P1, P2, dan P3 compared with P0. P2 was the lowest *Significant (p2D3 1 nM and 10 nM showed significant difference in CD40 expression compared to control. B. Treatment of 1,25(OH)2D3 10 nM showed significant difference in CD86 expression compared to control. C. Treatment of 1,25(OH)2D3 10 nM dan 100 nM showed significant difference in HLA-DR expression compared to control.

Figure 3

The difference of IL-12p70 secretion level in treatment groups of 1,25(OH)2D3 in DC. There were reduced level of IL-12p70 in groups P1, P2, and P3 compared with P0 (control). Group P2 (10 nM of 1,25(OH)2D3) showed significant difference compared to control.

Figure 4

The 1,25(OH)2D3 increased Th17 CD4 T cells. Expression of Th17 percentage after 1,25(OH)2D3 treatment with different doses (0, 1, 10, 100 nM) in CD4 T cells of patients with SLE. Upper right quadrant showed percentage of Th17 cells.

Figure 5

Expression of Th17 cells in CD4 T cells. There were reduced percentage of Th17 cells among groups P1, P2, and P3 compared with P0. P2 was the lowest. *Significant (p

Figure 6

The difference of IL-17A secretion level in treatment groups of 1,25(OH)2D3 in CD4 T cells. There was a decrease in IL-17A level in groups of P1, P2, dan P3 compared with P0 (control). Group P2 was the lowest (p=0.047). *Significant (p<0.05) compared with control.

Figure 7

The 1,25(OH)2D3 inhibited Treg CD4 T cells. Expression of Treg percentage after 1,25(OH)2D3 treatment with different doses (0, 1, 10, 100 nM) in CD4 T cells of patients with SLE. Upper right quadrant showed percentage of Th17 cells.

Figure 8

Expression of Treg cells in CD4 T cells. There was an increase percentage of Treg cells between group P1 and P3 compared with P0. There were no significancies compared to control.

Figure 9

The difference of TGF-β secretion in treatment groups of 1,25(OH)2D3 in CD4 T cells. There was a difference in TGF-β level among groups of P1, P2, dan P3 compared with P0 (control). There were no significancies compared to control.