Dexamethasone Suppresses Oxysterol-Induced Differentiation of Monocytic Cells

Abstract

Oxysterol like 27-hydroxycholesterol (27OHChol) has been reported to induce differentiation of monocytic cells into a mature dendritic cell phenotype. We examined whether dexamethasone (Dx) affects 27OHChol-induced differentiation using THP-1 cells. Treatment of monocytic cells with Dx resulted in almost complete inhibition of transcription and surface expression of CD80, CD83, and CD88 induced by 27OHChol. Elevated surface levels of MHC class I and II molecules induced by 27OHChol were reduced to basal levels by treatment with Dx. A decreased endocytosis ability caused by 27OHChol was recovered by Dx. We also examined effects of Dx on expression of CD molecules involved in atherosclerosis. Increased levels of surface protein and transcription of CD105, CD137, and CD166 by treatment with 27OHChol were significantly inhibited by cotreatment with Dx. These results indicate that Dx inhibits 27OHChol-induced differentiation of monocytic cells into a mature dendritic cell phenotype and expression of CD molecules whose levels are associated with atherosclerosis. In addition, we examined phosphorylation of AKT induced by 27OHChol and effect of Dx, where cotreatment with Dx inhibited the phosphorylation of AKT. The current study reports that Dx regulates oxysterol-mediated dendritic cell differentiation of monocytic cells.

Figure 1

Effects of Dx on the transcription and surface expression of mDC markers induced by 27OHChol. (a) THP-1 cells (1 × 10⁶ cells/60 mm culture dish) were cultured for 48 h with 2.5 μg/mL of 27OHChol with or without 0.1, 1, or 10 μM of Dx. Transcription of CD80, CD83, and CD88 was analyzed by real-time PCR. Data are expressed as mean ± SD (n = 3 replicates/group). ^∗∗∗ P < 0.001 versus control; ^### P < 0.001 versus 27OHChol. (b) THP-1 cells (1 × 10⁶ cells/60 mm culture dish) were cultured for 48 h in the presence of 2.5 μg/mL of 27OHChol with or without 1 μM of Dx. Cells were immunostained with antibodies against CD80, CD83, and CD88 and analyzed by flow cytometry. Results are representative of three independent experiments.

Figure 2

Effects of Dx on expression of MHC class I and II molecules induced by 27OHChol. After incubation for 48 h with 27OHChol (2.5 μg/mL) with or without Dx (1 μM), the stimulated THP-1 cells (1 × 10⁶ cells/60 mm culture dish) were immunostained for MHC classes I and II. Fluorescence was analyzed by flow cytometry. Results are representative of three independent experiments.

Figure 3

Effects of Dx on functional alteration of monocytic cells induced by 27OHChol. After incubation for 48 h with 27OHChol (2.5 μg/mL) with or without Dx (1 μM) and PMA (200 nM), THP-1 cells were treated with 1 mg/mL of FITC-conjugated dextran for 1 h. The cells were analyzed by flow cytometry. Results are representative of three independent experiments.

Figure 4

Effects of Dx on expression of atherosclerosis-associated CD molecules induced by 27OHChol. (a) After incubation with 27OHChol (2.5 μg/mL) for 48 h with or without 0.1, 1, or 10 μM of Dx, transcription of CD105, CD137, and CD166 was analyzed by real-time PCR. Data are expressed as mean ± SD (n = 3 replicates/group). ^∗∗∗ P < 0.001 versus control; ^### P < 0.001 versus 27OHChol. (b) THP-1 cells (1 × 10⁶ cells/60 mm culture dish) were cultured for 48 h with 2.5 μg/mL of 27OHChol with or without 1 μM of Dx. The harvested cells were immunostained with antibodies against CD105, CD137, and CD166 and analyzed by flow cytometry. Results are representative of three independent experiments.

Figure 5

Effect of Dx on phosphorylation of AKT induced by 27OHChol. After incubation with 27OHChol (2.5 μg/mL) for 48 h with or without 0.1, 1, or 10 μM of Dx, lysates from the cells were analyzed with ELISA kit for p-AKT. Data are expressed as mean ± SD (n = 3 replicates/group). ^∗∗∗ P < 0.001 versus control; ^## P < 0.05 versus 27OHChol.