The effects of 1,25(OH)2 D3 treatment on immune responses and intracellular metabolic pathways of bone marrow-derived dendritic cells from lean and obese mice.

Vitamin D affects differentiation, maturation, and activation of dendritic cells (DCs). Obesity-related immune dysfunction is associated with metabolic changes in immune cells. Objectives of the study are to investigate the effects of vitamin D and obesity on immune responses and markers related to immunometabolism of bone marrow-derived dendritic cells (BMDCs). Bone marrow cells (BMCs) were isolated from lean and obese mice, and BMDCs were generated by culturing BMCs with rmGM-CSF. BMDCs were treated with 1 or 10 nM of 1,25-dihydroxyvitamin D3 (1,25(OH)2 D3 ), and maturation was induced by LPS (50 ng/ml) stimulation for 24 hr. Cell phenotypes, cytokine productions, and expression of proteins and genes involved in Akt/mTOR signaling pathway and glycolytic pathway were determined. 1,25(OH)2 D3 treatment inhibited differentiation of BMDCs (CD11c+ %), expression of phenotypes related with DC function (MHC class II and CD86) and production of IL-12p70 in both lean and obese mice. The expression of PD-L1 and the ratio of IL-10/IL-12p70 were increased by 1,25(OH)2 D3 . With 1,25(OH)2 D3 treatment, Akt/mTOR signaling pathway was suppressed, and expression of genes related to glycolysis (Glut1, Pfkfb4, and Hif1A) was increased. The upregulation of glycolysis-related genes observed with 1,25(OH)2 D3 treatment seems to be associated with the induction of tolerogenic features of BMDCs from lean and obese mice, and Hif1A seems to have a potential role in conveying the effect of 1,25(OH)2 D3 on glycolysis.

The effects of dietary vitamin D supplementation and in vitro 1,25 dihydroxyvitamin D3 treatment on autophagy in bone marrow-derived dendritic cells from high-fat diet-induced obese mice.

Obesity is associated with the dysregulation of vitamin D metabolism and altered immune responses in bone marrow-derived dendritic cells (BMDCs). Vitamin D can affect the differentiation, maturation, and activation of dendritic cells (DCs) and regulate autophagy via vitamin D receptor signaling. Autophagy was shown to be involved in the functions of DCs. We investigated the effects of dietary vitamin D supplementation and in vitro 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) treatment on autophagy in BMDCs from control diet (CON)-fed lean and high-fat diet (HFD)-induced obese mice. C57BL/6 male mice were fed CON or HFD with 10% or 45% kcal fat, respectively, supplemented with 1,000 or 10,000 IU vitamin D/kg diet (vDC or vDS) for 12 weeks. BMDCs were generated by culturing bone marrow cells from the mice with 20 ng/mL rmGM-CSF and treated with 1 nM 1,25(OH)2D3. Maturation of BMDCs was induced by lipopolysaccharide (50 ng/mL) stimulation. Treatment with 1,25(OH)2D3 inhibited the expression of phenotypes related to DC function (MHC class Ⅱ, CD86, CD80) and production of IL-12p70 by BMDCs from control and obese mice, regardless of dietary vitamin D supplementation. LC3Ⅱ/Ⅰ and VPS34 protein levels increased, and p62 expression decreased, after 1,25(OH)2D3 treatment of the BMDCs in CON-vDC only. Vdr mRNA levels decreased following 1,25(OH)2D3 treatment of BMDCs in the HFD-vDC. In conclusion, autophagy flux was increased by 1,25(OH)2D3 treatment of the BMDCs in CON-vDC but not in the HFD-vDC group. This suggests that the decreased expression of Vdr following 1,25(OH)2D3 treatment might have affected autophagy flux in BMDCs from obese mice.