Vitamin D receptor silencing impairs the 1,25-dihydroxyvitamin D3-stimulated phosphorus absorption by suppressing NaPi-IIb gene expression in primary duodenal epithelial cells of broilers.

Our previous study detected the potential regulatory role of vitamin D receptor (VDR) in 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3)-stimulated phosphorus (P) absorption and the type IIb sodium-phosphate cotransporter (NaPi-IIb) gene expression in the ligated duodenum of broilers. In the present study, three experiments were carried out to further investigate whether and how VDR participated in 1,25-(OH)2D3-stimulated P absorption and NaPi-IIb transcription in broiler primary duodenal epithelial cells (BPDECs). In experiment 1, the BPDECs were treated with different concentrations (0, 30, 60, 120 or 240 pmol/L) of 1,25-(OH)2D3 for 87 min. In experiment 2, three VDR-specific siRNAs were designed and transfected into BPDECs to screen for the most effective siRNA against VDR. In experiment 3, with or without RNA interference against VDR, the cells were incubated in the medium with an added 1,25-(OH)2D3 level of 0 or 75 pmol/L for 87 min. The results showed that in experiment 1, as supplemental 1,25-(OH)2D3 levels increased, among the affected parameters (P ≤ 0.005), P absorption amount and rate, the mRNA and protein expression levels of VDR and NaPi-Ⅱb increased quadratically (P ≤ 0.009), and 75 pmol/L of 1,25-(OH)2D3 was recommended to be the optimal supplemental level for promoting P absorption, VDR and NaPi-IIb gene expression levels in BPDECs. In experiment 2, the si-326 was relatively the most effective in suppressing (P < 0.05) VDR mRNA expression level. In experiment 3, supplemental 1,25-(OH)2D3 enhanced (P < 0.05) the P absorption amount and rate, up-regulated (P < 0.05) mRNA and protein expression levels of NaPi-IIb and VDR, and increased (P < 0.05) the enrichment of VDR bound to NaPi-Ⅱb DNA promoter region in BPDECs. However, VDR silencing inhibited (P < 0.05) them, especially the promoting effect of 1,25-(OH)2D3 on the P absorption amount and rate as well as the NaPi-IIb protein expression level. Therefore, VDR mediates the 1,25-(OH)2D3-stimulated P absorption by promoting the NaPi-IIb gene expression via its transcriptional activity in BPDECs.