Figure 1. VDR is required for 1,25D3 mediated down-regulation of Has2 mRNA and protein.

(A) RNA was isolated from KO240, WT145 and KO^hVDR cells treated with 100 nM 1,25D3 for 6, 12, 24, or 48 hours. Has2 mRNA in control and 1,25D3 treated samples was assessed by the ΔΔC_t method and values were normalized against 18S and expressed as fold change (1,25D3 vs control) for each cell line. Bars represent mean ± standard deviation, ^* p < 0.05 control vs 1,25D3 treated at each time point as evaluated by Student’s t test. (B) Immunofluorescence for HAS2 (green) in WT145 and KO^hVDR cells treated with 100 nM 1,25D3 or vehicle for 48 hours. Nuclei were stained with DAPI (blue). Images were acquired on a Leica DMI6000 microscope with a TCS SP5 confocal laser scanner using Leica Application Suite software. (C) Lysates from WT145 and KO^hVDR cells treated with 100 nM 1,25D3 for 48 hours were blotted with antibodies against HAS2.