Vitamin D receptor (VDR) knock down experiments through siRNA. (A) MTT proliferation assay showing the effect of 100 nM 1,25D or vehicle after 72 h in cells transfected with a negative control siRNA (Neg-siRNA) and cells transfected with specific siRNA for VDR (VDR-siRNA). Results are expressed as % of control (Neg-siRNA transfected cells grown without exposure to 1,25D). (B) Q-PCR of VDR gene expression in cells transfected with neg-siRNA or VDR-siRNA normalized to GAPDH in the absence of 1,25D. (C) Western-blot for VDR and α-tubulin (loading control) in lysates from cells transfected with the negative siRNA or the VDR-siRNA. Band intensities are indicated below each lane. (D) Q-PCR of CYP24A1 gene expression in cells transfected with Neg-siRNA or VDR-siRNA. Data are mean ± SD of fold change in CYP24A1 expression in 1,25D treated cells vs. vehicle treated control. Statistical significance was assessed by two-tailed unpaired Student's t-test. Differences were considered statistically significant when p < 0.01 (*). p > 0.01 was considered not significant (ns).