siRNA-mediated knockdown of VDR induces expression of LVSCC-A1C mRNA and protein. A) Comparison of LVSCC-A1C mRNA levels.
VDR suppression resulted in increased LVSCC-A1C mRNA expression, but the effects of VDR suppression on LVSCC-A1C were normalized after vitamin D treatment.* LVSCC-A1C mRNA levels from VDR-silenced neurons were statistically higher than in other groups (p = 0,015, p = 0,034, p = 0,002, p = 0,024, respectively). ** LVSCC-A1C mRNA levels were statistically lower than in VDR siRNA-treated group (p = 0,013). B) Detection of LVSCC-A1C protein by western blot. Although LVSCC-A1C protein increased in VDR-silenced neurons, vitamin D treatment decreased LVSCC-A1C expression to control levels. Beta actin was used as loading control. C) Comparison of LVSCC-A1C protein band intensities relative to Beta actin. Western blot results were consistent with mRNA results. The absolute intensities were measured using Image J software, and the relative intensities were calculated from the ratio of LVSCC-A1C to Beta actin absolute intensities. * LVSCC-A1C protein levels from VDR-silenced neurons were statistically higher compared to control groups (p<0.01, p<0.01, p<0.05, respectively). ** LVSCC-A1C protein levels from vitamin D-treated VDR-silenced neurons were statistically lower compared to the VDR siRNA-treated group (p<0.001). Control: Untreated control group; Vehicle: Transfection reagent-treated control group; Non target siRNA: Non-target siRNA-treated negative control group; Cyc B siRNA: Cyclophilin B siRNA-treated positive control group; VDR siRNA: VDR siRNA-treated group and VDR siRNA+Vitamin D: Following 12 hours of VDR siRNA treatment, groups were treated with vitamin D. Data are presented as a mean SD.