Figure 5

Calcitriol regulates TGF-β-mediated EMT mRNA and protein expression changes in BEAS-2B cells. BEAS-2B cells were stimulated with the active form of Vitamin D, calcitriol (100 nM). After 24 hours, TGF-β1 (10 ng/ml) or TGF-β2 (10 ng/ml) was added to the cells for an additional 48 hours. Total RNA was isolated and assessed for the expression of A: E-cadherin, B: Snail, C: Vimentin, D: N-cadherin (left panels) by means of quantitative real-time PCR. Expression levels were normalized to the housekeeping 18 s and calculated as mean level of induction in comparison to 0.1% vehicle control cells. Data is presented as mean × SEM n =4. Total protein was isolated and assessed for the expression of A: E-cadherin, B: Snail, C: Vimentin, D: N-cadherin (right panels), and GAPDH by means of western blot analysis. Expression levels were normalized to the housekeeping gene GAPDH and calculated as mean level of induction in comparison to 0.1% vehicle control cells. Data is presented as mean × SEM n =3. Both qRT-PCR and western blot analysis were analyzed by one-way ANOVA *p <0.05, **p <0.01, and ***p <0.001 compared to control, #p <0.05, ##p <0.01, and ###p <0.001 compared to calcitriol, §p <0.05, §§p <0.01, and §§§p <0.001.