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Figure 4 | SpringerPlus

Figure 4

From: Bisphenol A stimulates human prostate cancer cell migration via remodelling of calcium signalling

Figure 4

Effects of BPA on Orai1 and STIM1 expression in prostate cancer cells. (A), Total RNA was extracted and semi-quantitative RT-PCR experiments performed to study the expression of Orai1 (406 bp) mRNA using specific primers. β-actin mRNA expression (212 bp) was used as an internal standard. PCRs were carried out as described in Methods using 40 ng ARN equivalent cDNA from CTL and treated samples. H2O and samples without reverse transcriptase (not shown) were used as negative controls. Amplified fragments were resolved on 1.5% agarose gel by electrophoresis and visualized by EtBr staining.M, 1-kb DNA ladder, a molecular weight marker. (B, C, D), Effects of BPA on the ion channels' protein expression in LNCaP cells. Following the 48 h BPA treatments, total proteins were extracted and 10 μg proteins were analysed on 10% acrylamide gel (SDS-PAGE), transferred to PVDF membranes. Immuno-blots were then performed as described in Methods for the detection of Orai1 and STIM1. β-actin expression was used for the loading control of the samples. Experiments were performed at least twice in two independent cell cultures and representative figures are presented. The fold variation of the Orai1 and STIM1 proteins in different samples normalized to β-actin expression is shown under the panel. (E), Immunofluorescence studies of the effects of BPA on the expression of Orai1 in LNCaP. Cells were cultured for 48 h in the complete medium containing steroids (CTL + ST) or in the same medium without steroids (CS-RPMI, CTL -ST) or in CS-RPMI containing 1 nM BPA. (F), Expression of Orai1 protein in human PCa tissue. PCa tissue Cryosections were processed by Immunofluorescence for the detection of Orai1. S, Stroma; A, Acinus.

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