Fig. 3.

SK-CO15 cells express Na⁺/H⁺ exchanger 3 (NHE3). A: expression of NHE1, NHE2, NHE3, and NHE8 in Caco-2 and SK-CO15 cells were determined by RT-PCR. A representative figure of 3 independent experiments is shown. Quantitative real-time RT-PCR of SK-CO15 (B) and Caco-2 (C) was performed to determine relative levels of NHE1, NHE2, and NHE3 transcripts. Threshold cycle values of NHE1–3 were normalized to that of β-actin. The relative abundance of NHEs was then calculated by normalizing to the expression level of NHE1 mRNA. D: NHE3 protein expression was determined using anti-NHE3 antibody EM450 in SK-CO15, Caco-2, Caco-2bbe (C2b), C2b/NHE3V, and OKP cells. β-actin was used as a loading control. E: SK-CO15 cells were transiently transfected with control shRNA (shCon) or NHE3-specific shRNA (shNHE3). The specificity of EM450 was determined by Western blot analysis, which showed a decrease in NHE3 protein band intensity in cells transfected with shNHE3. Cells transfected with pcDNA3.1/NHE3V were used as a control. β-actin was used as a loading control (n = 3).