Constitutional Nephrin Deficiency in Conditionally Immortalized Human Podocytes Induced Epithelial-Mesenchymal Transition, Supported by 𝜷-Catenin/NF-kappa B Activation: A Consequence of Cell Junction Impairment?

Figure 1

Phenotypic features of wild type (WT) human podocyte cell lines and 𝛼-SMA expression: effects of TGF-𝛽1 treatment. (a) The cells were grown as indicated in Section 2. Prior to the TGF-𝛽1 incubation, the cells were starved for 48 hr. The pictures show the cells after 48 hr of TGF-𝛽1 treatment used at 5 ng/mL. The optical images were obtained with a Will (Wetzelar) inverted microscopy at a 10x enlargement. (b) Western blot detection of 𝛼-SMA. Typically, 30 μg of total cell lysates was loaded on a 10% polyacrylamide gel. Western blot is described in Section 2. GAPDH expression was used to normalize protein loading. The anti-𝛼-SMA mouse monoclonal antibody was purchased from Sigma-Aldrich (St. Louis, MO, USA). The anti-GAPDH (sc-25778) rabbit polyclonal antibody was purchased from Santa Cruz Biotechnologies, (Santa Cruz, CA, USA). To quantify the protein bands, we scanned the blots and performed a densitometry analysis, using the software NIH ImageJ v. 6.4 (freeware, NIH, MD, USA). The figure shows a typical experiment of at least three performed under the same conditions.