A proteomic approach to immune-mediated epithelial-mesenchymal transition.

Increasing evidence suggests epithelial-mesenchymal transition (EMT) plays an important role in renal fibrosis. Initial renal injury enables the infiltration of mononuclear cells into the interstitium, and the resulting generation of inflammatory mediators that favour EMT may have a direct role in the development of renal fibrosis. The aim of this study was to investigate the proteome of renal tubular epithelial cells undergoing EMT in vitro. The human proximal tubular cell line (HK-2), exposed to conditioned medium from activated peripheral blood mononuclear cells (PBMC-CM), undergo phenotypic change, from an epithelial towards a fibroblastic phenotype, as evidenced by decreased E-cadherin and increased fibronectin protein expression. Further proteomic analysis, using 2-DE and Progenesis software, revealed the down-regulation of 4 proteins and up-regulation of 23 proteins. MS analysis allowed the positive identification of 15 differentially expressed proteins, including annexin A2, adipocyte plasma membrane-associated protein, T-complex protein 1, reticulocalbin-1 precursor and moesin among others. Western blotting and quantitative real-time PCR confirmed the increase in annexin A2 at the protein and gene level, respectively. Since annexin A2 and S100A6 exist as complexes in B-1 cells, we investigated the S100A6 gene expression further and show an increased expression in HK-2 cells following exposure to activated PBMC-CM. Therefore, we have identified several potential proteins that could play key roles in immune-mediated EMT.