The Role of the p38 MAPK Signaling Pathway in High Glucose-Induced Epithelial-Mesenchymal Transition of Cultured Human Renal Tubular Epithelial Cells Zhi-Mei Lv, Qun Wang, Qiang Wan, Jian-Gong Lin, Meng-Si Hu, You-Xia Liu, Rong Wang* Department of Nephrology, Provincial Hospital Affliated to Shandong University, JiNan, China Abstract Background: Epithelial-mesenchymal transition of tubular epithelial cells, which is characterized by a loss of epithelial cell characteristics and a gain of ECM-producing myofibroblast characteristics, is an essential mechanism that is involved in tubulointerstitial fibrosis, an important component of the renal injury that is associated with diabetic nephropathy. Under diabetic conditions, p38 MAPK activation has been reported in glomeruli and mesangial cells; however, studies on p38 MAPK in TECs are lacking. In this study, the role of p38 MAPK in AP-1 activation and in the EMT in the human proximal tubular epithelial cell line (HK-2) under high glucose concentration conditions is investigated. Methodology/Principal Findings: A vector for small interfering RNA that targets p38 MAPK was constructed; the cells were then either transfected with p38 siRNA or pretreated with a chemical inhibitor of AP-1 and incubated with low glucose plus TGF-b1 or high glucose for 48 h. Cells that were not transfected or pretreated and were exposed to low glucose with or without TGF-b1 or high glucose for 48 h were considered to be the controls. We found that high glucose induced an increase in TGF-b1. And high glucose-induced p38 MAPK activation was inhibited by p38 siRNA (P,0.05). A significant decline in E-cadherin and CK expression and a notable increase in vimentin and a-SMA were detected when exposed to low glucose with TGF-b1 or high glucose, and a significant raise of secreted fibronectin were detected when exposed to high glucose; whereas these changes were reversed when the cells were treated with p38 siRNA or AP-1 inhibitor (P,0.05). AP-1 activity levels and Snail expression were up-regulated under high glucose conditions but were markedly down-regulated through knockdown of p38 MAPK with p38 siRNA or pretreatment with AP-1 inhibitor (P,0.05). Conclusion: This study suggests that p38 MAPK may play an important role in the high glucose-induced EMT by activating AP-1 in tubular epithelial cells. Citation: Lv Z-M, Wang Q, Wan Q, Lin J-G, Hu M-S, et al. (2011) The Role of the p38 MAPK Signaling Pathway in High Glucose-Induced Epithelial-Mesenchymal Transition of Cultured Human Renal Tubular Epithelial Cells. PLoS ONE 6(7): e22806. doi:10.1371/journal.pone.0022806 Editor: Christos Chatziantoniou, Institut National de la Sante´ et de la Recherche Me´dicale, France Received January 14, 2011; Accepted July 7, 2011; Published July 29, 2011 Copyright: 2011 Lv et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was funded by National Natural Science Foundation of China (Grant 30971381). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: sdwangrong@sina.cn Introduction Glomerular mesangial expansion and podocyte loss are important early features of diabetic nephropathy, and tubuloin- terstitial injury and fibrosis are critical for the progression of diabetic nephropathy to kidney failure. It has been shown that tubulointerstitial fibrosis (TIF) is a more consistent predictor of functional impairment than glomerular damage. Accumulating evidence suggests that TECs play a pivotal role in TIF by undergoing EMT, which increases extracellular matrix (ECM) synthesis [1–3]. The EMT of tubular epithelial cells is proposed as an orchestrated, highly-regulated process that consists of four key steps [4]: (1) loss of epithelial cell adhesion; (2) de novo a-SMA expression and actin reorganization; (3) disruption of the tubular basement membrane; and (4) enhanced cell migration and invasion. The transforming growth factor-b (TGF-b) family of secreted factors regulates various biological processes, including cell proliferation, differentiation and apoptosis [5]. TGF-b can induce mesenchymal transdifferentiation in epithelial and endothelial cells through various signal pathways[6]. Thus in our study, to further confirm whether high glucose could induce EMT in TECs, we observed expression of TGF-b1 under high glucose conditions, given the fact that TGF-b is the key inducer of EMT [7,8]. p38 MAPK is a member of the MAPK family and is essential for the regulation of many cellular processes, including inflam- mation, cell differentiation, cell growth and cell death [9–11]. p38 MAPK mediates the signals that are relevant to thedevelopment of diabetic nephropathy. It is thought that p38 MAPK is a signal transducer in the underlying diabetic nephropathy pathways, and it has been proposed that the agents that inhibit the p38 MAPK signaling pathway may reduce the formation of the ECM in the glomerular mesangium and block the thickening of the glomerular basement membrane, thus preventing the development of diabetic nephropathy [12]. There is a wealth of data that supports the central role of the p38 MAPK signaling pathway in high glucose- PLoS ONE | www.plosone.org 1 July 2011 | Volume 6 | Issue 7 | e22806