An immortalized rat pancreatic stellate cell line RP-2 as a new cell model for evaluating pancreatic fibrosis, inflammation and immunity

Background Pancreatic stellate cells (PSCs) play a critical role in the pathogenesis of pancreatic fibrosis and have emerging functions as progenitor cells, immune cells or intermediaries in pancreatic exocrine secretion. Increasing evidence has shown that desmin as an exclusive cytoskeleton marker of PSC is only expressed in part of these cells. This study was to establish a desmin-positive PSC cell line and evaluate its actions on pancreatic fibrosis, inflammation and immunity. Methods The presence of cytoskeletal proteins, integrin α 5 β 1 or TLR4, was determined by immunocytochemistry while the production of desmin, collagen I, MMP-1, MMP-2, TIMP-2, or CD14 was evaluated by Western blotting. The levels of desmin, collagen I, IL-1 and IL-6 mRNA were determined by real-time quantitative PCR. The secretion of cytokines was detected by ELISA. Cell function was assessed using adhesion, migration, or proliferation assays. Results A stable activated rat PSC cell line (designated as RP-2) was established by RSV promoter/enhancer-driven SV40 large T antigen expression. RP-2 cells retained typical PSC properties, exhibited a myofibroblast-like phenotype and persistently produced desmin. The cells produced collagen I protein, matrix metalloproteinases and inhibitors thereof. RP-2 cells demonstrated typical PSC functions, including proliferation, adherence, and migration, the latter two of which occurred in response to fibronectin and were mediated by integrin α 5 β 1 . TLR4 and its response genes including proinflammatory cytokines (IL-1, IL-6, TNF-α) and chemotactic cytokines (MCP-1, MIP-1α, Rantes) were produced by RP-2 cells and activated by LPS. LPS-induced IL-1 or IL-6 mRNA expression in this cell line was fully blocked with MyD88 inhibitor. Conclusion RP-2 cells provide a novel tool for analyzing the properties and functions of PSCs in the pathogenesis of fibrosis, inflammation and immunity in the pancreas.