Expression Regulation Of IL-13Ra2 By IL-13 And Its Relevance To Collagen Synthesizing

Purpose:According to the latest report of world health organization, fibrosis has become a major public health problem in the world wide. In the last decade, the morbidity and mortality of fibrosis present a significant increase. In addition, clinical fibrosis cases continue to grow. Here an increasing number of scientists focus on the research of causes and mechanism of fibrosis. More and more science academy and medical companies devoted to the development of new anti-fibrosis drug which most of them are based specific expression receptor and molecular cell surface mark.Present study shows that fibrosis could be found in wound healing, nonresolving inflammation, cancer and atherosclerosis. IL-13 could promote extracellular matrix deposition in fibrosis which characterized by over expression and production of collagen. IL-13 has two different receptors:IL-13Rαl and IL-13Ra2. IL-13 Ral binds IL-13 with low affinity by itself, while paired with IL-4Ra form a heterodimer complex with high affinity and transducing IL-13 signals. IL-13Ra2 binds IL-13 with high affinity as 5,000 times as IL-13 Rαl. It may not have the function of signal transduction based on the conception that its intracellular tail is very short. There are three phenotype of IL-13Ra2:intracellular-IL-13Ra2, membrane-IL-13Ra2 and soluble-IL-13Ra2. It is not very clear about the conversion mechanism of phenotype. Meanwhile, the function of IL-13Ra2 in IL-13-dependent fibrosis is controversial.Here we detect and analysis cell proliferation, migration and collagen formation of IL-13 on human hepatic stellate cells and lung fibroblasts. In addition, we demonstrate the expression regulation of IL-13Ra2 by IL-13 and its relevance to collagen synthesizing, which in order to provide new experimental basis and theoretical reference about clinical fibrosis research and treatment.Methods:1. Fibrosis vitro model construction:Human hepatic stellate cell LX-2 and lung fibroblasts HFL1 exposure to different concentration of IL-13;2. Cell proliferation and migration analysis:MTT method and Transwell technology;3. IL-4Rα, IL-13Rα1, intracellular-and soluble-IL-13Ra2 detect:RT-polymerase chain reaction (PCR) and ELISA;4. Imaging of intracellular-and membrane-IL-13Ra2:immunofluorescence;5. Collagen type I mRNA expression detection:RT-PCR6. Total collagen content detection:Hydroxyproline assay.Results:Hepatic stellate cells1. IL-13 can promote proliferation, migration, collagen type I mRNA expression and increases total collagen protein content of hepatic stellate cell (LX-2) in a dose dependent manner;2. IL-13 does not regulate IL-13Rαl and IL-4Ra expression level of hepatic stellate cell;3. IL-13Ra2 does not express in hepatic stellate cell (LX-2); IL-13 couldn't up regulate IL-13Ra2 expression.Lung fibroblasts1. Compared with the control group, there are no significant variation on proliferation, migration, collagen type I mRNA expressions and total collagen protein content of lung fibroblasts HFL1 exposure to low concentration range (5 to 20 ng/ml)of IL-13;2. High concentrations of IL-13 (50,100 ng/ml) can rely on the way to dose significantly promotes proliferation, migration, the type I collagen mRNA expressions and collagen protein content of lung fibroblasts;3. IL-13 does not regulate IL-13Ral and IL-4Rαexpression level of lung fibroblasts HFL1;4. IL-13Ra2 was undetectable in unstimulated lung fibroblasts. Expression of IL-13Ra2 was induced by IL-13 at low concentrations (5,10 and 20 ng/ml in our study); however, we have also detected that when exposure to IL-13 at high concentration, lung fibroblast cells were show diminished (50 ng/ml in our study) or even marginal (100 ng/ml in our study) expression of IL-13Ra2; intracellular IL-13Ra2 main distribute in cytosol area near the nucleus which has the highest relative content in three phenotypes of IL-13Ra2; concentrations of soluble sIL-13Ra2 were about 1/2 to 1/3 to intracellular IL-13Ra2; membrane-type IL-13Ra2 has the least content among three phenotypes.Conclusion:IL-13 can promote proliferation, migration and collagen synthesizing of hepatic stellate cell and lung fibroblast cell (don't change IL-13Ral and IL-4Ra expression level). But these effects could affect by IL-13Ra2. At the time that IL-13Ra2 does not expression, IL-13 could promote fibrosis in a dose-dependent manner. By contrast, IL-13 function would be inhibited by IL-13Ra2 expression.