| Asthma is a chronic inflammatory disease of the lung characterized by airway infiltration by lymphocytes and eosinophils. The clinical expression of asthma is associated to Th1/Th2 imbalance. Animal models and clinical studies have indicated an important role for Th2 lymphocytes producing IL-4, IL-5 and IL-13. Through releasing cytokines mentioned above, Th2 cells recruit and activate allergic response in mast cells and eosinophils. Among all the cytokines released by Th2 lymphocytes, IL-13 not only induces airway hyperresponsiveness in animal models of asthma, but also produces several of the structural changes seen in chronic asthma, including goblet cell hyperplasia, airway smooth muscle proliferation, and subepithelial fibrosis. In asthma, the expression of IL-13 is increased, but the underlying mechanism remains unclarified.As one member of the protein kinases family, mitogen-activated protein (MAP) kinases are major components of pathways controlling embryogenesis, cell differentiation, cell proliferation, and cell death. The extracellular signal-regulated kinases (ERKs), as one of the well-characterized subfamilies of the MAPKs, are widely expressed and are involved in the regulation of meiosis, mitosis, and postmitotic functions in differentiated cells. Previous study demonstrated that ERK signaling pathway might be involved in the airway inflammation in asthma. However, what remains unclear is that whether ERK signaling pathway directly involves in the expression of IL-13 in peripheral blood Th2 cells of asthmatic rats models. Previous studies have confirmed PD98059 as a specific inhibitor of ERK and EGF as an activator. In this study, we focused on the role of ERK signaling pathway on the expression of IL-13 in asthmatic rats' lymphocytes, and the results should be helpful for further understanding of the pathogenesis in asthma. Objective1. To investigate the changes and activation of extracellular signal-regulated kinase (ERK) signaling pathway in asthmatic lymphocytes.2. To explore the role of extracellular signal regulated kinase (ERK) in the expression of Th cytokine, interleukin 13 (IL-13) by lymphocytes in asthma.Methods and Materials40 SD rats were randomly divided into 2 groups,normal control and asthmatic group. Peripheral blood monocytes (PBMCs) were isolated and purified from blood of each asthmatic SD rats and divided into 5 groups:control,asthmatic cells, asthmatic cells stimulated with epidermal growth factor (EGF),or with ERK inhibitor PD98059, or with PD98059 and EGF together. The expression of p-ERK protein was observed by immunocytochemical staining, the expression of ERK mRNA was observed by RT-PCR, IL-13 protein in supernatants was measured by ELISA.Results1.Compared to the percentage of cells with positive p-ERK protein of normal control group (3.09%±0.27%), the percentage of cells with positive p-ERK protein of asthmatic group (15.63%±1.04%)was significantly increased (n=5, p<0.01).2.Compared to the expression of ERK mRNA of normal control group (0.2396±0.034), the expression of ERK mRNA of asthmatic group (0.7119±0.052) was significantly increased (n=5, p<0.01).3.Compared to the percentage of cells with positive p-ERK protein of asthmatic group (15.63%±1.04%), the percentage of cells with positive p-ERK protein of EGF group(30.60%±1.96%) was significantly increased (n=5, p<0.01), while the percentage of cells with positive p-ERK protein of PD98059 group (9.08%±0. 38%) was decreased significantly (n=5, p<0.01). The percentage of cells with positive p-ERK protein of asthmatic PD+EGF group (17.11%±1.51%) showed no significant difference compared to that of the asthmatic group (n=5, p>0.05).4.Compared to the expression of ERK mRNA of asthmatic group (0.7119±0.052), the expression of ERK mRNA of asthmatic EGF group (0.9606±0.075) was increased significantly, while the expression of ERK mRNA of asthmatic PD98059 group (0.4547±0.037) was decreased significantly (n=5, p<0.01). The expression of ERK mRNA of asthmatic PD+EGF group (0.7606±0.049) showed no significant difference compared to that of asthmatic group (n=5, p>0.05).5. Compared to the expression of IL-13 protein of normal control group (7.5±1.6), the expression of IL-13 protein of asthmatic group (21.6±3.7) was significantly increased (n=5, p<0.01).6. Compared to the expression of IL-13 protein of asthmatic group (21.6±3.7), the expression of IL-13 protein of asthmatic EGF group (62.4±5.29) was increased significantly (n=5, p<0.01), while the expression of IL-13 protein of asthmatic PD98059 group (13.4±1.34) was decreased significantly (n=5, p<0.01). The expression of IL-13 protein of asthmatic PD+EGF group (22.0±1.5) showed no significant difference compared to that of asthmatic group (n=5, p>0.05). 7. There was a significant positive correlation between the percentage of cells of nuclei stained positive for p-ERK and the expression of IL-13 proteins (n=5,r =0.892, p<0.001). And also there was a significant positive correlation between the A value of ERK mRNA expression and the IL-13 expression (n=5,r =0.843, p<0.005).ConclusionIn asthma the ERK expression and activation level were raised, and so was the protein level of IL-13. ERK signaling pathway may be involved in the increased Th2 cytokine IL-13 in asthma. |
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