| Objective: To evaluate the effect of Interleukin-1βon the expression of Transforming Growth Factor-β1 in the bronchial epithelial cell and to investigate molecular mechanisms of the protective effect of Tripterygium Wilfordii Hook F(TWHF) on airway remodeling in Asthma. Methods: (1)Primary bronchial epithelial cell were obtained from rabbit trachea and cultured in vitro. After cell grew to confluence and were devided into groups. (2)The normal cultured bronchial epithelial cell were divided randomly into 4 groups: the normal control group, IL-1βtreatment A,B,C group. A, B, C groups were stimulated by IL-1β(0.1ng/ml, 1ng/ml, 10ng/ml, repectilvely) for 24 hours, the normal control group were incubated with equal serum free conditioned medium. Cell-coated dishes were attained, then the expression of TGF-β1 mRNA were measured by in site hybridization. A brownish yellow color was a positive sign of nucleic acid hybridization. (3)The normal cultured bronchial epithelial cell were divided randomly into 5 groups: the normal control group(incubated with equal serum free conditioned medium), IL-1βtreatment group(stimulated by IL-1β10ng/ml for 24 hours), both IL-1βand TWHF treatment A, B, C groups which were stimulated by IL-1β10ng/ml for 24 hours after the 0.5 hour of presence of TWHF(5μg/ml, 10μg/ml, 20μg/ml , respectively). In site hybridization and immunocyte-chemistry were preformed to detect the expression of TGF-β1 mRNA and protein, positive staining appeared brownish yellow. (4)The normal cultured bronchial epithelial cell were randomly divided into 4 groups: the normal control group (incubated with equal serum free conditioned medium), IL-1βtreatment group(stimulated by IL-1β10ng/ml for 24 hours), IL-1βtogether with TWHF treatment group (stimulated by IL-1β10ng/ml for 24 hours in the presence of TWHF 10μg/ml), TWHF treatment group(incubated with TWHF 10μg/ml). Then the total RNA of bronchial epithelial cell was extracted respectively for RT-PCR, in order to assess the expression of TGF-β1 mRNA. Result: (1) Cell Morphology and Identification. Bronchial epithelial cell grew in a typical epithelial fashion forming a fully confluent monolayer with cobblestone appearance and its characteristics were identified by positive immunostaining of keratin. (2) The effect of IL-1βon the expression of TGF-β1 mRNA. The expression of TGF-β1 mRNA in IL-1βtreatment A group(0.162±0.023), B group(0.318±0.013), C group(0.644±0.016)were significantly higher than that in the normal control group(0.098±0.020), P<0.01, and there were significant difference in A, B, C group in term of TGF-β1 mRNA expression , P<0.01. IL-1βtreatment cause dose-dependent stimulated of TGF-β1 mRNA expression. (3) The effect of TWHF on the expression of TGF-β1 mRNA and protein. The result of in situ hybridization: The expression of TGF-β1 mRNA in IL-1βtogether with TWHF treatment A group(0.452±0.030), B group(0.371±0.021), C group(0.194±0.005)were significantly lower than that in IL-1βtreatment group(0.644±0.016), P<0.01. TWHF treatment cause does-dependent inhibition of IL-1β-stimulated TGF-β1 mRNA expression. The result of immucytochemistry staining: The expression of TGF-β1 protein in IL-1βtogether with TWHF treatment A group(0.353±0.046), B group (0.245±0.034), C group(0.218±0.024)were significantly lower than that in IL-1βtreatment group(0.511±0.012), P<0.01. TWHF treatment cause does-dependent inhibition of IL-1β-stimulated TGF-β1 proteinexpression. A dose range of 5-20μg/ml, TWHF was effectively inhibitory to both TGF-β1 mRNA and protein. (4) The effect of IL-1βand TWHF on TGF-β1 mRNA expression by RT-PCR. IL-1βtreatment resulted in great increase in the expression of TGF-β1 mRNA, in contrasted to the normal control group, P<0.01. The expression of TGF-β1 mRNA in IL-1βtogether with TWHF treatment group were significantly lower than that in IL-1βtreatment group, P<0.01. There were no significant differences between the normal control group and alone TWHF treatment group in term of...
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