Original Articles: Mechanisms Of Allergy
IL-4 and IL-13 induce myofibroblastic phenotype of human lung fibroblasts through c-Jun NH2-terminal kinase–dependent pathway,☆☆

https://doi.org/10.1067/mai.2001.114702Get rights and content

Abstract

Background: Myofibroblasts play a role in the airway remodeling response of bronchial asthma. IL-4 and IL-13 are possibly involved in the airway remodeling response by inducing extracellular matrix production by fibroblasts. However, the roles of these cytokines in inducing the phenotypic modulation of human lung fibroblasts (HLFs) to myofibroblasts and the intracellular signal have not been determined. Objective: We examined the effect of IL-4 and IL-13 on inducing the phenotypic modulation of HLFs to myofibroblasts characterized by α-smooth muscle actin and examined the role of the mitogen-activated protein (MAP) kinase superfamily in inducing the myofibroblastic phenotype of the HLF to clarify these issues. Methods: Phosphorylation and activities of c-Jun NH2-terminal kinase (JNK), p38 MAP kinase, and extracellular signal-regulated kinase (Erk) were examined by using Western blotting and in vitro kinase assay. Expression of α-smooth muscle actin in IL-4– and IL-13–stimulated HLFs was analyzed by means of Western blotting. Results: The results showed that (1) IL-4 and IL-13 increased α-smooth muscle actin expression in a dose- and time-dependent manner; (2) IL-4 and IL-13 induced increases in JNK and Erk phosphorylation and activity but not p38 MAP kinase activity; (3) CEP-1347 and PD 98059 attenuated IL-4– and IL13–induced JNK and Erk activity, respectively; and (4) CEP-1347, but not PD 98059, attenuated IL-4– and IL-13–induced α-smooth muscle actin expression. Conclusion: These results indicate that IL-4 and IL-13 are capable of inducing the phenotypic modulation of HLFs to myofibroblasts, and JNK, but not p38 MAP kinase and Erk, regulates IL-4– and IL-13–induced phenotypic modulation of HLFs to myofibroblasts. (J Allergy Clin Immunol 2001;107:1001-8.)

Section snippets

Reagents

SB 203580 and PD 98059 were obtained from Carbiochem-Novabiochem Corporation (La Jolla, Calif) and New England Biolabs, Inc (Beverly, Mass). Anti–TGF-β1 antibody, anti–IL-4 antibody, and anti–IL-13 antibody were obtained from Genzyme TECHNE (Minneapolis, Minn). CEP-1347 was kindly provided by Cephalon Incorporated (West Chester, Pa). SB 203580, PD 98059, and CEP-1347 were dissolved in dimethylsulfoxide (DMSO).

Cell cultures

HLFs were obtained from Clonetics (San Diego, Calif). In the preliminary experiments

IL-4 and IL-13 induce increases in α-smooth muscle actin expression

The expression of α-smooth muscle actin in IL-4– and IL-13–stimulated HLFs increased in a dose-dependent manner (Fig 1, A-C ) and in a time-dependent manner (Fig 1, D-F ).

. IL-4 and IL-13 induce α-smooth muscle actin expression. HLFs were stimulated with medium or with various concentrations of IL-4 (A) or IL-13 (B) and cultured for 6 days. HLFs were cultured with 100 ng/mL IL-4 (A) or 100 ng/mL IL-13 (B) for 2, 4, and 6 days. The amounts of α-smooth muscle actin proteins were quantified with

Discussion

In the present study we examined the role of IL-4 and IL-13 in inducing the phenotypic modulation of HLFs to myofibroblasts and analyzed the intracellular signal–inducing myofibroblastic phenotype of HLF characterized by the expression of α-smooth muscle actin. The results showed the following: (1) IL-4 and IL-13 induced the phenotypic modulation of HLFs to myofibroblasts in a dose- and time-dependent manner; (2) anti–TGF-β1 antibody did not affect IL-4– and IL-13–induced phenotypic modulation

Acknowledgements

We thank Dr Yuzuru Matsuda (Kyowa-Hakko Kogyo Company, Limited) and Dr Jeffery Vaught and Dr Matthew Miller (Cephalon Incorporated) for the generous gift of CEP-1347.

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    Supported in part by a Grant-in-Aid for High-Tech Research Center from the Japanese Ministry of Education, Science, Sports, and Culture to Nihon University.

    ☆☆

    Reprint requests: Shu Hashimoto, MD, First Department of Internal Medicine Nihon University School of Medicine, 30-1 Oyaguchikamimachi, Itabashi-ku, Tokyo 173-8610, Japan.

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