title: Connective tissue growth factor induces collagen I expression in human lung fibroblasts through the Rac1/MLK3/JNK/AP-1 pathway.
abstract: Connective tissue growth factor (CTGF) plays an important role in lung fibrosis. In this study, we investigated the role of Rac1, mixed-lineage kinase 3 (MLK3), c-Jun N-terminal kinase (JNK), and activator protein-1 (AP-1) in CTGF-induced collagen I expression in human lung fibroblasts. CTGF caused concentration- and time-dependent increases in collagen I expression. CTGF-induced collagen I expression was inhibited by the dominant negative mutant (DN) of Rac1 (RacN17), MLK3DN, MLK3 inhibitor (K252a), JNK1DN, JNK2DN, a JNK inhibitor (SP600125), and an AP-1 inhibitor (curcumin). Treatment of cells with CTGF caused activation of Rac1, MLK3, JNK, and AP-1. The CTGF-induced increase in MLK3 phosphorylation was inhibited by RacN17. Treatment with RacN17 and the MLK3DN inhibited CTGF-induced JNK phosphorylation. CTGF caused increases in c-Jun phosphorylation and the recruitment of c-Jun and c-Fos to the collagen I promoter. Furthermore, stimulation of cells with the CTGF resulted in increases in AP-1-luciferase activity; this effect was inhibited by Rac1N17, MLK3DN, JNK1DN, and JNK2DN. Moreover, CTGF-induced α-smooth muscle actin (α-SMA) expression was inhibited by the procollagen I small interfering RNA (siRNA). These results suggest for the first time that CTGF acting through Rac1 activates the MLK3/JNK signaling pathway, which in turn initiates AP-1 activation and recruitment of c-Jun and c-Fos to the collagen I promoter and ultimately induces collagen I expression in human lung fibroblasts.
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Chien-Huang
authors.initials: CH
authors.lastname: Lin
authors.affiliation: Department of Pulmonary Medicine, Taipei Medical University - Wanfang Hospital, Taipei, Taiwan; School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Ming-Chih
authors.initials: MC
authors.lastname: Yu
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Wan-Hsuan
authors.initials: WH
authors.lastname: Tung
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Tzu-Ting
authors.initials: TT
authors.lastname: Chen
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Chung-Chi
authors.initials: CC
authors.lastname: Yu
authors.affiliation: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Chih-Ming
authors.initials: CM
authors.lastname: Weng
authors.affiliation: Department of Pharmacology, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Yan-Jyu
authors.initials: YJ
authors.lastname: Tsai
authors.affiliation: Department of Pulmonary Medicine, Taipei Medical University - Wanfang Hospital, Taipei, Taiwan; School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Kua-Jen
authors.initials: KJ
authors.lastname: Bai
authors.affiliation: Taipei Medical University Wangfang Hospital, Taipei, Taiwan.
authors.forename: Chuang-Ye
authors.initials: CY
authors.lastname: Hong
authors.affiliation: Taipei Medical University Wangfang Hospital, Taipei, Taiwan; Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Ming-Hsien
authors.initials: MH
authors.lastname: Chien
authors.affiliation: School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, Taiwan. Electronic address: bcchen@tmu.edu.tw.
authors.forename: Bing-Chang
authors.initials: BC
authors.lastname: Chen
mesh_terms: D000199:Actins / Q000378:metabolism; D002460:Cell Line; D024042:Collagen Type I / Q000378:metabolism*; D055513:Connective Tissue Growth Factor / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D008168:Lung / Q000166:cytology*; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008954:Models, Biological; D010766:Phosphorylation / Q000187:drug effects; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11
chemical_list: D000199:Actins; D024042:Collagen Type I; D055513:Connective Tissue Growth Factor; D020930:MAP Kinase Kinase Kinases; D018808:Transcription Factor AP-1; D020830:rac1 GTP-Binding Protein; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11; C541116:ACTA2 protein, human
keywords: AP-1; ASK1; CCAAT/enhancer-binding protein β; CTGF; ChIP; Collagen I; Connective tissue growth factor; ECM; ERK; FCS; Fibroblasts; IL-6; JAK; JNK; Janus kinase; Lung fibrosis; MAPK kinase kinase; MAPKKK; MAPKs; MEM; MLB; MLK3; Minimum Essential Medium; NEAAs; PBD; STAT; Signal transduction; TGF-β; WT; activator protein-1; apoptosis signal-regulating kinase 1; c-Jun N-terminal kinase; c/EBPβ; chromatin immunoprecipitation; connective tissue growth factor; extracellular matrix; extracellular signal-regulated kinase; fetal calf serum; interleukin-6; magnesium lysis buffer; mitogen-activated protein kinases; mixed-lineage kinase 3; non-essential amino acids; p21-binding domain; siRNA; signal transducer and activator of transcription; small interfering RNA; transforming growth factor-β; wild-type; α-SMA; α-smooth muscle actin