title: Down-regulation of collagen and connective tissue growth factor expression with hepatocyte growth factor in lung fibroblasts from white scleroderma patients via two signaling pathways.
abstract: OBJECTIVE
  To study the mechanisms by which hepatocyte growth factor (HGF) down-regulates collagen and connective tissue growth factor (CTGF) in scleroderma (systemic sclerosis [SSc]) lung fibroblasts.
  
  
  METHODS
  CTGF, type I collagen, and IkappaBalpha expression, together with MAPK phosphorylation, were studied by immunoblotting of lung fibroblasts derived from white SSc patients. Matrix metalloproteinase 1 (MMP-1) expression in cell culture medium samples was measured by enzyme-linked immunosorbent assay, MMP-1 activity was studied using an MMP-1 assay, and NF-kappaB DNA binding activity was determined using a transcription factor assay.
  
  
  RESULTS
  In lung fibroblasts from white SSc patients, HGF activated MAPK (ERK-1/2) signaling pathways and MMP-1, while it inhibited NF-kappaB and significantly down-regulated CTGF and collagen in a time- and dose-dependent manner. Small interfering RNA (siRNA)-mediated depletion of Grb2 expression disrupted c-Met receptor downstream signaling, which resulted in diminished HGF-induced ERK-1/2 phosphorylation and the recovery of HGF-inhibited expression of MMP-1, NF-kappaB, collagen, and CTGF. The MAPK inhibitor, U0126, blocked MMP-1 activity and restored HGF-inhibited collagen and CTGF accumulation. Inhibition of MMP activity by MMP inhibitor GM1489 and inhibition of MMP-1 expression by siRNA did not prevent HGF-induced ERK-1/2 phosphorylation and NF-kappaB activity, but significantly restored HGF-inhibited collagen and CTGF accumulation. NF-kappaB inhibitor BAY 11-7082 did not interfere with MAPK phosphorylation or MMP-1 expression and activation, but significantly inhibited NF-kappaB DNA binding activity and acted synergistically with HGF to completely diminish the expression of CTGF.
  
  
  CONCLUSIONS
  In lung fibroblasts from white SSc patients, HGF down-regulates the accumulation of CTGF via MAPK/MMP-1 and NF-kappaB signaling pathways, whereas collagen down-regulation is mediated mainly by a MAPK/MMP-1-dependent pathway.
authors: Medical University of South Carolina, Charleston, SC 29425, USA. bogatkev@musc.edu
authors: Galina S
authors: GS
authors: Bogatkevich
authors: Anna
authors: A
authors: Ludwicka-Bradley
authors: Kristin B
authors: KB
authors: Highland
authors: Faye
authors: F
authors: Hant
authors: Paul J
authors: PJ
authors: Nietert
authors: C Beth
authors: CB
authors: Singleton
authors: Richard M
authors: RM
authors: Silver
mesh_terms: D018929:Cell Culture Techniques; D024042:Collagen Type I / Q000096:biosynthesis*; D055513:Connective Tissue Growth Factor; D015536:Down-Regulation; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005347:Fibroblasts / Q000378:metabolism*; D017228:Hepatocyte Growth Factor / Q000096:biosynthesis*; D006801:Humans; D017874:Immediate-Early Proteins / Q000096:biosynthesis*; D036341:Intercellular Signaling Peptides and Proteins / Q000096:biosynthesis*; D008168:Lung / Q000378:metabolism; D020935:MAP Kinase Signaling System; D008297:Male; D020781:Matrix Metalloproteinase 1 / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D012595:Scleroderma, Systemic / Q000378:metabolism*; D044465:White People
chemical_list: C526317:CCN2 protein, human; D024042:Collagen Type I; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D016328:NF-kappa B; D055513:Connective Tissue Growth Factor; D017228:Hepatocyte Growth Factor; D020929:Mitogen-Activated Protein Kinase Kinases; D020781:Matrix Metalloproteinase 1