title: Th17 cells favor inflammatory responses while inhibiting type I collagen deposition by dermal fibroblasts: differential effects in healthy and systemic sclerosis fibroblasts.
abstract: BACKGROUND
  T helper (Th)-17 cells are increased in systemic sclerosis (SSc). We therefore assessed whether Th17 cells could modulate the inflammatory and fibrotic responses in dermal fibroblasts from healthy donors (HD) and SSc individuals.
  
  
  METHODS
  Fibroblasts were obtained from 14 SSc and 8 HD skin biopsies. Th17 clones were generated from healthy peripheral blood upon enrichment of CC chemokine receptor (CCR)-4/CCR6/CD161 expressing cells. Their cytokine production was assessed by flow cytometry and multiplex beads immunoassay. Fibroblast production of monocyte chemoattractant protein (MCP)-1, interleukin (IL)-8, matrix metalloproteinase (MMP)-1, tissue inhibitor of metalloproteinase (TIMP)-1, MMP-2 and type-I collagen was quantified by enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay (RIA), and changes in their transcription levels assessed by real-time PCR. Intracellular signals were dissected by western blot and the use of pharmacological inhibitors. IL-17A, tumor necrosis factor (TNF) and interferon-gamma (IFN-γ) blocking reagents were used to assess the specificity of the observed effects.
  
  
  RESULTS
  IL-17A increased MCP-1, IL-8 and MMP-1 production in a dose-dependent manner while having no effect on type I collagen in HD and SSc fibroblasts both at protein and mRNA levels. Nuclear factor-kappa B (NF-κB) and p38 were preferentially involved in the induction of MCP-1 and IL-8, while MMP-1 was most dependent on c-Jun N-terminal kinase (JNK). Supernatants of activated Th17 clones largely enhanced MCP-1, IL-8 and MMP-1 while strongly inhibiting collagen production. Of note, the production of MCP-1 and IL-8 was higher, while collagen inhibition was lower in SSc compared to HD fibroblasts. The Th17 clone supernatant effects were mostly dependent on additive/synergistic activities between IL-17A, TNF and in part IFN-γ. Importantly, the inhibition of type I collagen production induced by the Th17 clone supernatants was completely abrogated by blockade of IL-17A, TNF and IFN-γ mostly in SSc fibroblasts, revealing an intrinsic resistance to inhibitory signals in SSc.
  
  
  CONCLUSIONS
  Our findings demonstrate that in vitro Th17 cells elicit pro-inflammatory responses while restraining collagen production. Thus, the increased Th17 cell number observed in SSc may impact on the inflammatory component of the disease simultaneously potentially providing a protective role against fibrosis.
authors.forename: Nicolò Constantino
authors.initials: NC
authors.lastname: Brembilla
authors.forename: Elisa
authors.initials: E
authors.lastname: Montanari
authors.forename: Marie-Elise
authors.initials: ME
authors.lastname: Truchetet
authors.forename: Elena
authors.initials: E
authors.lastname: Raschi
authors.forename: Pierluigi
authors.initials: P
authors.lastname: Meroni
authors.forename: Carlo
authors.initials: C
authors.lastname: Chizzolini
mesh_terms: D015153:Blotting, Western; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism*; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism*; D017077:Culture Media, Conditioned / Q000378:metabolism / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D018836:Inflammation Mediators / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology; D020381:Interleukin-17 / Q000494:pharmacology; D016209:Interleukin-8 / Q000235:genetics / Q000378:metabolism*; D008297:Male; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism*; D011863:Radioimmunoassay; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012595:Scleroderma, Systemic / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D012867:Skin / Q000378:metabolism / Q000473:pathology; D058504:Th17 Cells / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: C111835:CCL2 protein, human; D018932:Chemokine CCL2; D024042:Collagen Type I; D017077:Culture Media, Conditioned; D018836:Inflammation Mediators; D020381:Interleukin-17; D016209:Interleukin-8; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; D020781:Matrix Metalloproteinase 1