title: Effects of macitentan and its active metabolite on cultured human systemic sclerosis and control skin fibroblasts.
abstract: OBJECTIVE
  To investigate the effects of the endothelin 1 (ET-1) receptor antagonists (ETRA) macitentan, its active metabolite ACT-132577, and bosentan on myofibroblast activation and extracellular matrix production induced by ET-1 in cultured systemic sclerosis (SSc) and control skin fibroblasts.
  
  
  METHODS
  Fibroblasts were obtained from skin biopsies of 6 patients with SSc and 5 healthy subjects. Some cultured cells were untreated or treated with macitentan, ACT-132577, or bosentan alone (10 μM). Other cultured cells were treated with ET-1 alone (100 nM) or with ETRA, and after 1 h, also with ET-1. After 48 h of treatment, myofibroblast activation was investigated to evaluate the α-smooth muscle actin (α-SMA) expression by immunofluorescence; type I collagen (COL-1) and fibronectin (FN) were investigated by immunocytochemistry, Western blotting, and quantitative real-time PCR (qRT-PCR). Statistical analysis was performed by the nonparametric Mann-Whitney U test.
  
  
  RESULTS
  In cultured SSc skin fibroblasts, only the treatment with macitentan significantly reduced the basal level of α-SMA expression (p = 0.03 vs untreated cells). Macitentan also significantly reduced the basal level of COL-1 synthesis, similarly to bosentan (p < 0.05 vs untreated cells). Macitentan or ACT-132577 antagonized the ability of ET-1 to further induce α-SMA expression (p = 0.03), COL-1, and FN synthesis (p = 0.03, p = 0.005); bosentan showed similar effects. These results obtained by immunofluorescence and immunocytochemistry were confirmed by Western blotting and qRT-PCR. The downregulatory effects exerted by ETRA were observed also in cultured human control skin fibroblasts.
  
  
  CONCLUSIONS
  Macitentan and ACT-132577 seem to downregulate in vitro the profibrotic myofibroblast phenotype induced by ET-1 in cultured human SSc skin fibroblasts.
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital. mcutolo@unige.it.
authors.forename: Maurizio
authors.initials: M
authors.lastname: Cutolo
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Paola
authors.initials: P
authors.lastname: Montagna
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Renata
authors.initials: R
authors.lastname: Brizzolara
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Vanessa
authors.initials: V
authors.lastname: Smith
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Elisa
authors.initials: E
authors.lastname: Alessandri
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Barbara
authors.initials: B
authors.lastname: Villaggio
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Alberto
authors.initials: A
authors.lastname: Sulli
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Pietro Paolo
authors.initials: PP
authors.lastname: Tavilla
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Carmen
authors.initials: C
authors.lastname: Pizzorni
authors.affiliation: From Research Laboratory and Academic Division of Clinical Rheumatology, Research Laboratory of Nephrology, Department of Internal Medicine, and the Department of Health Science, Unit of Dermatology, University of Genoa, Genoa, Italy; and the Department of Rheumatology, Ghent University Hospital, Ghent, Belgium.M. Cutolo, MD, Full Professor, Director; P. Montagna, BS, PhD; R. Brizzolara, BS, PhD; E. Alessandri, MD; A. Sulli, MD, Assistant Professor; C. Pizzorni, MD, Assistant Professor; S. Soldano, BS, PhD, Research Laboratory and Academic Division of Clinical Rheumatology; B. Villaggio, BS, Research Laboratory of Nephrology, Department of Internal Medicine; P.P. Tavilla, MD, Department of Health Science, Unit of Dermatology, University of Genoa; V. Smith, MD, PhD, Department of Rheumatology, Ghent University Hospital.
authors.forename: Stefano
authors.initials: S
authors.lastname: Soldano
mesh_terms: D000199:Actins / Q000378:metabolism; D000368:Aged; D000077300:Bosentan; D015536:Down-Regulation / Q000187:drug effects; D065130:Endothelin A Receptor Antagonists / Q000494:pharmacology*; D005260:Female; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D008297:Male; D008875:Middle Aged; D011743:Pyrimidines / Q000494:pharmacology*; D012595:Scleroderma, Systemic / Q000378:metabolism / Q000473:pathology*; D012867:Skin / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D013449:Sulfonamides / Q000494:pharmacology*
chemical_list: D000199:Actins; D065130:Endothelin A Receptor Antagonists; D011743:Pyrimidines; D013449:Sulfonamides; C572762:aprocitentan; D000077300:Bosentan; C533860:macitentan
keywords: ENDOTHELIN 1; ENDOTHELIN RECEPTOR ANTAGONISTS; EXTRACELLULAR MATRIX; HUMAN SKIN FIBROBLASTS; SKIN FIBROSIS; SYSTEMIC SCLEROSIS