Claudio Corallo1,2, Annalisa Santucci3,4, Giulia Bernardini3,4, Natale Figura3,4, Roberto Leoncini3,4, Giulia Riolo3,4, Antonio Montella3,4, Chiara Chirico3,4, Ranuccio Nuti3,4, Nicola Giordano3,4. 1. From the Department of Medicine, Surgery, and Neurosciences; the Department of Biotechnology, Chemistry and Pharmacy; and the Department of Medical Biotechnology, University of Siena, Siena, Italy. corallo.claudio@gmail.com. 2. C. Corallo, PhD, Department of Medicine, Surgery and Neurosciences, University of Siena; A. Santucci, PhD, Department of Biotechnology, Chemistry and Pharmacy, University of Siena; G. Bernardini, PhD, Department of Biotechnology, Chemistry and Pharmacy, University of Siena; N. Figura, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; R. Leoncini, PhD, Department of Medical Biotechnology, University of Siena; G. Riolo, PhD, Department of Medical Biotechnology, University of Siena; A. Montella, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; C. Chirico, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; R. Nuti, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; N. Giordano, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena. corallo.claudio@gmail.com. 3. From the Department of Medicine, Surgery, and Neurosciences; the Department of Biotechnology, Chemistry and Pharmacy; and the Department of Medical Biotechnology, University of Siena, Siena, Italy. 4. C. Corallo, PhD, Department of Medicine, Surgery and Neurosciences, University of Siena; A. Santucci, PhD, Department of Biotechnology, Chemistry and Pharmacy, University of Siena; G. Bernardini, PhD, Department of Biotechnology, Chemistry and Pharmacy, University of Siena; N. Figura, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; R. Leoncini, PhD, Department of Medical Biotechnology, University of Siena; G. Riolo, PhD, Department of Medical Biotechnology, University of Siena; A. Montella, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; C. Chirico, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; R. Nuti, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena; N. Giordano, MD, Department of Medicine, Surgery, and Neurosciences, University of Siena.
Abstract
OBJECTIVE: To identify using proteomic analysis the proteins of altered abundance in the affected and unaffected limited cutaneous systemic sclerosis (lcSSc) skin fibroblasts. METHODS: Excision biopsies (3 mm) were obtained from the affected and unaffected skin of 5 patients with lcSSc. Dermal fibroblasts were isolated enzymatically. Two-dimensional gel electrophoresis was used to separate and define proteins in affected and unaffected fibroblast lysates. Proteins of altered abundance were identified by mass spectrometry. Differences among skin samples were confirmed also by immunohistochemistry (IHC) and by quantitative real-time PCR (qRT-PCR) for type I collagen (Col-1) and vimentin (VIM). RESULTS: Proteomic analysis revealed different expressions of proteins involved in cytoskeleton organization (27%), extracellular matrix remodeling (11%), response to oxidative stress (22%), energy metabolism (19%), protein metabolism (5%), cellular homeostasis (5%), signal transduction (3%), and protein transcription, synthesis, and turnover (8%). IHC analysis showed that SSc-affected epidermis is thickened and the dermis is strongly reactive to Col-1 and VIM (typical markers of activated myofibroblasts) compared to SSc-unaffected skin, whose stainings are comparable to those of control healthy skin. Overexpression of Col-1 and VIM mRNA levels in affected lcSSc fibroblasts compared to unaffected lcSSc ones was confirmed by qRT-PCR. CONCLUSION: Consistent with previous studies, these findings are important for 2 reasons: first, because they reveal the opposite behavior of dermal fibroblasts in the unaffected and affected skin areas of the same patient with lcSSc; second, because they demonstrate the histological/histochemical similarities between unaffected skin from patients with lcSSc and healthy control skin.
OBJECTIVE: To identify using proteomic analysis the proteins of altered abundance in the affected and unaffected limited cutaneous systemic sclerosis (lcSSc) skin fibroblasts. METHODS: Excision biopsies (3 mm) were obtained from the affected and unaffected skin of 5 patients with lcSSc. Dermal fibroblasts were isolated enzymatically. Two-dimensional gel electrophoresis was used to separate and define proteins in affected and unaffected fibroblast lysates. Proteins of altered abundance were identified by mass spectrometry. Differences among skin samples were confirmed also by immunohistochemistry (IHC) and by quantitative real-time PCR (qRT-PCR) for type I collagen (Col-1) and vimentin (VIM). RESULTS: Proteomic analysis revealed different expressions of proteins involved in cytoskeleton organization (27%), extracellular matrix remodeling (11%), response to oxidative stress (22%), energy metabolism (19%), protein metabolism (5%), cellular homeostasis (5%), signal transduction (3%), and protein transcription, synthesis, and turnover (8%). IHC analysis showed that SSc-affected epidermis is thickened and the dermis is strongly reactive to Col-1 and VIM (typical markers of activated myofibroblasts) compared to SSc-unaffected skin, whose stainings are comparable to those of control healthy skin. Overexpression of Col-1 and VIM mRNA levels in affected lcSSc fibroblasts compared to unaffected lcSSc ones was confirmed by qRT-PCR. CONCLUSION: Consistent with previous studies, these findings are important for 2 reasons: first, because they reveal the opposite behavior of dermal fibroblasts in the unaffected and affected skin areas of the same patient with lcSSc; second, because they demonstrate the histological/histochemical similarities between unaffected skin from patients with lcSSc and healthy control skin.
Authors: Paraskevi Chairta; Paschalis Nicolaou; Kleitos Sokratous; Christine Galant; Frédéric Houssiau; Anastasis Oulas; George M Spyrou; Marta E Alarcon-Riquelme; Bernard R Lauwerys; Kyproula Christodoulou Journal: Arthritis Res Ther Date: 2020-05-07 Impact factor: 5.156