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Slit2/Robo4 axis may contribute to endothelial cell dysfunction and angiogenesis disturbance in systemic sclerosis
  1. Eloisa Romano1,
  2. Mirko Manetti2,
  3. Irene Rosa1,2,
  4. Bianca Saveria Fioretto1,
  5. Lidia Ibba-Manneschi2,
  6. Marco Matucci-Cerinic1,
  7. Serena Guiducci1
  1. 1 Department of Experimental and Clinical Medicine, Division of Rheumatology, University of Florence, Scleroderma Unit, Azienda Ospedaliero-Universitaria Careggi (AOUC), Florence, Italy
  2. 2 Department of Experimental and Clinical Medicine, Section of Anatomy and Histology, University of Florence, Florence, Italy
  1. Correspondence to Dr Eloisa Romano, Department of Experimental and Clinical Medicine, Division of Rheumatology, University of Florence, Florence, 50139, Italy; eloisaromano{at}libero.it

Abstract

Objective In systemic sclerosis (SSc), early microvascular injury is followed by impaired angiogenesis and peripheral capillary loss. Here, we investigated the possible contribution of the neurovascular guidance molecule Slit2 and its Roundabout (Robo) receptors to SSc-related endothelial cell dysfunction.

Methods Circulating Slit2 levels were measured in patients with SSc and healthy controls. Slit2, Robo1 and Robo4 expression was investigated in SSc and healthy skin biopsies and explanted dermal microvascular endothelial cells (MVECs). Slit2/Robo4 function in MVEC angiogenesis was studied by cell viability, wound healing and capillary-like tube formation assays.

Results Circulating Slit2 was significantly increased in either SSc or patients with a very early diagnosis of SSc (VEDOSS) compared with controls. Interestingly, serum Slit2 levels were raised in patients with VEDOSS with nailfold videocapillaroscopy (NVC) abnormalities, while they were similar in VEDOSS with normal NVC and controls. In SSc, Slit2 and Robo4 expression was upregulated in clinically affected skin and explanted MVECs in respect to controls. The angiogenic performance of healthy MVECs was significantly reduced after challenge with recombinant human Slit2 or SSc sera. These inhibitory effects were significantly attenuated when SSc sera were preincubated with an anti-Slit2 blocking antibody. In vitro angiogenesis was severely compromised in SSc-MVECs and could be significantly ameliorated by Slit2 neutralisation or ROBO4 gene silencing. Slit2/Robo4 axis interfered with angiogenesis through the inhibition of Src kinase phosphorylation.

Conclusions In SSc, increased circulating levels of Slit2 and activation of the Slit2/Robo4 antiangiogenic axis may contribute to peripheral microangiopathy since the very early phase of the disease.

  • systemic sclerosis
  • qualitative research
  • autoimmune diseases
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Footnotes

  • ER and MM contributed equally.

  • Handling editor Josef S Smolen

  • Contributors Study conception and design: ER, MM, MM-C and SG. Acquisition of data: ER, MM, IR, BSF and SG. Interpretation of data: ER, MM, IR, LI-M and SG. Manuscript preparation: ER, MM, IR and MM-C.

  • Funding The authors have not declared a specific grant for this research from any funding agency in the public, commercial or not-for-profit sectors.

  • Competing interests None declared.

  • Patient consent Obtained.

  • Ethics approval The study was approved by the local institutional review board at the Azienda Ospedaliero-Universitaria Careggi (AOUC), Florence, Italy.

  • Provenance and peer review Not commissioned; externally peer reviewed.

  • Presented at Part of this work has been presented at the EULAR 2018 congress and published as a conference abstract: Ann Rheum Dis 2018;77(Suppl 2):97–8. doi: 10.1136/annrheumdis-2018-eular.4610.

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