Combined VEGF/PDGF inhibition using axitinib induces αSMA expression and a pro-fibrotic phenotype in human pericytes

  • Jakob Siedlecki
  • Ben Asani
  • Christian Wertheimer
  • Anna Hillenmayer
  • Andreas Ohlmann
  • Claudia Priglinger
  • Siegfried Priglinger
  • Armin Wolf
  • Kirsten Eibl-Lindner
Basic Science

Abstract

Purpose

Large trials on anti-VEGF/PDGF (vascular endothelial/platelet-derived growth factor) combination therapy have been established to improve management of neovascular activity in age-related macular degeneration. Targeting pericytes, PDGF is thought to induce vessel regression and reduce fibrovascular scarring. The fate of pericytes exposed to anti-VEGF/PDGF combination therapy is not clear. Therefore, this study was designed to study the influence of anti-VEGF/PDGF on pericyte phenotype and cellular behavior.

Methods

Human pericytes from placenta (hPC-PL) were treated with axitinib, a tyrosine kinase inhibitor targeting VEGFR1–3 and PDGFR. Toxic effects were excluded using live/dead staining. Phenotypic changes were evaluated using phalloidin staining for actin cytoskeleton and the expression of stress fibers. MRNA and protein expression levels of α-smooth muscle actin (αSMA) as a marker of proto-myofibroblastic transition were evaluated with real-time PCR and Western blotting. Influences of fibrotic cellular mechanisms were evaluated with a scratch wound migration and a collagen gel contraction assay.

Results

Treatment with 0.5, 1, and 2.5 μg/ml axitinib strongly induced a proto-myofibroblast-like actin cytoskeleton with a marked increase in stress fibers. Quantitative real-time PCR and Western blotting revealed these changes to be linked to dose-dependent increases in αSMA mRNA and protein expression. However, fibrotic cellular mechanisms were significantly reduced in the presence of axitinib (scratch wound closure: up to − 78.4%, collagen gel contraction: up to − 37.4%).

Conclusions

Combined anti-VEGF/PDGF inhibition seems to induce a proto-myofibroblast-like phenotype in human pericytes in vitro, but reduce profibrotic cellular mechanisms due to prolonged anti-PDGF inhibition.

Keywords

Age related macular degeneration Choroidal neovascularization VEGF PDGF Axitinib Fibrosis 

Notes

Funding information

No funding was received for this research.

Compliance with ethical standards

This article does not contain any studies with human participants or animals performed by any of the authors.

Conflict of interest

All authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest (such as honoraria; educational grants; participation in speakers’ bureaus; membership, employment, consultancies, stock ownership, or other equity interest; and expert testimony or patent-licensing arrangements), or non-financial interest (such as personal or professional relationships, affiliations, knowledge, or beliefs) in the subject matter or materials discussed in this manuscript.

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Jakob Siedlecki
    • 1
  • Ben Asani
    • 1
  • Christian Wertheimer
    • 1
  • Anna Hillenmayer
    • 1
  • Andreas Ohlmann
    • 1
  • Claudia Priglinger
    • 1
  • Siegfried Priglinger
    • 1
  • Armin Wolf
    • 1
  • Kirsten Eibl-Lindner
    • 1
  1. 1.Department of OphthalmologyLudwig-Maximilians-UniversityMunichGermany

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