July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Substrate stiffness influences cell density, transcriptome and angiomodulatory properties of retinal pigment epithelium cells
Author Affiliations & Notes
  • Clara Gimpel
    Eye Center, Medical Center - University of Freiburg, Freiburg, Germany
  • René Poettke
    Eye Center, Medical Center - University of Freiburg, Freiburg, Germany
  • Gunther R Schlunck
    Eye Center, Medical Center - University of Freiburg, Freiburg, Germany
  • Footnotes
    Commercial Relationships   Clara Gimpel, None; René Poettke, None; Gunther Schlunck, None
  • Footnotes
    Support  Deutsche Ophthalmologische Gesellschaft (german ophthalmological society) training grant
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 1923. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Clara Gimpel, René Poettke, Gunther R Schlunck; Substrate stiffness influences cell density, transcriptome and angiomodulatory properties of retinal pigment epithelium cells. Invest. Ophthalmol. Vis. Sci. 2019;60(9):1923.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

Purpose : Physiological and pathological processes such as ageing or the aggregation of basal deposits can alter the mechanical properties of Bruch’s membrane. Little is known about how these changes influence the surrounding cells. Therefore, it was our goal to further characterize the influence of substrate stiffness on retinal pigment epithelium (RPE) cells.

Methods :
ARPE-19 cells were cultured on hydrogels of differing physiological Young’s moduli and tissue culture plastic. In a pilot study, the RPE transcriptomes were analyzed using MACE (massive analysis of cDNA ends) sequencing techniques and digital PCR.
RPE cell supernatants were used for cell proliferation (MTT) and migration assays in human umbilical vein endothelial cells (HUVECs). RPE densities were assessed in phase contrast micrographs obtained before cell harvesting.

Results :
4110 of the 20046 mRNAs detected by MACE reached an average normalized expression of more than 50 copies/million. 417 of them were differentially expressed on substrates of differing stiffness. 193 mRNAs were enhanced on soft gels (log2 fc≥ 1, p<0.05), among them several genes associated with angiogenesis, such as CD44 and CTGF (connective tissue growth factor). Among the 224 genes overexpressed on stiffer substrates (log2 fc ≤-1, p<0.05) were antiangiogenic factors like thrombospondin-1 and PEDF (pigment epithelium derived factor). These sequencing data were validated by digital PCR.
HUVECs stimulated with supernatant from RPE cells on soft gels reproducibly showed a higher proliferation rate than controls treated with supernatant from cells grown on plastic (1.60-fold, p=0.007, n=5). In the migration assay HUVECs treated with supernatant from cells on soft gels migrated faster than the ones treated with supernatant from stiff gels (1.56-fold, n.s.) or plastic (1.82-fold, p=0.028, single experiment).
Furthermore, soft substrates did not support cell plating at very high densities and rendered confluent cell cultures with a higher cell size variability than stiff substrates.

Conclusions : Our findings indicate that basal deposits may influence RPE cells and their angiomodulatory potential through their biomechanical properties. Since the RPE plays a crucial part in the pathogenesis of age-related macular degeneration, this aspect could contribute to the disease and may be relevant for tissue engineering-based therapeutic applications.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×