July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Characterization of bioorthogonally crosslinked collagen gels with encapsulated corneal stromal stem cells
Author Affiliations & Notes
  • Sarah Hull
    Chemical Engineering, Stanford University, Palo Alto, California, United States
    Ophthalmology, Stanford University, Palo Alto, California, United States
  • Gabriella Fernandes-Cunha
    Ophthalmology, Stanford University, Palo Alto, California, United States
  • Ilham Putra
    Ophthalmology, University of Illinois at Chicago, Illinois, United States
  • Medi Eslani
    Ophthalmology, University of Illinois at Chicago, Illinois, United States
  • Ali R Djalilian
    Ophthalmology, University of Illinois at Chicago, Illinois, United States
  • Sarah Heilshorn
    Materials Science and Engineering, Stanford University, Palo Alto, California, United States
    Chemical Engineering, Stanford University, Palo Alto, California, United States
  • David Myung
    Ophthalmology, Stanford University, Palo Alto, California, United States
    Chemical Engineering, Stanford University, Palo Alto, California, United States
  • Footnotes
    Commercial Relationships   Sarah Hull, None; Gabriella Fernandes-Cunha, None; Ilham Putra, None; Medi Eslani, None; Ali Djalilian, None; Sarah Heilshorn, None; David Myung, Stanford University (P)
  • Footnotes
    Support  National Eye Institute (NIH K08EY028176 and P30‐EY026877), Stanford SPARK Translational Research Grant, Research to Prevent Blindness (RPB) Foundation Core Grant, Eversight
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 4110. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Sarah Hull, Gabriella Fernandes-Cunha, Ilham Putra, Medi Eslani, Ali R Djalilian, Sarah Heilshorn, David Myung; Characterization of bioorthogonally crosslinked collagen gels with encapsulated corneal stromal stem cells. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4110.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose : A therapeutic strategy that can rapidly and suturelessly restore transparent stromal tissue to the cornea without the need for cadaveric tissue could be of great benefit to those suffering from or at risk of corneal blindness. We have developed a bio-orthogonally crosslinked, collagen-based hydrogel and hypothesize that delivering corneal stromal stem cells (CSSCs) within this gel can facilitate remodeling of the applied matrix and maintain corneal transparency over time.

Methods : Bioorthogonal collagen gels were formed using strain-promoted azide-alkyne cycloaddition (SPAAC), a biocompatible form of copper-free click chemistry. Briefly, type I collagen was functionalized with either an azide or cyclooctyne using NHS ester chemistry and then these two components were combined to produce a gel. The gels’ mechanical properties were modulated by blending in different amounts of unmodified collagen to change the overall covalent crosslinking density. CSSCs were encapsulated within the gels and changes in transparency and gel mechanics were monitored over time using UV/Vis spectroscopy and microrheology, respectively. In addition, CSSC morphology and keratocyte differentiation markers were determined using qPCR and immunohistochemistry on cells encapsulated within gels of varying crosslink density.

Results : We demonstrate that we can tune gel mechanical properties by changing crosslinking density to produce gels with storage moduli between 10-200 Pa. The crosslinked collagen gels with and without encapsulated CSSCs showed transparency over 90% in the visible light range and were able to maintain transparency over a two-week culture time. Encapsulation of primary CSSCs within SPAAC-crosslinked collagen gels yielded cells that expressed aldehyde dehydrogenase 3A1 (ALDH3A1), keratan sulfate, hepatocyte growth factor, and keratinocyte growth factor in the absence of alpha smooth muscle actin (alpha-SMA). In contrast, culturing primary CSSCs within physical collagen gels resulted in more spindle-like morphology, expression of alpha-SMA, and lower overall levels of ALDH3A1.

Conclusions : Our results indicate that CSSCs can be successfully encapsulated within bioorthogonally crosslinked gels, that the gel constructs can maintain transparency and mechanical strength over time, and that CSSCs can express keratocyte markers and secrete keratocyte-specific factors when cultured within the gels.

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.

×