April 2014
Volume 55, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2014
Co-culture of Human Tenon Fibroblast and macrophages as a novel in-vitro model for conjunctival scarring
Author Affiliations & Notes
  • Garima Sharma
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
    UCL School of Pharmacy, London, United Kingdom
  • Hanieh Khalili
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
    UCL School of Pharmacy, London, United Kingdom
  • He Li
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
  • Ashkan Khalili
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
  • Alastair Lockwood
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
    UCL School of Pharmacy, London, United Kingdom
  • Derek Gilroy
    Div Medicine, University College London, London, United Kingdom
  • Steve Brocchini
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
    UCL School of Pharmacy, London, United Kingdom
  • Peng Khaw
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
  • Maryse Bailly
    National Institute for Health Research (NIHR) Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, United Kingdom
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 3203. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Garima Sharma, Hanieh Khalili, He Li, Ashkan Khalili, Alastair Lockwood, Derek Gilroy, Steve Brocchini, Peng Khaw, Maryse Bailly; Co-culture of Human Tenon Fibroblast and macrophages as a novel in-vitro model for conjunctival scarring. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3203.

      Download citation file:


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

      ×
  • Supplements
Abstract
 
Purpose
 

Glaucoma filtration surgery is one of the most effective treatments for lowering intraocular pressure. However, conjunctival contraction and scarring can lead to failure of this surgery. It is established that persistent inflammation after surgery can cause aggressive scarring. The fibroblast populated collagen gel is a well-established assay to study contraction in vitro, but it lacks the complex interaction between inflammatory cells and fibroblasts, which mediate wound-healing process in vivo. To provide a more realistic model of conjunctival scarring and develop more effective treatments, we developed a novel in-vitro model with co-culture of fibroblasts and inflammatory cells.

 
Methods
 

The U937 monocyte-like human cell line was differentiated into macrophage-like cells using phorbol 12-myristate 13- acetate (PMA). An antibody against CD 68 was used to confirm macrophage differentiation. Macrophages were co-cultured with human Tenon’s capsule fibroblasts (HTFs) in three-dimensional collagen gels and contraction was measured over 7 days using digital photography.

 
Results
 

PMA promoted the differentiation of monocytes into CD68-positive macrophages (Figure 1). While neither U937 cells, nor differentiated macrophages alone contracted collagen gels, co-culture of macrophages with HTFs at a 2:1 ratio led to a 50% increase in the contraction potential of HTFs for the first 5 days compared to HTFs alone (Figure 2).

 
Conclusions
 

Inflammatory cells significantly increased the contractibility of HTF as measured in collagen contraction assay. We propose that this new model, recapitulating both aspects of scarring - inflammation and contraction may prove useful in increasing our understanding of the complex wound environment for the development of combination therapies against scarring.

 
 
CD 68 staining of U937 cells following PMA - induced differentiation to macrophages.
 
CD 68 staining of U937 cells following PMA - induced differentiation to macrophages.
 
 
Co-culture of HTF with macrophage increases contraction. Collagen gel contraction for HTF co-cultured with or without macrophage (1:2).
 
Co-culture of HTF with macrophage increases contraction. Collagen gel contraction for HTF co-cultured with or without macrophage (1:2).
 
Keywords: 765 wound healing • 557 inflammation • 474 conjunctiva  
×
×

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.

×