April 2011
Volume 52, Issue 14
ARVO Annual Meeting Abstract  |   April 2011
Imaging of Regulatory T Cells in a Murine Uveitis Model
Author Affiliations & Notes
  • Doran B. Spencer
    Casey Eye Institute, Oregon Health & Science University, Portland, Oregon
  • Steve R. Planck
    Casey Eye Institute, Oregon Health & Science University, Portland, Oregon
  • James T. Rosenbaum
    Casey Eye Institute, Oregon Health & Science University, Portland, Oregon
  • Footnotes
    Commercial Relationships  Doran B. Spencer, None; Steve R. Planck, None; James T. Rosenbaum, None
  • Footnotes
    Support  NIH EY13609, EY10572 and RPB
Investigative Ophthalmology & Visual Science April 2011, Vol.52, 3012. doi:
  • Views
  • Share
  • Tools
    • Alerts
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Doran B. Spencer, Steve R. Planck, James T. Rosenbaum; Imaging of Regulatory T Cells in a Murine Uveitis Model. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3012.

      Download citation file:

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

  • Supplements

Purpose: : Autoimmune uveitis is a T cell-mediated disease. Regulatory T cells (Treg, Foxp3+) represent an anti-inflammatory cell subset of presumed significant therapeutic potential whose role in uveitis remains largely unknown. Using non-invasive, intravital microscopy, we examined the infiltration of Treg and inflammatory, T effector (Teff) cells into the iris in a murine uveitis model.

Methods: : Donor mice (DO11.10, T cell-specific for ovalbumin (OVA)) were immunized with OVA (100 µg) via intraperitoneal (i.p.) injection. After one month, splenocytes were harvested and cultured for 5 days (OVA, 2 µg/ml). CD4+ Teffs were purified, stained with a fluorescent red dye (CMTMR) and transferred (2x107) intravenously (i.v.) into recipient Foxp3-GFP or Foxp3-GFP-DTR (diphtheria toxin receptor) mice. Recipients were subsequently challenged (at 0 h) via intravitreal (i.vit.) injection of 50 µg of OVA or control protein (BSA) with 200 ng endotoxin. For ablation studies of Tregs, animals were injected i.p. with 1 µg of of DT 2 and 1 days prior to i.v. transfer of Teffs and i.vit. challenge. Infiltrating T cells were recorded via videomicroscopy in vivo. The behavior of Foxp3-GFP cells in the normal eyes and skin was also observed.

Results: : At baseline, Foxp3-GFP cells were observed in the limbus but not the cornea nor iris of recipient animals (up to 22/high power field (hpf, 200x)) whereas they were present throughout the skin (12/hpf). Infiltrating Teff (red) and Foxp3-GFP (green) cells accumulated in the iris of the OVA-challenged eye, peaking at 24 hours post injection (142/hpf; BSA-challenged contralateral eye 3/hpf). The fluorescent cell density remained high for 3 days before disappearing by 7 days. Fewer than 5% of the T cells in the iris were observed to move during several observation periods between 24-72 h, without significant Teff/Foxp3-GFP interaction. In contrast, the majority of Teffs and Foxp3-GFP cells in the cornea exhibited significant motility. The ratio of Foxp3-GFP to Teff cells was 1.5 at 24-48 h. Preliminary ablation studies of Foxp3-GFP-DTR cells during uveitis induction do not demonstrate an effect of Foxp3-GFP absence on the overall numbers of Teffs, nor their time of infiltration or persistence in the iris.

Conclusions: : Intravital examination of our model demonstrates the presence of many Foxp3-GFP cells during the inflammatory process. This system presents the first in vivo images of Tregs in the eye, and is a promising platform for examining the cellular and molecular determinants of Teff and Treg function in uveitis.

Keywords: inflammation • uveitis-clinical/animal model 

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.