April 2014
Volume 55, Issue 13
ARVO Annual Meeting Abstract  |   April 2014
Intravital multiphoton visualization identifies inter-networking between nerves and bone marrow derived cells of the mouse cornea
Author Affiliations & Notes
  • Tomas Blanco
    Ophthalmology, Duke University, School of Medicine, Durham, NC
  • Daniel R Saban
    Ophthalmology, Duke University, School of Medicine, Durham, NC
    Immunology, Duke School of Medicine, Durham, NC
  • Footnotes
    Commercial Relationships Tomas Blanco, None; Daniel Saban, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 4057. doi:
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      Tomas Blanco, Daniel R Saban; Intravital multiphoton visualization identifies inter-networking between nerves and bone marrow derived cells of the mouse cornea. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4057.

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

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Purpose: We have previously shown that (BAC-transduced) CX3CR1-Cre Rosa-tdTomato (or GFP) mice faithfully report bone marrow (BM)-derived cells in the cornea, which populate the tissue in the anticipated manner. Multiphoton visualization revealed the presence of reporter (tdTomato or GFP)-positive structures that interconnected BM-derived cells in a complex network that stretches across the width and depth of the normal cornea. The current study sought to further understand the form and function of these interconnecting structures.

Methods: Intravital imaging of mouse cornea was done via multiphoton microscopy (range 900-960 nm at 40 nm/sec, 80 MHz rep rate, 140 fsec pulse width) and image processing via Imaris Bitplane's core scientific software. We used progeny of CX3CR1-Cre mice x Rosa-GFP or Rosa-tdTomato mice. Thy-1 eYFP transgenic mice were used for nerve visualization, and confirmed via confocal microscopy of Tuj1 stained cornea explants. Bone marrow chimeras were generated with host CX3CR1-Cre Rosa-tdTomato mice and donor CX3CR1-Cre Rosa-GFP cells (or vice versa). Also, wildtype or Thy-1 eYFP hosts received donor CX3CR1-Cre Rosa-tdTomato cells.

Results: Tuj1 staining revealed BM derived cells in physical contact with corneal nerves throughout the cornea. Chimeric Thy-1 eYFP hosts with CX3CR1-Cre Rosa-tdTomato donors also showed this. Even more striking, donor tdTomato+ BM derived cells that populated the cornea were associated with the emergence of interconnecting tdTomato+ structures, which co-localized with corneal nerves (Thy-1). Chimeric CX3CR1-Cre Rosa-tdTomato hosts with CX3CR1-Cre Rosa-GFP donors showed corneal nerves that co-localized with host reporter (tdTomato), donor reporter (GFP), or both. Furthermore, donor BM derived cells laden with host tdTomato reporter were also readily detectable.

Conclusions: We conclude that BM derived cells populate the cornea in tight physical contact with corneal nerves. Furthermore, chimeras showed that BM derived cell-expressed reporter (tdTomato or GFP) co-localizes with corneal nerves, and that reporter protein is bi-directionally transported between adjacent BM derived cells. Thus, the two respective networks, i.e. of corneal nerves and BM-derived cells that populate the cornea, appear to interact in parallel. Future studies are required to understand the immunologic function of such inter-networking.

Keywords: 480 cornea: basic science • 423 antigen presentation/processing • 555 immunomodulation/immunoregulation  

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