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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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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.
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.
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.
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.
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