Abstract
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