Previous studies have suggested that corneal stromal keratocytes express the CD34 antigen. We wished to investigate CD34 antigen expression in normal mouse cornea using dual and triple-staining techniques. Whole-mount preparations of mouse and rat corneas were examined with confocal microscopy using single, dual, or triple immunostaining to study their morphology, phenotype, and distribution. Single-cell suspensions from normal mouse corneas were also prepared and analyzed by flow cytometry. After short-term culture of corneal stromal explants, nonadherent cells were harvested and cyto-spins were prepared and stained for different markers. Combined staining for F-actin and leukocyte differentiation markers clearly showed that the corneal stroma contains a population of CD45(+) resident bone marrow-derived cells, whereas most cells were CD45-F-actin(+) keratocytes. A significant proportion (two thirds) of CD45(+) cells in the normal corneal stroma expressed CD34(+), whereas no CD45(-) cells (i.e., keratocytes) coexpressed CD34. In addition, CD34(+) cells were CD11c(-) and CD11b(+). Fewer than 10 % of the CD34(+) cells also coexpressed Sca-1(+), but no CD34(+) cells coexpressed major histocompatibility complex (MHC) class II+. In contrast, the remaining population of CD45(+)CD34(-) cells in the corneal stroma expressed CD11b, MHC class II+ but not CD11c and were found mostly in the anterior and peripheral part of stroma. These cells are in intimate contact with corneal keratocytes, which stained only for F-actin and were negative for all leukocyte markers. Very few CD45+ cells expressed the B220 marker, suggesting a plasmacytoid dendritic cell phenotype. Flow cytometry analyses confirmed the morphometric data showing that 68% of CD45(+) cells coexpress CD34 and CD11b, whereas 22% are CD11b(+)CD34(-). We conclude that the normal mouse cornea contains two populations of bone marrow-derived leukocytes, both of which are distinct from stromal keratocytes. The larger population resembles CD34(+) hemopoietic stem cells, whereas the smaller population are CD34(-)CD11b(+) MHC class II+ macrophages. A very small percentage comprises plasmacytoid dendritic cells.
|Number of pages||8|
|Publication status||Published - Apr 2005|
- hemopoietic stem cell
- dendritic cell
- HUMAN KERATOCYTES