Abstract
Purpose: :
We examined the chronology of the determination and differentiation of ocular surface epithelia using tet-On triple transgenic mouse lines.
Methods: :
Three Pax6-rtTA (P6Ros) driver mouse lines, (e.g., P6Ros1, P6Ros2 and P6Ros11) were generated with ocular surface tissue-specific Pax6 promoter. Triple transgenic P6Ros/TC/Rosa26mTmG mice were obtained by cross breeding of P6Ros, tet-O-Cre (TC) and Rosa26mTmG (mTmG). Pax6 expressing cells of triple transgenic mice changed from red to green EGFP by deleting tomato-red transgene upon dox induction. The experimental mice were fed dox at different gestations, e.g., embryonic (E)0, E12.5 & E16.5 to postnatal day P0 & P21, and E0 to E12.5 & E16.5. The expression pattern of EGFP was determined by fluorescent microscopy, and immune fluorescence staining with anti-Pax6 antibodies.
Results: :
Fluorescence microscopy revealed similar expression patterns of EGFP from all triple transgenic experimental mice of the three different P6Ros driver mice. P6Ros11 driver had the highest expression. EGFP positive cells were found in basal cells of corneal epithelium, acini of lacrimal gland, and lens epithelial and fiber cells of triple transgenic mice continuously fed dox starting from E0, E12.5 and E16.5 to P21. Induction during embryonic development from E0 to E12.5, a few EGFP positive cells were found in corneal and conjunctival epithelium, and acini of lacrimal gland of triple transgenic embryos. Induction from E0 to E16.5, the EGFP positive cells were found in corneal epithelium, bulbar but not palpebral conjunctiva, and acini of lacrimal glands. Immunofluorescence staining with anti-Pax6 antibodies indicating that EGFP positive cells were Pax6 positive. No EGFP positive cells were found in acini of Meibomian gland.
Conclusions: :
Our data suggested that during embryonic development ocular surface peridermal epithelium derived from surface ectoderm expresses Pax6 including conjunctiva, cornea and lacrimal glands. Pax6 expression gradually became limited to basal cells of corneal epithelium, and lacrimal glands, indicating that Pax6 expressing cells were not stem cells, rather they are characteristic of transient amplifying cells.
Keywords: gene/expression • cornea: epithelium • lacrimal gland