A conditional Pax6 depletion study with no morphological effect on the adult mouse corneal epithelium.

Objective: The corneas of heterozygous Pax6 mice develop abnormally and deteriorate further after birth but it is not known whether the postnatal deterioration is predetermined by abnormal development. Our objective was to identify whether depletion of Pax6 in adult mice caused any corneal abnormalities, similar to those in Pax6 mice, where Pax6 levels are low throughout development and adulthood. We used two tamoxifen-inducible, Cre-loxP experimental strategies to deplete Pax6 either ubiquitously or in a restricted range of cell types.

Analysis of compound heterozygotes reveals that the mouse floxed Pax6 (tm1Ued) allele produces abnormal eye phenotypes.

Analysis of abnormal phenotypes produced by different types of mutations has been crucial for our understanding of gene function. Some floxed alleles that retain a neomycin-resistance selection cassette (neo cassette) are not equivalent to wild-type alleles and provide useful experimental resources. Pax6 is an important developmental gene and the aim of this study was to determine whether the floxed Pax6 (tm1Ued) (Pax6 (fl) ) allele, which has a retained neo cassette, produced any abnormal eye phenotypes that would imply that it differs from the wild-type allele.

Abnormal corneal epithelial maintenance in mice heterozygous for the micropinna microphthalmia mutation Mp.

We investigated the corneal morphology of adult Mp/+ mice, which are heterozygous for the micropinna microphthalmia mutation, and identified several abnormalities, which implied that corneal epithelial maintenance was abnormal. The Mp/+ corneal epithelium was thin, loosely packed and contained goblet cells in older mice. Evidence also suggested that the barrier function was compromised.

Lineage tracing in the adult mouse corneal epithelium supports the limbal epithelial stem cell hypothesis with intermittent periods of stem cell quiescence.

The limbal epithelial stem cell (LESC) hypothesis proposes that LESCs in the corneal limbus maintain the corneal epithelium both during normal homeostasis and wound repair. The alternative corneal epithelial stem cell (CESC) hypothesis proposes that LESCs are only involved in wound repair and CESCs in the corneal epithelium itself maintain the corneal epithelium during normal homeostasis. We used tamoxifen-inducible, CreER-loxP lineage tracing to distinguish between these hypotheses.

Evaluating alternative stem cell hypotheses for adult corneal epithelial maintenance.

In this review we evaluate evidence for three different hypotheses that explain how the corneal epithelium is maintained. The limbal epithelial stem cell (LESC) hypothesis is most widely accepted. This proposes that stem cells in the basal layer of the limbal epithelium, at the periphery of the cornea, maintain themselves and also produce transient (or transit) amplifying cells (TACs).

Hemizygous Le-Cre transgenic mice have severe eye abnormalities on some genetic backgrounds in the absence of LoxP sites.

Eye phenotypes were investigated in Le-Cre(Tg/-); Pax6(fl/+) mice, which were expected to show tissue-specific reduction of Pax6 in surface ectoderm derivatives. To provide a better comparison with our previous studies of Pax6(+/-) eye phenotypes, hemizygous Le-Cre(Tg/-) and heterozygous Pax6(fl/+)mice were crossed onto the CBA/Ca genetic background. After the Le-Cre transgene had been backcrossed to CBA/Ca for seven generations, significant eye abnormalities occurred in some hemizygous Le-Cre(Tg/-); Pax6(+/+) controls (without a floxed Pax6(fl) allele) as well as experimental Le-Cre(Tg/-); Pax6(fl/+) mice.

In utero exposure to low doses of environmental pollutants disrupts fetal ovarian development in sheep.

Epidemiological studies of the impact of environmental chemicals on reproductive health demonstrate consequences of exposure but establishing causative links requires animal models using 'real life' in utero exposures. We aimed to determine whether prolonged, low-dose, exposure of pregnant sheep to a mixture of environmental chemicals affects fetal ovarian development. Exposure of treated ewes (n = 7) to pollutants was maximized by surface application of processed sewage sludge to pasture.