bromochloroacetic-acid and Aniridia

PAX6 heterozygosity (PAX6(+/-)) causes aniridia and aniridia-related keratopathy (ARK) in humans, but the pathway from gene dosage deficiency to clinical disease has not been fully characterized. Recently, the authors suggested a model of a chronic wound state exacerbated by oxidative stress, showed the barrier function of Pax6(+/-) corneas is compromised and suggested Pax6(+/-) corneas show the molecular signature of a perpetual wound-healing state.. Pax6(+/-) mice were used as a model for Pax6-related corneal diseases and in vivo wound-healing assays. Immunohistochemistry and electron microscopy analyses were performed on mutant and wounded corneas.. This work reports defects in keratin, desmoplakin, and actin-based cytoskeletal structures in Pax6(+/-) cells. During wild-type corneal reepithelialization, cell fissures and desquamation, intracellular vesicles, intercellular gaps, and filopodialike structures were apparent, similar to the phenotypes seen in "unwounded" Pax6(+/-) corneal epithelia. Pax6(+/-) cells and wounded wild-type cells showed changed patterns of desmoplakin and actin localization. Protein oxidation and ERK1/2 and p38 MAPK phosphorylation were barely detected in the basal cells of intact wild-type corneal epithelia, but they were found in basal wild-type cells near the wound edge and throughout Pax6(+/-) corneal epithelia.. These data show that cell junctions and cytoskeleton organization are dynamically remodeled in vivo by wounding and in Pax6(+/-) corneas. This apparent wound-healing phenotype contributes to the clinical aspects of ARK.

Topics: Actins; Animals; Aniridia; Cell Adhesion; Cells, Cultured; Corneal Diseases; Cytoskeletal Proteins; Desmoplakins; Epithelium, Corneal; Eye Injuries; Eye Proteins; Female; Homeodomain Proteins; Keratins; Male; Mice; Mice, Inbred CBA; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase 6; p38 Mitogen-Activated Protein Kinases; Paired Box Transcription Factors; PAX6 Transcription Factor; Phosphorylation; Repressor Proteins

Heterozygosity for a PAX6 deficiency (PAX6+/-) results in low levels of the PAX6 transcription factor and causes aniridia. Corneal changes in aniridia-related keratopathy (ARK) include peripheral pannus and epithelial abnormalities, which eventually result in corneal opacity and contribute to visual loss. The corneal abnormalities of Pax6+/- mice provide an excellent model for the corneal changes seen in PAX6+/- humans. The aim of the present study was to investigate the contributions of different factors (including altered cell proliferation, abnormal epithelial differentiation and incursion of the conjunctival epithelium) that may underlie the pathogenesis of the corneal changes caused by low levels of Pax6 in heterozygous Pax6+/Sey-Neu (Pax6+/-) mice. BrdU incorporation showed enhanced proliferation of Pax6+/- corneal epithelium compared to wild-type controls and analysis of p63 (a marker of high proliferative potential) revealed a slight increase in frequency of p63-positive basal corneal epithelial cells in Pax6+/- mice. Immunohistochemical investigation of K12 (a Pax6-regulated marker of corneal epithelial differentiation) in 2-52-week-old mice showed that K12 expression was delayed and down-regulated in the Pax6+/- corneal epithelium, implying that differentiation of the Pax6+/- corneal epithelium was delayed and abnormal. Goblet cells were identified within the peripheral corneal epithelium of the Pax6+/- eyes, but some were surrounded by cells expressing K12, suggesting they may have arisen in situ in the corneal epithelium. These findings suggest that low levels of Pax6 may be directly responsible for failure or delay of proper differentiation of the corneal epithelial cells, but the proliferative component of the mutant epithelium is probably not impaired. This abnormal differentiation suggests that ARK is not entirely attributable to a limbal stem cell deficiency.

Topics: Animals; Aniridia; Cell Differentiation; Cell Proliferation; Corneal Diseases; Disease Models, Animal; Epithelium, Corneal; Eye Proteins; Homeodomain Proteins; Keratin-12; Keratins; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Mutant Strains; Paired Box Transcription Factors; PAX6 Transcription Factor; Phosphoproteins; Repressor Proteins; Trans-Activators

To investigate corneal abnormalities in heterozygous Pax6(+/Sey-Neu) (Pax6(+/-), small eye) mice and compare them with aniridia-related keratopathy in PAX6(+/-) patients.. Fetal and postnatal corneal histopathology, adult corneal thickness, and the distribution of K12-immunostained cells were compared in wild-type and Pax6(+/-) mice.. Prenatally, the corneal epithelium was thinner in Pax6(+/-) fetuses than wild-type littermates, but the stroma appeared irregular, hypercellular, and thickened. The anterior chamber angle was obliterated, and the iris was hypoplastic from early developmental stages. The adult Pax6(+/-) corneal epithelium was thinner, had fewer layers, and included goblet cells, indicating repopulation from conjunctival epithelium. The ocular surface was often roughened, with epithelial vacuolation and lens tissue within the stroma. The corneal stroma was thicker centrally, with an irregular lamellar alignment. Many adult Pax6(+/-) corneas were vascularized or contained cellular infiltrates, but some remained clear. Corneal degeneration was age-related: Older Pax6(+/-) mice had prominent subepithelial pannus and more goblet cells in the peripheral corneal epithelium. Cytokeratin 12 stained very weakly in the peripheral and superficial corneal epithelium in 12-month-old Pax6(+/-) mice.. Corneal abnormalities in Pax6(+/-) mice are similar to those in aniridia-related keratopathy in PAX6(+/-) patients. This extends the relevance of this mouse model of human aniridia to include corneal abnormalities. Incursion of goblet cells suggests impaired function of Pax6(+/-) limbal stem cells, abnormal expression of cytokeratin 12 may result in greater epithelial fragility, and corneal opacities in older mice may reflect poor wound-healing responses to accumulated environmental insults.

Topics: Animals; Aniridia; Anterior Eye Segment; Cornea; Corneal Diseases; Disease Models, Animal; Eye Proteins; Female; Homeodomain Proteins; Humans; Immunoenzyme Techniques; Keratins; Male; Mice; Microphthalmos; Paired Box Transcription Factors; PAX6 Transcription Factor; Repressor Proteins

A candidate tumor suppressor gene, WT-1, is believed to have an important role in the pathogenesis of Wilms' tumor, especially that occurring in patients with congenital aniridia.. To obtain a stable tumor line to work with, Wilms' tumor tissue was serially transplanted in athymic nude mice. Biopsied Wilms' tumor tissue, derived from an aniridia patient, was transplanted subcutaneously to an athymic nude mouse, and then transplanted serially. Histopathologic and molecular biologic studies were performed on the xenotransplants.. The aniridia patient showed partial deletion in one short arm of chromosome 11, which bears the WT-1 gene. The tumor was successfully transplanted in the nude mouse. Although the tumor contained blastemic, organoid, and stromal histologic elements, the organoid element began to decrease after more than 20 passages. Cytogenetic analysis revealed an additional abbreviation of one long arm of chromosome 6. Dot blot analysis showed that the copy number of WT-1 gene was decreased to half the amount in the tumor, in spite of the WT-1 transcript with normal size detected by Northern blotting.. The tumor is expected to bear one WT-1 gene with minute abnormalities as well as one congenitally deleted gene. This tumor line is useful when examining the effect caused by introduction of WT-1 gene to Wilms' tumor in vivo.

Topics: Alleles; Animals; Aniridia; Biopsy; Chromosome Deletion; Chromosome Mapping; Chromosomes, Human, Pair 11; Genes, Wilms Tumor; Humans; Immunohistochemistry; Infant; Keratins; Kidney Neoplasms; Male; Mice; Mice, Nude; Microscopy, Electron; Mucin-1; Mucoproteins; Neoplasm Proteins; S100 Proteins; Transplantation, Heterologous; Uromodulin; Vimentin; Wilms Tumor