Increased apoptosis and abnormal wound-healing responses in the heterozygous Pax6+/- mouse cornea.
OBJECTIVE
Corneal wound healing involves a cascade of interactions between the epithelium and stroma. Pax6 is upregulated, and early events include epithelial cell migration and apoptosis of superficial keratocytes. The mouse heterozygous Pax6 (Pax6+/-) corneal phenotype mimics human aniridia-related keratopathy (ARK), and some aspects of wound healing have been shown to be abnormal, including matrix metalloproteinase (MMP)-9 expression. The purpose of this study was to test whether the Pax6+/- genotype affects corneal wound-healing responses, including stromal cell apoptosis, epithelial cell migration rate, and MMP secretion in culture.


METHODS
Pax6+/- and wild-type (Pax6+/+) mice were killed and their corneas wounded by epithelial debridement. Whole eyes were cultured in organ culture and corneal epithelial healing rates and keratocyte apoptosis were quantified by topical fluorescein staining and TUNEL, respectively. Dissociated corneal epithelial cells from Pax6+/- and wild-type mice were cultured, and the activities of secreted MMP-9 were determined by zymography.


RESULTS
Wound-healing rates during the first 6 hours were significantly faster for larger wounds and for Pax6+/- corneas. Compared with wild-type, wounded Pax6+/- eyes showed significantly more stromal cell apoptosis, and cultured Pax6+/- corneal epithelial cells produced lower MMP-9 activity.


CONCLUSIONS
The cumulative effect of abnormal wound-healing responses, characterized by increased stromal cell apoptosis and reduced levels of MMP-9 secretion may contribute to the corneal changes in the Pax6+/- mice. Possible contributions of elevated stromal cell apoptosis and other abnormal wound-healing responses to ARK are discussed.
Ramaesh
Thaya
T
Division of Reproductive and Developmental Sciences, Genes and Development Group, University of Edinburgh, Scotland, United Kingdom.
Ramaesh
Kanna
K
Leask
Rosemary
R
Springbett
Anthea
A
Riley
Simon C
SC
Dhillon
Baljean
B
West
John D
JD
D000818:Animals; D015783:Aniridia / Q000235:genetics; D017209:Apoptosis; D002465:Cell Movement; D003319:Corneal Stroma / Q000473:pathology*; D019573:Epithelium, Corneal / Q000502:physiology*; D005136:Eye Proteins / Q000235:genetics*; D005260:Female; D005347:Fibroblasts / Q000473:pathology; D005786:Gene Expression Regulation / Q000502:physiology*; D005838:Genotype; D018398:Homeodomain Proteins / Q000235:genetics*; D020287:In Situ Nick-End Labeling; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008856:Microscopy, Fluorescence; D009924:Organ Culture Techniques; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors / Q000235:genetics*; D012097:Repressor Proteins / Q000235:genetics*; D014945:Wound Healing / Q000502:physiology*
D005136:Eye Proteins; D018398:Homeodomain Proteins; D000071841:PAX6 Transcription Factor; C000605730:PAX6 protein, human; D051761:Paired Box Transcription Factors; C000605732:Pax6 protein, mouse; D012097:Repressor Proteins; D020780:Matrix Metalloproteinase 9