Alterations in the corneal stroma of transgenic mice carrying a truncated collagen XII gene.

Abstract

Abstract: : Purpose: To determine the corneal dominant negative phenotype resulting from a decrease in FACIT collagen XII expression. Methods:Corneas were dissected from five 8 week old transgenic mice and two 8 week old wild type mice for RNA isolation or for fixation prior to embedding in epon. Relative reverse transcription polymerase chain reaction was used to quantitate mRNAs for collagens I, XII and XIV. Electron microscopy was performed on sectioned corneas to examine fibril diameters and fibril spacing. Results:Wild type and transgenic mice expressed normal levels of full length collagen XII mRNA. In addition, transgenic mice expressed truncated collagen XII mRNAs at levels about 75% of the normal level. The amount of collagen XII protein secreted by transgenic corneal cells was decreased. By relative PCR, fibrillar type I collagen mRNA levels were increased by 18.4%, and FACIT type XIV collagen mRNA levels by 45%. The corneal stromal fibrils in transgenic and wild type mice were both an average of 22 nm in diameter, and the range of fibril distributions was the same. However, interfibrillar spacing was dramatically altered in the transgenic mice. The average center to center distance between fibrils in the wild type mouse was 50 nm. In the transgenic mice, this distance was 42 nm, a difference of about 16%. Conclusions: The transgene caused dominant interference of molecular assembly of collagen XII chains, resulting in a decrease of secreted collagen XII protein. This resulted in corneal fibrils being more closely spaced in transgenic mouse corneas than in normal corneas. These results show that, unlike collagen XIV, collagen XII is not a fibril diameter regulator, but instead appears to play a role in the organization of neighboring fibrils.

Keywords: cornea: basic science • extracellular matrix • transgenics/knock–outs