Constitutive Activation of Ras in the Lens Results in the Loss of E–Cadherin Expression and Lens Capsule Degradation in Transgenic Mice

Abstract

Abstract: : Purpose: Ras is a small GTP–binding protein in the signal transduction pathways which can be activated by growth factors. We previously reported that expression of a constitutively activated H–Ras in the lens can induce overproliferation but not premature differentiation of the lens epithelial cells in transgenic mice. The purpose of this study is 1) to report that sustained activation of H–Ras in the lens of the transgenic mice can also result in the loss of E–cadherin in lens epithelium and degradation of the lens capsule, and 2) to investigate the molecular mechanisms involved in these processes. Methods: Transgenic mice expressing a constitutive active mutant of H–Ras in lens were generated using a heterogenic lens–specific promoter which consists of the Δ1–crystallin enhancer fused to the mouse αA–crystallin promoter. Lens epithelial cell detachment from the lens capsule and breakdown of the lens capsule in the transgenic mice were analyzed by histology. E– and N–cadherin expression was examined by in situ hybridization. Semi–quantitative RT–PCR was performed to compare expression levels of matrix metalloproteinase (MMPs). Results: Three lines of transgenic mice with different expression levels for the oncogenic H–Ras were used in this study. Lens capsule breakdown and invasion of the hyaloid blood vessels into the lens of transgenic mice began at embryonic day 13 to 15. The onset of these changes was dependent on the transgene expression level. RT–PCR showed that the mRNA levels were up–regulated in the transgenic lens for MMP–(2, 9, and 14), vascular cell growth factor (VEGF) and hypoxia inducible factor (HIF)–1α. In situ hybridization showed that E–cadherin expression in the lens epithelial cells was significantly inhibited while N–cadherin expression was upregulated in transgenic mice. Conclusions: Constitutive activation of Ras disrupts the signaling pathways which are required for normal lens epithelial cell integrity and development.

Keywords: cell adhesions/cell junctions • transgenics/knock-outs • signal transduction