Abstract
Purpose: :
Previous studiesa indicate integrins play important roles during murine lens development. To investigate the role of integrin signaling via ILK in lens development we localized ILK and examined the effects of conditionally deleting Ilk using the Cre/LoxP approach.
Methods: :
Immunofluorescence using a monoclonal antibody to ILK was used to examine the spatiotemporal expression of ILK during lens development (E12.5-P21). The MLR10 mouse line, which expresses Cre in fibers and epithelium, was used to conditionally delete Ilk in the developing lens. Embryonic and postnatal tissues from mutant and wild-type mice were analyzed by PCR, histology and immunofluorescence.
Results: :
ILK was detected in both lens fibers and epithelial cells from E12.5-E15.5, but was restricted to epithelial cells during fetal and postnatal development. Intercrosses of MLR10 with floxed ILK mice generated mutant mice (Cre+/ILKfl/fl) that had mild microphthalmia due to an abnormal lens. Loss of ILK protein was evident from E13.5. However, distinct lens structural changes were not evident until E17.5, with abnormal nuclear migration in lens fibres and, by P2, a dramatic loss of central, but not equatorial epithelial cells. By P10, there was extensive fibre cell vacuolisation and capsule rupture. Analyses of postnatal lenses reveal decreased cell proliferation by BrdU incorporation, cyclin D1 and phospho-histone-3 staining. Abnormal fiber differentiation was indicated by persistent Pax6 and decreased c-maf, α-, and β-crystallin, but not Prox1 expression. PAS, laminin and collagen IV staining revealed deficient ECM deposition into the lens capsule. TUNEL staining revealed increased apoptosis in the epithelium at P2 followed by widespread fiber cell death at P10.
Conclusions: :
These data indicate that Ilk plays key roles in regulating lens epithelial cell survival and proliferation. Many of the changes in the fiber cells are likely to be secondary to effects on epithelial cells in the germinative zone.a. Wederell E, de Iongh RU. (2006). Extracellular matrix and integrin signaling in lens development and cataract. Sem Cell Dev Biol, 17:759-76.
Keywords: cell adhesions/cell junctions • development • cell survival