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Yujin Zhang, Lung-Kun Yeh, Winston W Kao, Chia-Yang Liu; Wnt/b-catenin signaling controls corneal stroma maturation by repressing TGFbs expression. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2202.
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© ARVO (1962-2015); The Authors (2016-present)
Wnt/b-catenin signaling plays critical roles during the morphogenesis of multiple tissues. In this study, we examined the hypothesis that perturbation of Wnt/b-catenin signaling in corneal stromal keratocytes (Ctnnb1cskΔ/Δ or Lrp5/6 cskΔ/Δ) could impact stroma formation.
Kera-rtTA/ tet-O-Cre/Ctnnb1f/f (KR/TC/Ctnnb1f/f) triple transgenic mice were induced with doxycycline (Dox) from embryonic 12.5 (E12.5) to various gestation stages e.g., E16.5, E18.5 and postnatal day 0 (P0) and neonates were collected. The specimens were subjected to transmission electron microscopy (TEM), histology and immunohistochemistry examination. KR/TC/Lrp5cskΔ/cskΔ/Lrp6cskΔ/cskΔ quadruple mice were generated for examining the effects of interrupted Wnt signaling. Primary corneal stroma fibroblasts from the Ctnnb1f/f mice were cultured and transduced by Ad-Cre or Ad-EGFP virus. Expression profiles of cytokines after virus infection were determined by cDNA micro-array, RT-qPCR and western blotting.
Ctnnb1cskΔ/cskΔ mutant mice exhibited precocious maturation during development in comparison to that of wild-type mice at birth, P0. TEM revealed the collagen fibrils and stromal cells are better organized in mutant neonates. Moreover, cytokine cDNA array screening showed that TGFb2 and TGFb3 were up regulated by 3 and 75 fold, separately in cultured Ctnnb1cskΔ/Δ corneal fibroblasts and verified by RT-qPCR and western blotting. Likewise, CDK inhibitor p21 gene was up-regulated and the extracellular matrix (ECM) components such as Col1a2 and keratocan (Kera) were also increased. Single Lrp5cskΔ/Δ or Lrp6cskΔ/Δ mutant appeared normal, while Lrp5cskΔ/Δ/Lrp6cskΔ/Δ (Lrp5/6cskΔ/Δ ) double mutant manifested precocious corneal stroma maturation similar to the phenotype observed in Ctnnb1cskΔ/Δ mice.
Our data supports the notion that cross-talk between Wnt/β-catenin and TGFb signaling play critical roles in corneal morphogenesis during development.
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