TGF Beta-Smad3 Signaling Is Required for Epithelial-Mesenchymal Transition of Lens Epithelium

Abstract

Abstract: : Purpose: To determine the role of TGF ß-Smad3 signaling in epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs) in response to injury in mice. Methods: The animal protocol was approved by National Cancer Institute/NIH. Adult Smad3-/- (KO) and wild type (WT) mice (67 KO and 70 WT mice) were generally and topically anesthetized and the anterior lens capsule of the right eye was pierced by a hypodermic needle. After intervals of healing from 6 hr up to 8 weeks the animals were killed. The eyes were processed for immunohistochemistry for α-smooth muscle actin (αSMA), a marker of late stage EMT, lumican, collagen I and BrdU, and in situ hybridization for Snail, a marker of early stage EMT. Smad3-null lenses were organ-cultured with or without TGFß2 (10 ng/ml) for 10 days and examined similarly. Results: LECs in WT mice underwent a fibroblast-like shape change at day 5 post-injury, whereas they maintained an epithelial-like morphology up to 8 weeks post-injury in KO mice. Expression of αSMA mRNA and protein was up-regulated from day 5 until week 4 in WT, but was not detectable in KO mice at any time. Lumican and collagen I were up-regulated in healing epithelial cells of WT mice at day 1 or 3, respectively, but not in KO mice. Snail mRNA began to be expressed in LECs around the injury from day 1 in WT, but was never expressed in the cells of KO mice. In organ culture, TGFß2 induced EMT of WT LECs with αSMA expression, whereas Smad3-null LECs maintained an epithelial morphology. Conclusions: Smad3 signaling is required for EMT in LECs in response to injury in mice.

Keywords: signal transduction • growth factors/growth factor receptors • wound healing