Under Diabetic Conditions Reactive Oxygen Species Inhibits Corneal Epithelial Cell Migration and Tight Junction Formation via Akt Signaling

Abstract

Basement membrane is composed of ECM proteins that have viscoelastic properties. When the viscoelasticity is mimicked in vitro, epithelial cells coalesce by ''dragging'' the ECM protein through the PDMS substrate. This mechanosensing of viscoelasticity is achieved through the translocation of vinculin from the focal adhesions to the cell-cell junctions and is sensitive to the level of vinculin in the cell. Apart from the composition of cell-matrix and cell-cell adhesion complexes within the cell, we find that other biophysical and biochemical cues from environment affect the cell response on a viscoelastic substrate. By varying the interfacial force between ECM protein, fibronectin, and the PDMS substrate through physisorption or covalent linkage, we found that increasing the adhesion force hinders the coalescence of cells on a viscoelastic substrate as if on a soft-elastic substrate, suggesting the role of ECM-substrate interaction for in vitro models. Also, stronger cell-cell adhesions cause coalescence when coated with either fibronectin or collagen-1 alone but not when coated with Matrigel, consisting of collagen-IV, laminin, and other ECM proteins. To gain further insights into this phenomenon, we are using quantitative super-resolution microscopy to investigate how the difference in ECM anchoring is sensed by the cell at the molecular level and traction force microscopy to quantify the ECM remodeling and substrate deformation. These results on the viscoelastic substrate would provide new insights into in vivo basement membrane and cell-cell dynamics in general, and help to have better in vitro model, mimicking in vivo ECM arrangement and viscoelasticity, where these are crucial.