Motivation: vasculature is one of the most dynamic and inherently mechanically loaded tissues with extrinsic mechanical inputs ranging from pulsatile blood flow, shear stress and cyclic strain. These inputs arise from fluid flow and contraction activity of surrounding smooth muscle cells, pushing and pulling forces resulting from transmigration of immune cells, as well as hydrostatic compression and ECM stiffening. These dynamic forces indisputably modulate both cell-cell and cell-ECM adhesive networks of the vasculature. In that respect, vascular integrity, homeostasis, and remodeling require exquisite dynamic coordination between adhesion sites of the neighboring endothelial cells and those formed between endothelial cells and the underlying basement membranes. Vascular endothelial cadherin (VE-Cadherin) adhesion receptor is the central component of cell-cell adhesion junctions that is necessary for maintenance of a stable vascular system during development and is in control of vascular permeability and angiogenesis in adult vasculature.
Aims: first, perform integrated analysis of the biochemical and biophysical states of endothelial monolayers subjected to a spectrum of exogenously applied strain regimes. Second, decipher the mechano-sensory elements responsible for orchestrating the mechano-adaptation response of endothelial monolayers. Third, implicate mechanically driven signaling cascades required for maintenance of endothelial integrity.
Methods: FlexCell tension system, atomic force microscopy, confocal microscopy, and morphometric analyses were utilized to characterize and quantify the dynamic effects of differential strain regimes on VE-Cadherin junctions and mechanosignaling, and tissue-level architecture, rheology, dynamics, and tension. Mass spectrometry-based phospho-proteomic and biotin ligase tagged VE-Cadherin proteomics analyses were utilized to identify global and VE-Cadherin proximal mechanosensing proteins and siRNA approach was utilized to modulate the expression of the identified strain-regulated proteins and decipher their role in the mechano-response.
Results & conclusions: stretch-activated ion channel-triggered calcium signaling mediates endothelial mechano-response to exogenously applied strain by initiating biphasic waves of Rho and Rac signaling. This facilitates remodeling of cell-cell and cell-ECM adhesions as an immediate response to strain and subsequently promotes mechano-adaptation by modifying the biophysical state of the actin cytoskeleton.