Basement membrane stiffening promotes retinal endothelial activation associated with diabetes

Abstract

Endothelial activation is a hallmark of the high-glucose (HG)-induced retinal inflammation associated with diabetic retinopathy (DR). However, precisely how HG induces retinal endothelial activation is not fully understood. We hypothesized that HG-induced upregulation of lysyl oxidase (LOX), a collagen-cross-linking enzyme, in retinal capillary endothelial cells (ECs) enhances subendothelial basement membrane (BM) stiffness, which, in turn, promotes retinal EC activation. Diabetic C57BL/6 mice exhibiting a 70 and 50% increase in retinal intercellular adhesion molecule (ICAM)-1 expression and leukocyte accumulation, respectively, demonstrated a 2-fold increase in the levels of BM collagen IV and LOX, key determinants of capillary BM stiffness. Using atomic force microscopy, we confirmed that HG significantly enhances LOX-dependent subendothelial matrix stiffness in vitro, which correlated with an ∼2.5-fold increase in endothelial ICAM-1 expression, a 4-fold greater monocyte-EC adhesion, and an ∼2-fold alteration in endothelial NO (decrease) and NF-kB activation (increase). Inhibition of LOX-dependent subendothelial matrix stiffening alone suppressed HG-induced retinal EC activation. Finally, using synthetic matrices of tunable stiffness, we demonstrated that subendothelial matrix stiffening is necessary and sufficient to promote EC activation. These findings implicate BM stiffening as a critical determinant of HG-induced retinal EC activation and provide a rationale for examining BM stiffness and underlying mechanotransduction pathways as therapeutic targets for diabetic

Biofabrication Strategies and Engineered In Vitro Systems for Vascular Mechanobiology

“…[ 57,115 ] In general, higher BM elasticity leads to increased EC spreading, focal adhesion formation, traction force generation, higher EC contractility, and disruption of cell–cell junctions. [ 116 ] Stiffness‐mediated YAP/TAZ translocation to EC nuclei also occurs concurrently with changes in EC phenotype, actin cytoskeletal reorganization, and Rho‐GTPase activation. [ 98 ] Stiffness‐dependent sensitivity of ECs to growth factors, inflammatory molecules, and other small molecules also affects platelet adhesion, endothelial‐mesenchymal transition (Endo‐MT), and migration.…”

“…Obesity‐associated hypertension and insulin resistance causes increases in arterial and capillary stiffness, thickening of the basement membrane, an increase in vessel diameter, and dysregulated vasomodulation. [ 116,344 ] Vasomodulation is adversely affected by impaired vasodilation (reduced secretion of and hyposensitivity to vasodilators) or impaired vasoconstriction (elevated secretion of and hypersensitivity to vasoconstrictors), leading to reduced blood flow and reduced functional hyperemia in response to exercise, food intake, or increase in local metabolism. [ 344,346 ] This chain of events also correlates with microvascular rarefaction (reduction of blood vessel density), reduced perfusion to peripheral limbs and tissues, and hypoxia, and eventually leads to local ischemia and peripheral vascular disease.…”

“…[ 441 ] Higher blood viscosity, lower flowrate and shear stress, and high blood glucose associated with age and diabetes can cause retinal vein occlusion, basement membrane stiffening, reduced molecular transport, reduced partial oxygen pressure and local hypoxia in the inner retina. [ 116,441,442 ] Hindered bloodflow is associated with increased risk of thrombus formation, pericyte dropout, metabolic and vasomodulatory imbalance, and enhanced leukocyte adhesion and infiltration. [ 31,443–445 ] These effects can also be exacerbated with elevated PP due to hypertension.…”

“…Increased hypoxia induces proangiogenic signaling and switching of retinal vasculature from a stable, quiescent phenotype to an activated, angiogenic phenotype leading to neovascular sprouting and BRB dysfunction, which ultimately lead to diabetic retinopathy (DR), age‐related macular degeneration (AMD) and macular edema. [ 116,434,436 ]…”

Directional Fluid Transport across Organ–Blood Barriers: Physiology and Cell Biology

“…As in the BBB, pericytes and glial cells also participate in the establishment and maintenance of the iRBB (reviewed in (Klaassen et al 2013; Arboleda-Velasquez et al 2015)). In addition, changes in the physical properties of retinal EC basement membrane associated to diabetic retinopathy are causative of EC alterations and barrier dysfunction (Chronopoulos et al 2010; Yang et al 2016). …”