Stabilization of brain microvascular endothelial barrier function by shear stress involves VE‐cadherin signaling leading to modulation of pTyr‐occludin levels

Abstract

Blood-brain barrier (BBB) regulation involves the coordinated interaction of intercellular adherens and tight junctions in response to stimuli. One such stimulus, shear stress, has been shown to upregulate brain microvascular endothelial cell (BMvEC) barrier function, although our knowledge of the signaling mechanisms involved is limited. In this article, we examined the hypothesis that VE-cadherin can transmit shear signals to tight junction occludin with consequences for pTyr-occludin and barrier function. In initial studies, chronic shear enhanced membrane localization of ZO-1 and claudin-5, decreased pTyr-occludin (in part via a dephostatin-sensitive mechanism), and reduced BMvEC permeability, with flow reduction in pre-sheared BMvECs having converse effects. In further studies, VE-cadherin inhibition (VE-cad ΔEXD) blocked shear-induced Rac1 activation, pTyr-occludin reduction, and barrier upregulation, consistent with an upstream role for VE-cadherin in transmitting shear signals to tight junctions through Rac1. As VE-cadherin is known to mediate Rac1 activation via Tiam1 recruitment, we subsequently confirmed that Tiam1 inhibition (Tiam1-C580) could elicit effects similar to VE-cad ΔEXD. Finally, the observed attenuation of shear-induced changes in pTyr-occludin level and barrier phenotype following Rac1 inhibition (NSC23766, T17N) establishes a downstream role for Rac1 in this pathway. In summary, we describe for the first time in BMvECs a role for VE-cadherin in the transmission of physiological shear signals to tight junction occludin through engagement of Tiam1/Rac1 leading to barrier stabilization. A downstream role is also strongly indicated for a protein tyrosine phosphatase in pTyr-occludin modulation. Importantly, these findings suggest an important route of inter-junctional signaling cross-talk during BBB response to flow.

Effect of cellular senescence on the albumin permeability of blood-derived endothelial cells

“…Correspondingly, exposure of confluent monolayers to shear stress for 24 h led to a decrease in tyrosine phosphorylation and an associated decrease in permeability (49). Incubation of ECs with the protein tyrosine phosphatase activity inhibitor dephostatin led to smaller reductions in endothelial permeability after exposure to shear stress (49). In the present study, we found that there was significantly less phosphorylated occludin in hCB-ECs compared with HAECs, which is consistent with the reduced permeability of hCB-ECs.…”

“…Treatment of human umbilical vein ECs with tyrosine phosphatase inhibitors led to selective occludin proteolysis and redistribution and increased permeability (47). When bovine brain microvascular ECs were exposed to shear stress for just a few hours, hydraulic permeability and the phosphorylation of tyrosine sites in occludin increased (49). Correspondingly, exposure of confluent monolayers to shear stress for 24 h led to a decrease in tyrosine phosphorylation and an associated decrease in permeability (49).…”

“…When bovine brain microvascular ECs were exposed to shear stress for just a few hours, hydraulic permeability and the phosphorylation of tyrosine sites in occludin increased (49). Correspondingly, exposure of confluent monolayers to shear stress for 24 h led to a decrease in tyrosine phosphorylation and an associated decrease in permeability (49). Incubation of ECs with the protein tyrosine phosphatase activity inhibitor dephostatin led to smaller reductions in endothelial permeability after exposure to shear stress (49).…”

Thrombomodulin and the vascular endothelium: insights into functional, regulatory, and therapeutic aspects

“…It exhibits considerable phenotypic heterogeneity across different vascular beds (4), its prime function being to regulate systemic blood flow and tissue perfusion rates through adjustment of vessel diameter and vascular tone (closely interacting with the underlying smooth muscle cells and pericytes), with a vasodilatory phenotype predominating in healthy vessels (52). It also acts as a highly selective barrier controlling the paracellular (and transcellular) exchange of fluids, ions, and macromolecules between circulating blood and tissues (11), a homeostatic function maintained by the coordinated interaction of interendothelial adherens and tight junction complexes (172). In a further dimension to this barrier role, endothelial cells (which normally maintain an anti-inflammatory phenotype) can mediate the recruitment and extravasation (diapedesis) of proinflammatory leukocytes to areas of tissue damage and infection via the lumenal expression/release of cell adhesion molecules (selectins, integrins, PECAM-1) and cytokines (TNF-␣, IL-1, IL-6, MCP-1) (33, 41).…”

Constitutive excessive erythrocytosis causes inflammation and increased vascular permeability in aged mouse brain

“…Indeed recent work suggests that shear stress may be the most critical factor affecting gene expression in endothelial cells and during inflammation. Data from in vitro (Stone et al, 2003;Walsh et al;Warabi et al, 2004) and in vivo (Xu et al, 2004;Yamawaki et al, 2003) studies indicate that low or low oscillating endothelial shear stress upregulates molecular and vascular responses that may be instrumental to progression of vascular-related diseases. Excessive erythrocytosis compromises vascular homeostasis as a result of significantly increased hematocrit and decreased plasma levels (Neunteufl et al, 2001).…”