Endothelial FN (Fibronectin) Deposition by α5β1 Integrins Drives Atherogenic Inflammation

Al‐Yafeai, Zaki; Yurdagul, Arif; Peretik, Jonette M.; Alfaidi, Mabruka; Murphy, Patrick A.; Orr, A. Wayne

doi:10.1161/atvbaha.118.311705

Cited by 60 publications

(50 citation statements)

References 46 publications

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“…At atherosclerosis prone regions, NF-κB signaling plays a major role in driving endothelial activation 4 . Signaling through α5β1 mediates NF-κB activation and enhanced proinflammatory gene expression in response to both oxLDL 8,9 and shear stress 16,[29][30][31][32] . Since integrin inhibitors (small molecule inhibitors, blocking antibodies) blunt shear stress and oxLDL-induced NF-κB activation, we and others have postulated that new integrin-matrix interactions likely drive endothelial responses to both stimuli.…”

Section: Talin1 L325r Mutation Blocks Endothelial Inflammation On Fibmentioning

confidence: 99%

“…In addition to inflammation, α5β1 integrins regulate fibronectin deposition in response to oxLDL and shear stress 9,32 . However, it is unclear whether basal integrin activation or inducible integrin activation is required to mediate fibronectin matrix deposition.…”

Section: Aberrant Fibronectin Fibrillogenesis In Endothelial Cells Exmentioning

confidence: 99%

“…However, disturbed flow-induced endothelial activation is significantly enhanced in the presence of a fibronectin-rich matrix, whereas endothelial activation is reduced on basement membrane proteins (collagen IV, laminin) [5][6][7] . Additionally, matrix content plays a vital role in oxidized LDL (oxLDL)-mediated proinflammatory responses, where a fibronectin-rich matrix enhances endothelial activation in response to oxLDL 8,9 . Integrins, the largest family of receptors for extracellular matrix (ECM) proteins, are heterodimers of α and β subunits that mediate both celldependent extracellular matrix remodeling and matrix-dependent changes in cell phenotype 10 .…”

Section: Introductionmentioning

confidence: 99%

“…Integrins, the largest family of receptors for extracellular matrix (ECM) proteins, are heterodimers of α and β subunits that mediate both celldependent extracellular matrix remodeling and matrix-dependent changes in cell phenotype 10 . Blocking fibronectin-binding integrins suppresses endothelial activation by oxLDL and by disturbed flow both in vitro and in vivo 8,9,11,12 , indicating an important role for these integrin in this context. Therefore, cell-matrix interactions serve as an essential component of early atherogenic endothelial activation.…”

Section: Introductionmentioning

confidence: 99%

“…The fibronectin-binding integrin α5β1 plays a clear role in atherogenic endothelial activation. Atheroprone areas show elevated α5 mRNA and protein expression 14,15 , and both disturbed flow and oxLDL induce 51-dependent fibronectin matrix assembly and endothelial proinflammatory responses including NF-B activation and ICAM-1/VCAM-1 expression 8,9,16 . Showing selective deletion of α5 integrins in the endothelium reduces atherogenic inflammation, and mice expressing an α5/α2 integrin chimera similarly reduces early atherosclerotic plaque formation and atherogenic inflammation, indicating an important role for 51 integrin signaling in early atherosclerosis 16 .…”

Section: Introductionmentioning

confidence: 99%

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Integrin Affinity Modulation Critically Regulates Atherogenic Endothelial Activation in vitro and in vivo

Al‐Yafeai

Peretik

Pearson

et al. 2020

Preprint

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While vital to platelet and leukocyte adhesion, integrin activation's role in adherent cells remains controversial. In endothelial cells, atheroprone hemodynamics and oxidized lipoproteins promote endothelial integrin activation in vitro, and integrin inhibitors reduce endothelial activation to these stimuli in vitro and in vivo. However, integrin activation's importance to endothelial phenotype remains unknown. We now show that integrin activation-deficient endothelial cells (talin1 L325R) adhere and spread, suggesting that integrin activation is dispensable for endothelial cell adhesion. However, talin1 L325R endothelial cells fail to support integrin activation, fibronectin deposition, and proinflammatory responses to atheroprone hemodynamics and oxidized lipoproteins. Rescuing integrin activation in talin1 L325R cells partially restores fibronectin deposition, whereas NF-B activation and maximal fibronectin deposition require both integrin activation and other integrin-independent signaling.Similarly, atheroprone hemodynamics fail to promote inflammation and macrophage recruitment in endothelial-specific talin1 L325R mice. These studies demonstrate a vital role for integrin activation in regulating endothelial phenotype. Materials and MethodsCell culture-Mouse lung endothelial cells were isolated from Talin1 fl/L325R mice (gift of Brian Petrich, Emory, Atlanta, GA) as previously described 21 ). Briefly, lung tissue was harvested, minced, and pushed through a 16G needle and subjected to enzymatic digestion to obtain a single cell suspension. After sorting with magnetic beads coupled to ICAM2 antibodies (eBiosource), cells were transformed using a retrovirus expressing temperature-sensitive large T-antigen. The floxed allele of talin1 was deleted using adenovirus harboring either GFP (green fluorescence protein)-Cre or GFP alone and then sorted for GFP positivity. Lung endothelial cells were grown in DMEM with 10% fetal bovine serum, 1% penicillin/streptomycin, and 1% glutamax. For endothelial shear stress treatments, cells were plated on 38 x 75 mm glass slides (Corning) at confluency, and slides were assembled into a parallel plate flow chamber as previously described 24 . For acute shear stress, cells were exposed to 12 dynes/cm2 flow for up to 1 hour. For chronic oscillatory flow (model of disturbed flow), cells were exposed to ±5 dynes/cm2 with 1 dyne/cm2 forward flow for media exchange. Human LDL (Intracell) was oxidized by dialysis in PBS containing 13.8 M Cu2SO4 followed by 50 M EDTA as previously described 25 . Computationally designed transmembrane α-helical peptides (CHAMP) peptides were a gift of Dr. William DeGrado (University of California -San Francisco) 26 .Western blot-Cell lysis and western blot was done as previously described 25 . Briefly, cells were lysed in 2X laemmli buffer, separated by SDS-PAGE gels and then transferred to polyvinylidene difluoride (PVDF) membranes (Bio-Rad, Hercules, CA). Consequently, membranes were blocked in 5% dry milk in TBST for an hour, then incubated with pr...

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Section: Talin1 L325r Mutation Blocks Endothelial Inflammation On Fibmentioning

confidence: 99%

Section: Aberrant Fibronectin Fibrillogenesis In Endothelial Cells Exmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Integrin Affinity Modulation Critically Regulates Atherogenic Endothelial Activation in vitro and in vivo

Al‐Yafeai

Peretik

Pearson

et al. 2020

Preprint

Self Cite

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Rare genetic variation in fibronectin 1 (FN1) protects against APOEε4 in Alzheimer’s disease

Bhattarai,

Gunasekaran,

Belloy

et al. 2024

Acta Neuropathol

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The risk of developing Alzheimer’s disease (AD) significantly increases in individuals carrying the APOEε4 allele. Elderly cognitively healthy individuals with APOEε4 also exist, suggesting the presence of cellular mechanisms that counteract the pathological effects of APOEε4; however, these mechanisms are unknown. We hypothesized that APOEε4 carriers without dementia might carry genetic variations that could protect them from developing APOEε4-mediated AD pathology. To test this, we leveraged whole-genome sequencing (WGS) data in the National Institute on Aging Alzheimer's Disease Family Based Study (NIA-AD FBS), Washington Heights/Inwood Columbia Aging Project (WHICAP), and Estudio Familiar de Influencia Genetica en Alzheimer (EFIGA) cohorts and identified potentially protective variants segregating exclusively among unaffected APOEε4 carriers. In homozygous unaffected carriers above 70 years old, we identified 510 rare coding variants. Pathway analysis of the genes harboring these variants showed significant enrichment in extracellular matrix (ECM)-related processes, suggesting protective effects of functional modifications in ECM proteins. We prioritized two genes that were highly represented in the ECM-related gene ontology terms, (FN1) and collagen type VI alpha 2 chain (COL6A2) and are known to be expressed at the blood–brain barrier (BBB), for postmortem validation and in vivo functional studies. An independent analysis in a large cohort of 7185 APOEε4 homozygous carriers found that rs140926439 variant in FN1 was protective of AD (OR = 0.29; 95% CI [0.11, 0.78], P = 0.014) and delayed age at onset of disease by 3.37 years (95% CI [0.42, 6.32], P = 0.025). The FN1 and COL6A2 protein levels were increased at the BBB in APOEε4 carriers with AD. Brain expression of cognitively unaffected homozygous APOEε4 carriers had significantly lower FN1 deposition and less reactive gliosis compared to homozygous APOEε4 carriers with AD, suggesting that FN1 might be a downstream driver of APOEε4-mediated AD-related pathology and cognitive decline. To validate our findings, we used zebrafish models with loss-of-function (LOF) mutations in fn1b—the ortholog for human FN1. We found that fibronectin LOF reduced gliosis, enhanced gliovascular remodeling, and potentiated the microglial response, suggesting that pathological accumulation of FN1 could impair toxic protein clearance, which is ameliorated with FN1 LOF. Our study suggests that vascular deposition of FN1 is related to the pathogenicity of APOEε4, and LOF variants in FN1 may reduce APOEε4-related AD risk, providing novel clues to potential therapeutic interventions targeting the ECM to mitigate AD risk.

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Depletion of Arg/Abl2 improves endothelial cell adhesion and prevents vascular leak during inflammation

Amado-Azevedo¹,

Stalborch

Valent³

et al. 2021

Angiogenesis

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Endothelial barrier disruption and vascular leak importantly contribute to organ dysfunction and mortality during inflammatory conditions like sepsis and acute respiratory distress syndrome. We identified the kinase Arg/Abl2 as a mediator of endothelial barrier disruption, but the role of Arg in endothelial monolayer regulation and its relevance in vivo remain poorly understood. Here we show that depletion of Arg in endothelial cells results in the activation of both RhoA and Rac1, increased cell spreading and elongation, redistribution of integrin-dependent cell-matrix adhesions to the cell periphery, and improved adhesion to the extracellular matrix. We further show that Arg is activated in the endothelium during inflammation, both in murine lungs exposed to barrier-disruptive agents, and in pulmonary microvessels of septic patients. Importantly, Arg-depleted endothelial cells were less sensitive to barrier-disruptive agents. Despite the formation of F-actin stress fibers and myosin light chain phosphorylation, Arg depletion diminished adherens junction disruption and intercellular gap formation, by reducing the disassembly of cell-matrix adhesions and cell retraction. In vivo, genetic deletion of Arg diminished vascular leak in the skin and lungs, in the presence of a normal immune response. Together, our data indicate that Arg is a central and non-redundant regulator of endothelial barrier integrity, which contributes to cell retraction and gap formation by increasing the dynamics of adherens junctions and cell-matrix adhesions in a Rho GTPase-dependent fashion. Therapeutic inhibition of Arg may provide a suitable strategy for the treatment of a variety of clinical conditions characterized by vascular leak.

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Endothelial FN (Fibronectin) Deposition by α5β1 Integrins Drives Atherogenic Inflammation

Cited by 60 publications

References 46 publications

Integrin Affinity Modulation Critically Regulates Atherogenic Endothelial Activation in vitro and in vivo

Integrin Affinity Modulation Critically Regulates Atherogenic Endothelial Activation in vitro and in vivo

Rare genetic variation in fibronectin 1 (FN1) protects against APOEε4 in Alzheimer’s disease

Depletion of Arg/Abl2 improves endothelial cell adhesion and prevents vascular leak during inflammation

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