Mechanotransduction in the Vascular System : Focus on Shear and Integrin-ECM Summer Topic Review.

Mechanotransduction in the Vascular System :Focus on Shear and Integrin-ECM

Summer Topic Review

Why do we care?

Mechanical forces sensed by the vasculature determine

blood vessel structure both acutely (through vasodilation) and chronically (though vascular remodeling)

gene expression and phenotypic status of endothelial and mural cells

Through these biological effects, mechanical forces

regulate susceptibility to inflammation and atherosclerotic plaque formation

affect endothelial cell junctions, migration, permeability and probably extravasation

Regulation of Vascular Structure by Flow

Chick Yolk Sac, Thoma 1893

•Vessels with high-flow enlarge

•Those with low-flow regress John Hunter, 1728 - 1793

Shear Stress

Well defined role in:

Vessel enlargement/regressionVasodilation (acute dilation of blood vessel)Atherosclerosis/inflammationDevelopmental vascular remodellingArteriovenous specificationAnd a suggested role in aortic hemogenesis in the embryo

Frictional force of blood flow on the vessel lining (endothelium)

Circumferential Stress

Well defined role in:

Switch from synthetic to contractile and proliferative to quiescentMyogenic tone (contraction of smooth muscle cells in response to lumen pressure)Supporting a likely role in vessel wall growth in hypertension

Outward force of blood pressure on the vessel wall (endothelium and mural cells)

Yamamoto et al. Nat. Med. 2008

Discrete Molecular Responses to Shear and Circumferential Stress

Yamamoto et al. Nat. Med. 2008

Discrete Molecular Responses to Shear and Circumferential Stress

Rabbit hindlimb angiogramEitenmüller et al. Circ. Res. 2006

Shear not Circumferential Stress Promotes Vessel GrowthSurgical Fistula

Pressure shear = increase growth

Regulation of Arteriovenous Specification

le Noble et al. Development 2004

Atherosclerotic Plaques Preferentially Develop in Areasof Chronic Low/Oscillatory Shear Stress

Dai et al.PNAS 2004

the endothelium is themediator of shear stresseffects on the vasculature

Chronic Low/Oscillatory Shear Stress Induces an “Activated” or “Atheroprone” Endothelial Phenotype

Dai et al.PNAS 2004

Pre-treatment

+ IL-1

Klf2 is Induced by Chronic Laminar Shear and Mediates Atheroprotective Gene Expression Patterns

Kush et al. JCI 2006

Dekker et al. Blood 2002

Acute Changes in Shear Regulate Permeability of the Endothelium

Electrical resistance of porcine endothelial cells in culture is reduced by removing shear stress. Resistance is further decreased by calcium chelation.

Chronic Changes in Shear Regulate Permeability of the Endothelium

1200 70

Cucullo et al. J. Cereb. Blood Flow & Met. 2007

Shear inhibits endothelial cell death upon serum starvation.

Dimmeler et al. FEBS 1996

Chronic Shear Stress Inhibits Endothelial Apoptosis

Akimoto et al. Circ. Res. 2000

Chronic Shear Inhibits Endothelial Proliferation

Acute Loss of Shear may Encourage Emboli/Cell Extravasation

Similar time-course of emboli and tumor cell extravasation in murine cerebrovasculature

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Molecular Transducers of Endothelial Shear Stress

Time-line of Shear Induced Changes

Hahn & Schwartz, Nat. Rev. Mol. & Cell. Biol. 2009

Cytoskeletal Reorganization

Increased leukocyte recruitment

Vasodilation

Chronic Shear Stress Regulates Endothelial Cell-Cell Adhesion

Noria et al. Circ Res. 1999

Porcine aortic endothelial cells upon application of FSS

Acute Shear Stress Induces Cytoskeletal Stress

Helmke, Biophys. Journal 2003

Confocal optical section of GFP-vimentinBefore and after shear

Chronic Shear Causes Re-alignment of Focal Adhesions

Davies et al. JCI 1993

Acute Shear-mediated Integrin aV/b3 Activation

Bovine aortic endothelial cells under shear for 5 minutes. Wow-1 binds activated Integrin aV/b3. b3 shows total b3 integrin.

Tzima et al. EMBO 2001

RGD Peptide Inhibits Acute Shear Induced Vasodilation

Muller et al. Circ. Res. 1997Isolated coronary arterioles

Genetic Ablation of a1 Integrin Inhibits Acute Shear Induced Vasodilation

Loufrani et al. Am. J. Phys. 2008

Bacteria-derived Peptide (pUR4/FUD) Blocks Fn Matrix Assemblyand Interferes with Vascular Remodeling In Vivo

Chiang et al. ATVB 2011

Vascular basement membranes contain

laminins (predominantly laminin-8/ laminin411 and laminin-10/ laminin511)Type IV collagens perlecannidogenscollagen XVIIIvon Willebrand factor.

In development, and during pathological remodeling additional ECM components include

FibronectinFibrinogen

“Quiescent” vs “Activated” Endothelial ECM

Orr et al. JCB 2005

Acute NF-kappa B (p65 subunit) Activation by Shear is ECM Dependent

BAE cells plated for 4 hrs were sheared for 60 min

Fn Is Deposited in Regions Chronically Exposed to Low/Oscillatory Shear

Orr et al. JCB 2005

In vitro Fn Deposition is Increased by Chronic Low/Oscillatory Shear

Feaver et al. Circ. Res. 2010

Modification of Fn Fibrils Which “Hide” EIIIA Domain Suppresses Shear-induced NF-kappa B Signaling by Increasing p38

Klein et al. JCS 2003

Orr et al. JCB 2005

EIIIA Fn isoform

total Fn

EIIIA Fn isoform

IgG control

Nor

mal

Rat

Art

ery

Wou

nded

Rat

Art

ery

Media

Intima

EIIIATotal

EIIIATotal

Increased Expression of EIIIA Isoform in Pathological Vascular Response

Protein Immunofluorescence RNA Dot Blot Hybridization

Human Atherosclerotic Lesion,10ug RNA 2x dilutions

Glukhova et al. JCB 1989

Ciliated mouse aortic endothelial cells are increase Ca++ release in response to increased shearwhen incubated in low or no-shear conditions, but not when incubated in high-shear.

Endothelial Cilia mediates Ca++ Release

Nauli et al. Circulation 2008

Shear Activates Purified G proteins imbedded in Liposomes

Cholesterol (which decreases membrane fluidity) reduces GTPase activation

Gudi et al. PNAS 1998

A Mechano-sensory Complex of VE-cadherin/VEGFR2/PECAM-1Regulates a Subset of Shear-induced Events

Tzima et al Nature 2005

Endothelial Integrin aV/b3 Activation by Shear Depends on the Mechano-sensory Complex

Ve-cadherin/PECAM1/VEGFR2

PI3 kinase

Src

aV/b3 activation (detected by WOW-1 antibody)

kinase inhibitors PP2 or SU6656

kinase inhibitors LY294002 or wortmannin

VE-cadherin-/- or PECAM-/- cells

Magnetic bead conjugated with PECAM-1 (but not VE-cadherin or CD44) induces WOW-1 signalupon mechanical pull on bovineaortic endothelial cells.

Tzima et al Nature 2005

Take-away Points

Shear stress exerts acute and chronic effects on the vasculature with important implications in human development and disease

-Acute increase promotes vasodilation-Acute and chronic decrease promotes permeability, proliferation, apoptosis-Low/oscillatory shear promotes inflammation and atherosclerosis

Acutely responsive signaling pathways critical in these responses have been identified

- PECAM/VE-cadherin/VEGFR2 complex- Integrin activation

Chronic changes have also been identified

-KLF2 -Fibronectin (EIIIA and EIIIB)

Open Questions

How do changes in ECM composition and integrin array induced by altered shear feed back into the in vivo consequences?

- New integrin adhesions are required for shear-mediate cytoskeletal reorganization- Crude pharmacologic inhibition of FN matrix assembly inhibits inflammatory receptor expression, recruitment, and proliferative response to reduced FSS

Can the different response to shear among vessels be attributed in part to differences in ECM composition?

What portion of this is genetically controlled, and what portion is environmentally controlled?