SHEAR STRESS IS SHEAR STRESS IS DIFFERENTIALLY DIFFERENTIALLY

REGULATED REGULATED AMONG INBRED AMONG INBRED

RAT STRAINSRAT STRAINSJamila Ibrahim PhD, Jody K. Jamila Ibrahim PhD, Jody K.

Miyashiro PhD, Bradford C. Berk Miyashiro PhD, Bradford C. Berk MD, PhDMD, PhD

Circ Res, May 2003Circ Res, May 2003

Importance of Vascular Importance of Vascular Remodelling (1)Remodelling (1)

Vascular remodelling is considered to play a central Vascular remodelling is considered to play a central role in the aetiology of major clinical disorders such role in the aetiology of major clinical disorders such as atherosclerosis, restenosis and hypertensionas atherosclerosis, restenosis and hypertension

Remodelling is an important determinant of blood Remodelling is an important determinant of blood flow in vascular disease. At sites of atherosclerotic flow in vascular disease. At sites of atherosclerotic plaque formation, outward remodelling restricts plaque formation, outward remodelling restricts while inward remodelling accelerates luminal while inward remodelling accelerates luminal narrowing thereby compromising blood flow narrowing thereby compromising blood flow

Remodelling at sites of atherosclerosis and plaque Remodelling at sites of atherosclerosis and plaque accumulation is critically important clinically and is accumulation is critically important clinically and is likely to occur by some of the same pathways as likely to occur by some of the same pathways as flow-induced remodellingflow-induced remodelling

Regulation of Shear Regulation of Shear StressStress

Blood flow through the vasculature generates Blood flow through the vasculature generates shear stress, a tangential dragging force shear stress, a tangential dragging force exerted by flow on the endothelial surface of exerted by flow on the endothelial surface of blood vesselsblood vessels

In accordance with Poisseulle’s Law, blood In accordance with Poisseulle’s Law, blood vessels subjected to abnormal flow and shear vessels subjected to abnormal flow and shear stress will respond to normalise this stress by stress will respond to normalise this stress by regulating vascular diameter, by a process regulating vascular diameter, by a process known as vascular remodelingknown as vascular remodeling

Increased shear stress is normalised by Increased shear stress is normalised by outward vascular remodelling and reduced outward vascular remodelling and reduced shear stress by inward vascular remodellingshear stress by inward vascular remodelling

Vascular remodelling at Vascular remodelling at sites of plaque formationsites of plaque formation

Reduced Flow Increased Flow

Plaque formationPlaque free vessel

Cross-section ofBlood Vessel

Importance of Vascular Importance of Vascular RemodellingRemodelling

(2)(2) Shear stress regulation by vascular Shear stress regulation by vascular

remodelling is likely to be complex remodelling is likely to be complex and polygenic and therefore efforts to and polygenic and therefore efforts to identify the key regulatory genes identify the key regulatory genes require appropriate animal models require appropriate animal models

The ability of vessels to compensate The ability of vessels to compensate for plaque burden, a process that for plaque burden, a process that requires outward remodelling, may requires outward remodelling, may critically depend on specific genescritically depend on specific genes

Experimental model (1)Experimental model (1) We have developed a rat model We have developed a rat model

involving ligation of the external and involving ligation of the external and internal carotid arteries, as depicted internal carotid arteries, as depicted in the schematic belowin the schematic below

Left Common Carotid Artery

ExternalCarotid Artery

InternalCarotid Artery

Occipital Artery

Superior Thyroid Artery

Experimental model (2)Experimental model (2) This model is well tolerated by rats, This model is well tolerated by rats,

allowing a surgical success rate of 100%allowing a surgical success rate of 100% Flow is reduced in the left common Flow is reduced in the left common

carotid artery by ~90% of baseline flow, carotid artery by ~90% of baseline flow, and flow in the right common carotid and flow in the right common carotid artery is ~150% of baseline flow following artery is ~150% of baseline flow following arterial ligationarterial ligation

We have examined the influence of We have examined the influence of 28days of chronic flow alteration by 28days of chronic flow alteration by arterial ligation on vessel diameter, blood arterial ligation on vessel diameter, blood flow, and shear stress flow, and shear stress

FIGURE 2C

% ∆

She

ar st

ress

(NT

G)

Right carotid arteries(high flow)

Left carotid arteries(low flow)

% ∆

She

ar st

ress

(NT

G)

Percent change in shear stress following chronic flow alteration

-80

-60

-40

-20

0

GH Fischer SHR-SP BN-100

-20

0

20

40

GH Fischer SHR-SP BN

80

60

-40

-60

GH = genetically hypertensive ratsSHR-SP = stroke-prone spontaneously hypertensive ratsBN = brown norway ratsNTG = nitroglycerin

FIGURE 3B

Left carotid arteries(low flow)

% ∆

Out

er d

iam

eter

(NT

G)

GH Fischer SHR-SP BN

-25

-20

-15

-10

-5

0

5

10

-30

-35

% ∆

Out

er d

iam

eter

(NT

G)

SHR-SP BNGH Fischer

0

5

10

15

20

25

30

35

40

45

Right carotid arteries(high flow)

Percent change in outer diameter following chronic flow alteration

GH = genetically hypertensive ratsSHR-SP = stroke-prone spontaneously hypertensive ratsBN = brown norway ratsNTG = nitroglycerin

∆ Flow, ml/min

∆ Flow, ml/min

SHR-SP

GH

∆ Sh

ear

stre

ss, d

yne/

cm2

FIGURE 4A

-20

-15

-10

-5

0

5

10

15

20

Control

+ NTG

SHR-SP

GH

∆ Sh

ear

stre

ss, d

yne/

cm2

-20

-15

-10

-5

0

5

10

15

20

1 3 5

2 4 6

2 4 6

1 3 5

Again, for a given change in flow, GH and SHR-SP rats exhibit different shear stress responses. GH rat vessels are relatively more responsive to NTG than SHR-SP vessels.

Relationships between flow and shear stress in RCA (increased flow)

For a given change in flow, GH and SHR-SP rats exhibit different shear stress responses. GH rats are better regulators of shear stress than SHR-SP.

FIGURE 4B

∆ Flow, ml/min

∆ Flow, ml/min

∆ O

uter

dia

met

er, m

m∆

Out

er d

iam

eter

, mm

Control

0

0.1

0.2

0.3

0.4

0.5

0.6

0 1 2 3 4 5 6

SHR-SP

GH

+ NTG

0

0.1

0.2

0.3

0.4

0.5

0.6

0 1 2 3 4 5 6

SHR-SP

GH

Relationships between flow and outer diameter in RCA (increased flow)

The slopes of the lines do not differ from control (upper) for GH and SHR-SP in the presence of NTG, indicating that the vascular structural changes are not dependent on differences in vascular tone

GH rat outer diameters are markedly more sensitive to flow than SHR-SP outer diameters

Carotid artery endothelial cells shown en face following immunostaining for eNOS. After 28 days of increased blood flow, there was an increase in eNOS expression in GH carotid arteries compared with carotid arteries exposed to control flow (compare b with f). In SHR-SP carotid arteries, however, increased flow produced a relatively attenuated increase in eNOS expression compared with corresponding controls (compare d with e). Moreover, GH RCA express greater eNOS compared with SHR-SP RCA following chronic flow increase (b vs. d).

Major findings (1)Major findings (1) Shear stress regulation and vascular remodelling Shear stress regulation and vascular remodelling

following chronic flow alteration vary considerably following chronic flow alteration vary considerably between rat strainsbetween rat strains

We identified a rat strain with a poor ability to We identified a rat strain with a poor ability to regulate shear stress (SHR-SP), and a strain with a regulate shear stress (SHR-SP), and a strain with a good ability to regulate shear stress (GH)good ability to regulate shear stress (GH)

These findings suggest SHR-SP and GH rat strains These findings suggest SHR-SP and GH rat strains may exhibit:may exhibit: differential sensitivity to chronic blood flow differential sensitivity to chronic blood flow

alterationalteration differences in signal transduction mechanisms differences in signal transduction mechanisms

activated by flow activated by flow differences in the chronic vascular response to differences in the chronic vascular response to

flow changes that mediate remodelling of the flow changes that mediate remodelling of the blood vesselblood vessel

Major findings (2)Major findings (2) Further work utilising these rat strains and Further work utilising these rat strains and

experimental model will permit identification of experimental model will permit identification of the key regulatory mechanisms and genes the key regulatory mechanisms and genes participating in flow mediated physiological participating in flow mediated physiological processes such as shear stress and vascular processes such as shear stress and vascular remodellingremodelling

Specifically, identification of genes that augment Specifically, identification of genes that augment or restrict remodelling is now possible. In the long or restrict remodelling is now possible. In the long term, this strategy may determine novel term, this strategy may determine novel therapeutic targets to augment remodelling and therapeutic targets to augment remodelling and restore blood flow, and thus improve the restore blood flow, and thus improve the management and treatment of cardiovascular or management and treatment of cardiovascular or cerebrovascular disease with a critical vascular cerebrovascular disease with a critical vascular remodelling componentremodelling component