The Renin-angiotensin System in Obesity and Vascular Diseases

Lisa A. Cassis, PhD

Professor and Chair

Department of Molecular and Biomedical Pharmacology

University of Kentucky

No relevant financial relationships exist.

Harriett Dustan

• Pioneer in clinical cardiovascular research

• 1st person to give sodium nitroprusside to humans,

• Thiazide diuretics potentiate blood pressure lowering of other antihypertensives,

• Definition of hemodynamics of primary aldosteronism,

• Pathophysiology of obesity-related hypertension

In Memoriam, Circulation 100:2122-2123, 1999

Acknowledgments: Lab/Trainees

The Ever Evolving RAS Angiotensinogen

Angiotensin I (1-10)

Renin

Angiotensin II (1-8)

ACE

Angiotensin III (2-8)

Aminopeptidase A

Angiotensin IV (3-8)

Aminopeptidase N

Ang1-9 Ang1-7 ACE2 ACE

Prorenin ?

(Pro)Renin Receptor

Activates Renin

Signals

Research Program

Angiotensin II

Adipose Tissue Atherosclerosis

AAA Hypertension

EC

Muscle

Adventitia

Adipose

Ao

Ao

Ao

Ao

Ao

Ao

Ao

Ao

Ao Ao Ao

Ao

Ao

Ao

Ao

Expression of RAS components during adipocyte differentiation

Undiff4hrs 8hrs 12hrs D1 D2 D3 D4 D5 D6 D7 D8 D9 D100.0

0.5

1.0

1.5

2.0

*

*

*

*

**

* *

*,P<0.05

AO

/18S

Undiff 4hrs 8hrs 12hrs 1d 2d 3d 4d 5d 6d 7d 8d 9d 10d0.00

0.01

0.02

0.03

0.04

**

AT

1a/1

8S

*

*,P<0.05

Angiotensinogen

AT1 receptor

ACE2

UD 4hrs8hrs12hrs D1 D2 D3 D4 D5 D6 D7 D8 D9 D100.000

0.025

0.050

0.075

0.100

***

*

*,P<0.05

AC

E2/1

8s

Undifferentiated 1 2 3 4 5 6 7 8 9 10

Cocktail

Gupte et al., AJP 295: R781-8, 2008

Prevalence Obesity* Trends Among U.S. Adults

(*BMI 30)

No data <10% 10-14% 15-19% 20-24% 25-29% >30%

Source: CDC, Behavioral Risk Factor Surveillance System, February 2010

1999 1990 2009

OBESITY

↑Angiotensin II

Hypertension

↑Angiotensinogen (AGT)

Frederique Yiannikouris, PhD

Neocassette Exon 2

Lox P Lox P FRT FRT

Lox P Lox P

Exon 3

Flp

Ap2-Cre

Lox P flanked allele

Deleted allele

Lox P

FRT

Exon 2 Exon 3 Neo

Lox P

Exon 3

a

b

c

0

10

20

30

40

Day 0 Day 8

Agtfl/fl

AgtaP2

*

*,**

**A

GT

(n

g/m

l)

AGT mRNA abundance AGT protein

Agtfl/fl, wild type, AgtaP2 adipocyte AGT deficient

Adipocyte AGT deficiency has no effect on body weight, fat mass, or glucose tolerance

A

0

10

20

30

40

50

LF HF

** *,**

*

AgtaP2

Agtfl/fl

Fat

mass (

% l

ean

)

0

10

20

30

40

50

60

70

80

LF HF

** **

Agtfl /fl

AgtaP2

Lean

mass (

% B

W)

B

0 2 4 6 8 10 12 14 160

Agtfl/fl, LF

AgtaP2, LF

Agtfl/fl, HFAgtaP2, HF

*****

* ***

***

20

30

40

50

Time

Bo

dy w

eig

ht

(g)

Agtfl/fl = controls AgtaP2 = Adipocyte AGT deficient

Adipocyte deficiency of AGT ablates obesity-hypertension

0

LF HF

Agtfl/fl

AgtaP2 *

**

100

110

120

130

140

SB

P (

mm

Hg

)

Hypertension, in press, 2012

Reductions in plasma AngII in obese adipocyte AGT-deficient mice are paralleled by reduced adipose AngII content

0

50

100

150

200

250

300

Agtfl/fl

AgtaP2

LF HF

*

**

An

g II (p

g/m

l)

0

5000

10000

15000

20000

25000

Agtfl/fl

AgtaP2

LF HF

An

gII (

pg

x t

ota

l fa

t

ma

ss

)

PLASMA ADIPOSE TISSUE

Summary

• Deficiency of AGT in adipocytes prevents obesity-induced increases in plasma AngII and obesity-related hypertension

• Tissue production of AngII (e.g., adipose) can be a significant source of circulating AngII in the setting of obesity and in the development of obesity-induced hypertension

Hypertension, in press, 2012

OBESITY

↑Angiotensin II (AGT)

Undiff 4hrs 8hrs 12hrs 1d 2d 3d 4d 5d 6d 7d 8d 9d 10d0.00

0.01

0.02

0.03

0.04

**

AT

1a/1

8S

*

*,P<0.05

Why do adipocytes have AT1aR? What does AngII do to an adipocyte? Is this influenced by obesity and does it contribute to obesity-associated diseases?

Kelly Putnam

Endocrinology, epub ahead of press, 2012

Adipocyte AT1aR

Liver Kidney Heart Brain Spleen BAT WAT0.0

0.1

0.2

AT1aRfl/fl

AT1aRaP2

*

* *

0.2

1.2

AT

1a

R:1

8s

X FLPE

LoxP3

FRT

LoxP1

Exon 3 AT1aRfl/fl

Exon 3 Neocassette

FRT FRT

LoxP1 LoxP2 LoxP3

X aP2-Cre

AT1aRaP2

Floxed AT1aR Gene

LoxP1-3

Adipocyte deletion of the AT1aR

Adipocyte AT1aR deficiency has no effect on the development of obesity

0 5 10 15

20

40

60

AT1aRaP2 HF

AT1aRfl/fl HF

AT1aRaP2 LF

AT1aRfl/fl LF

Weeks on diet

We

igh

t (g

)

0 AT1aRfl/fl

AT1aRaP2

AT1aRfl/fl

AT1aRaP2

0

20

40

60

80

100

% Lean mass

% Fat mass

LF HF

* *

% o

f b

od

y m

as

s

, but……

0

1000

2000

3000

4000

5000

6000

7000

0

20

40

60

80AT1aR

fl /fl LF

AT1aRaP2

LF

Adipocyte size (m2)

# o

f c

ell

s

LF HF0

1000

2000

3000

4000AT1aR

fl /fl

AT1aRaP2

***

Me

an

ad

ipo

cyte

siz

e (

m2)

AT1aRfl/fl

AT1aRaP2

0.0

0.2

0.4

0.6

0.8

1.0

*

Ab

so

rb

an

ce

(5

10

nm

)

AT1aRfl/fl

AT1aRaP2

0

200

400

600

*

PP

AR:1

8s

Summary

• Adipocytes have AT1aR, but they play no major role in the development of obesity

• In lean mice, deficiency of AT1aR on adipocytes decreases adipocyte differentiation, resulting in hypertrophy of remaining adipocytes

OBESITY

ACE2 AngII Ang-(1-7)

UD 4hrs8hrs12hrs D1 D2 D3 D4 D5 D6 D7 D8 D9 D100.000

0.025

0.050

0.075

0.100

***

*

*,P<0.05

AC

E2/1

8s

X

Sean Thatcher, PhD Assistant Professor

Manisha Gupte, PhD

Hypertension prevalence is greater in men than women before menopause: Are females protected

against obesity hypertension?

Age Men (%) Women (%)

20-34 9.2 2.2

35-44 21.1 12.6

45-54 36.2 36.2

55-64 50.2 54.4

65-74 64.1 70.8

75 and older 65.0 80.2

All 31.8 30.3

CDC

Menopause

0 5 10 150

515

20

25

30

35

40

45

50

55Male, LFMale, HF

Female, LF

Female, HF

*

**

**

**

* *

* *

**

**

*

Bo

dy w

eig

ht

(gm

)

Male Female0

5

10

15

20

25LF

HF *

*,**

**

Fat

mass (

g)

Female mice gain more weight and fat mass than males

65%

97%

86%

236%

*, P<0.05 compared to LF

*, P<0.05 compared to LF **, P<0.05 compared to male Male Female

0

50

100

150LF

HF **

**

SB

P N

igh

t (m

mH

g)

but are protected from obesity-hypertension

Arteriosclerosis, Thrombosis and Vascular Biology 32:1392-9, 2012

The AngII/Ang-(1-7) balance is different between obese males and females, and ACE2 deficiency promotes

hypertension in both sexes

Male Female0.0

0.2

0.4

0.6

0.8 LF

HF

***

**

Pla

sm

a A

ng

-(1-7

) (n

g/m

l)

Male Female0

50

100

150

200Ace2

+/+, HF

Ace2-/-

, HF

*,****

*

SB

P N

igh

t (m

mH

g)

Summary

• ACE2 is important in regulating the AngII/Ang-(1-7) balance in the development of obesity-hypertension

• Females rule!, they are protected against obesity-hypertension potentially through an ACE2-dependent mechanism

Research Program

Angiotensin II

Adipose Tissue Atherosclerosis

AAA Hypertension

AAA

• 13th leading cause of death in the United States

• Risk factors – Male Gender

– Smoking

– Age >65

– Family history

– Hypertension

– Obesity

• No pharmacologic treatments for AAA • Surgical repair is the only therapeutic option to prevent

rupture (> ~ 5.0 cm)

Allison et al. J Vasc Surg 2008; 48:121-7, Golledge et al. Circulation 2007; 116:2275-2279.

Angiotensin II

28 days Osmotic mini-pump

apoE-/- LDLr-/-

AAA

Atherosclerosis

AngII-induced Vascular Pathology

Daugherty A, Manning MW, Cassis LA. J Clin Invest. 2000 Jun;105(11):1605-12.

Risk Factors: Effects of Male Gender

Hypothesis:

Sex hormones mediate gender differences in AngII-induced vascular diseases by regulating the AT1a receptor

Xuan Zhang, PhD

Androgen is the primary regulator of AAA susceptibility in male mice through regulation of

aortic AT1aR

Sham Orx

% A

AA

Incid

ence

0

20

40

60

80

100

*

Male Female

% In

cid

ence

0

20

40

60

80

100

intact 1 week 5 weeks 1 week 5 week0.0

0.1

0.2

0.3

0.4

0.5

0.6

thoracic

abdominal

Castration Castration+DHT

AT

1a/1

8S

Male/Female AAA susceptibility Testosterone is the mediator

Aortic Development

Adapted from Majesky M. ATVB 2007

AR

Can we turn a female into a male with enhanced

AAA susceptibility by exposing her to testosterone

early in life?

Day 1

Testosterone (400 µg/mouse)

or Vehicle

10-12 weeks

•Aortic gene expression

•AngII infusion

Question

Circulation Research 25:373-85, 2012.

Neonatal testosterone strikingly promotes AngII-induced AAAs in adult female mice

500 750 10000

20

40

60

80vehicletestosterone

AngII infusion rate (ng/kg/min)

AA

A in

cid

en

ce

(%

)

*,**

A

C

B

vehicle testosterone

saline AngII0.0

0.5

1.0

1.5

2.0

2.5vehicletestosterone

Maxim

al exte

rnal

ab

do

min

al ao

rtic

dia

mete

r (m

m)

*,**

*

vehicle testosterone0.00

0.01

0.02

0.03

0.04

0.05thoracic

abdominalA

T1aR

/18s r

ati

o

ApoE-/-

*,**

Summary

• Testosterone has pronounced effects during development to influence the vasculature.

• AT1a receptors are a target of testosterone in smooth muscle cells to influence vascular remodeling in aneurysm formation.

• Vascular disease is sexually dimorphic.

Acknowledgments: Grant Support

• NIH HL73085 (LAC)

• NIH HL107326 (LAC)

• NIH P01 HL080100 (AD, LAC)

• NIH P42 ES007380 (BH, LAC)

• NIH T32 DK007778 (LAC)

• NIH P20 GM103527 (LAC)

• AHA: 0815419D (MG), 11PRE6760002 (KP), Pre0815513D (XZ), 12PRE12050430 (RS)

Alan Daugherty