Absence of S100A1 in mice is associated with a hypertensive phenotype dependent on both gender and endothelial function

Absence of S100A1 in mice is associated with a hypertensive phenotype dependent on both

gender and endothelial function

Jean-Francois Desjardins; Krystyna Kuliszewska; Adrian Quan; Duncan J. Stewart; Subodh Verma; Thomas G.

Parker

Presenter disclosure information

Jean-Francois DesjardinsAbsence of S100A1 in mice is associated with a

hypertensive phenotype dependent on both gender and endothelial function

Disclosures : none

IntroductionS100A1S100A1 Small dimeric EF-hand Ca2+-binding protein (20kDa)

Structurally related to Calmodulin Binds target proteins and regulate a variety of cellular processes

Phosphorylation and enzyme activity Highly expressed in cardiomyocytes (0.2% total soluble

protein content) Positive regulator of cardiac performance (Most et al., (2001)

Proc Natl Acad Sci USA 98: 13889-94. ) Regulation of Ca2+ transient

SERCA2 and RyR2

Introduction

S100A1S100A1 Expressed in endothelial and smooth

muscle cells (ECs and SMCs) ECsECs

Expression is increased in a rat model of cerebral basilar artery vasospasm (Lefranc et al., (2005) Neuropathol Appl Neurobiol 31(6): 649.)

SMCsSMCs Associates with the SR and actin stress fibers

(Mandinova et al., (1998) J Cell Sci 111: 2043.)

Objectives To determine the impact of S100A1

deficiency on systemic BP

To assess the role of the endothelium in vascular function in S100A1 KO animals

To evaluate S100A1 and eNOS interaction

Methods•S100A1 KO S100A1 KO **•WT (C57BL/6)WT (C57BL/6)

* Du et al., (2002) Mol Cell Biol 22(8):2821.

Human aortic Human aortic endothelial cellsendothelial cells

Ex vivoEx vivo vessel myograph isolated resistance and conduit vessels in KO and WT

In vivoIn vivo micromanometer catheter hemodynamics following coronary artery ligation or sham surgery impact of gender and anaesthesia was analysed

In vitroIn vitro eNOS/S100A1 co-immunoprecipitation and immunofluorescence co-localisation assays

S100A1 KO mice demonstrate poor survival and higher MAP following MI

0 10 20 300

50

100

WT

S100A1 KO

Time (days)

Surv

ival

(%)

n=49-56

P<0.01**

0

20

40

60

80

100

120

Sham MI

MAP

(mm

Hg)

WTS100A1KO

#, P<0.05 vs.respective-WT (n=6-18)

#

#M

AP

(mm

Hg)

MA

P (m

mH

g)

ShamSham MIMI0

20

40

60

80

100

120

Sham MI

MAP

(mm

Hg)

WTS100A1KO

#, P<0.05 vs.respective-WT (n=6-18)

#

#M

AP

(mm

Hg)

MA

P (m

mH

g)

ShamSham MIMI

Male S100A1 KO exhibit higher mortality and MAP than female S100A1 KO mice

0 10 20 300

50

100

Female KO

Male KO

Time (days)

Surv

ival

(%)

P<0.05

n=20-36

**

0

20

40

60

80

100

120

140

Males Females

MAP

(mm

Hg)

WTS100A1KO

* P<0.05 vs. female KO

# P<0.05 vs.respective-WT (n=6-18)

*

#

#

MalesMales FemalesFemales

MA

P (m

mH

g)M

AP

(mm

Hg)

0

20

40

60

80

100

120

140

Males Females

MAP

(mm

Hg)

WTS100A1KO

0

20

40

60

80

100

120

140

Males Females

MAP

(mm

Hg)

WTS100A1KO

* P<0.05 vs. female KO

# P<0.05 vs.respective-WT (n=6-18)

*

#

#

MalesMales FemalesFemales

MA

P (m

mH

g)M

AP

(mm

Hg)

The hypertensive phenotype is attenuated in S100A1 KO mice but persists under 2%

isoflurane anaesthesia

0102030405060708090

100

Males Females

MAP

(mm

Hg)

WTS100A1KO

* P<0.05 vs. female KO

#, P<0.05 vs.respective-WT (n=8-20)

*#

#

0102030405060708090

100

Males Females

MAP

(mm

Hg)

WTS100A1KO

* P<0.05 vs. female KO

#, P<0.05 vs.respective-WT (n=8-20)

*#

#

Male KO mice exhibit cardiac hypertrophy

3.0

4.0

5.0WTS100A1 KO

MalesMales

HW

HW

// BW

(mg

BW (m

g //g)g) *

* P<0.05

n=18-23

3.0

4.0

5.0WTS100A1 KO

3.0

4.0

5.0WTS100A1 KO

3.0

4.0

5.0WTS100A1 KO

3.0

4.0

5.0WTS100A1 KO

MalesMales

HW

HW

// BW

(mg

BW (m

g //g)g) *

* P<0.05

n=18-23

* P<0.05

n=18-23

Male S100A1 KO mice exhibit non-characteristic increase in systolic pressure following IP injection

of the 2-adrenergic agonist xylazine

0 5 10 15 20 25

-25

0

25

50WT n=5S100A1 KO n=4

Time (min)

L

VSP

(mm

Hg)

* P<0.01

*

** * *

0 5 10 15 20 25

-25

0

25

50WT n=5S100A1 KO n=4

Time (min)

L

VSP

(mm

Hg)

* P<0.01

*

** * *

0 5 10 15 20 25

-200

-150

-100

-50

0

Time (min)

HR

(bpm

)

Endothelium-dependent relaxation in isolated vessels is reduced in male

S100A1 KO

Mesenteric arteries

-8 -7 -6 -5

0

25

50

75

100

WT (n=5)S100A1 KO (n=5)

log Acetylcholine (M)

% R

elax

atio

n

**PP<0.05<0.05

* **

**

Aortas

-8 -7 -6 -5

0

25

50

75

100

WT (n=5)S100A1 KO (n=4)

log Acetylcholine (M)

% R

elax

atio

n

**PP<0.05<0.05

**

**

Mesenteric arteries

-8 -7 -6 -5

0

25

50

75

100

WT (n=5)S100A1 KO (n=5)

log Acetylcholine (M)

% R

elax

atio

n

**PP<0.05<0.05

* **

**

Aortas

-8 -7 -6 -5

0

25

50

75

100

WT (n=5)S100A1 KO (n=4)

log Acetylcholine (M)

% R

elax

atio

n

**PP<0.05<0.05

**

**

S100A1 KO and WT vessels exhibit similar vasoconstriction in response to

phenylephrine

Mesenteric arteriesMesenteric arteries AortasAortas

S100A1 and eNOS partly co-localize in cultured human aortic endothelial

cells (HAECs)

eNOSeNOS S100A1S100A1 mergedmerged

x60 x60 magmag

S100A1 and eNOS co-immunoprecipitates in cultured HAECs

IP: eNOSIP: eNOS

WB: eNOS, S100A1WB: eNOS, S100A1

eNOSeNOS

140 kDa140 kDa

S100A1S100A1

36 kDa36 kDaEGTA EGTA CCaa

2+2+

12hr12hr

Summary and conclusions• S100A1 plays a role in the regulation of basal blood

pressure in a gender specific manner

• Endothelium-dependent relaxation of isolated resistance and conduit vessels is reduced in male S100A1 KO animals

• eNOS and S100A1 co-immunoprecipitate in HAECs and partly co-localize around the nucleus

• S100A1 may be important not only in cardiac function in disease but also in the vascular responses