Improved Vascular Remodeling and Endothelial Function in Transglutaminase 2 Knock-Out Mice Infused with Angiotensin II L.Sada 1 ,C.Savoia 1 ,M.Briani 1 ,E Arrabito 1 ,S.Michelini 1 ,L.Pucci 1 , T.Bucci 1 ,C.Nicoletti 2 ,E Candi 3 , EL Schiffrin, M Volpe 1 . 1 Clinical and Molecular Medicine Department, Sapienza University of Rome, Italy 2 DAHFMO-Unit of Histology and Med. Embr., Sapienza University of Rome, Italy 3 Dep. of Exp. Medicine and Surgery, Fac. of Med. Unicers. of Rome Tor Vergata, Itay
Transcript
Improved Vascular Remodeling and Endothelial Function in
Transglutaminase 2 Knock-Out Mice Infused with Angiotensin II
L.Sada1,C.Savoia1,M.Briani1,E Arrabito1,S.Michelini1,L.Pucci1, T.Bucci1,C.Nicoletti2,E Candi3, EL Schiffrin, M Volpe1.
1 Clinical and Molecular Medicine Department, Sapienza University of Rome, Italy
2DAHFMO-Unit of Histology and Med. Embr., Sapienza University of Rome, Italy
3Dep. of Exp. Medicine and Surgery, Fac. of Med. Unicers. of Rome Tor Vergata, Itay
Transglutaminases (TGs) in the Vascular system
SMC
EC
• Type 1 (keratinocyte TG)• Factor XIII (plasma TG) • Type 2 (Tissue type TG)
Bakker et al., J Vasc Res, 2008
Epidermal differentiationWound healing
TGs catalyze covalent cross-linking between reactive lysine and glutamine residues of protein polymers
• Activation of PLC• Regulation of cell cycle progression
GTPaseTGase
Background
• TGs are involved in flow-induced vascular remodeling in ratcremaster arteries. Bakker et al., Circ res, 2005
• TGs are involved in aldosterone-induced vascular remodeling in mesenteric arteries and in aorta.
Yamada et al. Cardiovascular research,2008
• Tissue Transglutaminase is involved in endothelin 1-induced hypertrophy in cultured neonatal rat cardiomyocytes. Li et al. Hypertension, 2009
• We previously demonstrated that angiotensin II (Ang II) may positively regulate TG2 expression in vascular smooth muscle cells from SHR.
AIM
To determine whether Ang II may induce vascular remodeling in part
through TG2
• TG2 Knock-out mice (TG2-K/O, 8 weeks old) and age matched wild type (WT) mice were treated or not with Ang II (400 ng/Kg/min) for 14 days.
• Blood pressure (BP) was measured by tail-cuff method.
• Functional, structural and mechanical studies were performed on segments of pressurized (45 mmHg) mesenteric arteries.
• Vascular reactive oxygen species (ROS) level in the aorta was avaluated by dihydroethidium (DHE) staining.
• The expression of eNOS in aorta was evaluated by immunoblotting.
Methods
• BP was higher in TG2-K/O mice compared to WT (120.3±1.3 mmHg vs 88.3±1.9 mmHg, P<0.05).Ang II infusion significantly increased BP only in WT (+28% vs untreated WT, P<0.05), whereas BP was unchanged in TG2-K/O after Ang II infusion.
• TG2-K/O presented reduced M/L as compared to WT (4.8±0.3% vs 6.5±0.2%, P<0.05). Ang II infusion increased M/L only in WT (+13% vs untreated WT, P<0.05). M/L resulted unchanged in TG2-K/O after Ang II infusion. CSA was similar in all groups.
Results
• Endothelium-dependent relaxation was similarly preserved in untreated WT, TG2-K/O and Ang II-treated TG2-K/O. Ang II infusion impaired acetylcholine-induced relaxation only in WT (-50% vs untreated WT, P<0.05). L-NAME blunted acetylcholine-induced relaxation in all groups except in Ang II-treated WT
• SNP-dependent relaxation was similar in all groups.
Results
• eNOS expression was similar in untreated WT and untreated TG2-K/O. eNOS significantly increased only in TG2-K/O treated with Ang II
• ROS production was similar in untreated WT and untreated TG2-K/O. Ang II significantly increased ROS in WT (2-fold increase), and significantly decreased ROS in TG2-K/O
Results
WT TG2-K/O WT+AngII
TG2-K/O+Ang II
0
50
100
150*
*
mm
Hg
Blood Pressure in WT and TG2-K/O mice treated or not with angiotensin II
WT TG2-K/O
WT+AII
TG2-KO+AII
0.0
2.5
5.0
7.5
10.0 *
*
M/L
rati
o (
%)
0
5000
10000
15000
CS
A (
μm
2)
Media-to-Lumen Ratio and Cross-sectional area
of mesenteric arteries from WT and TG2-K/O mice
WT TG2-K/O
WT+AII
TG2-KO+AII
-9 -8 -7 -6 -5 -40
50
100
WT
TG2-KO
WT+Ang II
TG2-KO+Ang II
*
Acetylcholine (log M)
% o
f re
laxa
tion
-8 -7 -6 -5 -4 -30
50
100
WT
TG2-KO
WT+Ang II
TG2-KO+Ang II
SNP (log M)
% o
f re
laxati
on
Endothelium-dependent and -independent relaxation in mesenteric arteries from WT and
TG2-K/O mice
% o
f re
laxati
on
-9 -8 -7 -6 -5 -40
25
50
75
100 WT_(Acetylcholine)
WT (Acetylcholine+LNAME)
Acetylcholine (log M)
*
-9 -8 -7 -6 -5 -4
0
25
50
75
100 TG-2K/O (Acetylcholine)
TG-2K/O (Acetylcholine+LNAME)
*
Acetylcholine (log M)
% o
f re
laxa
tion
-9 -8 -7 -6 -5 -40
25
50
75
100 WT+Angi II (Acetylcholine)
WT+Angi II (Acetylcholine+LNAME)
Acetylcholine (log M)
% o
f re
laxati
on
Dose response curves to Acetilcholine ± LNAME in mesenteric arteries from WT and TG2-K/O mice
treated or not with angiotensin II
-9 -8 -7 -6 -5 -40
25
50
75
100TG2KO+Angi II (Acetyocholine+LNAME)
TG2-KO+Ang II ( Acetylcholine)
*
Acetylcholine (log M)
% o
f re
laxati
on
WT TG2 -K/O WT+AII
TG2-K/O+AII
0
100
200
eNOS expression in aorta from WT and TG2-K/O mice
beta actin
eNOS
eN
OS
/β-a
cti
n (
% C
TR
L) *
WT
WT + Ang II
TG2-K/O + Ang II
TG2 K/O
WT TG2-K/O WT+AII
TG2-K/O+AII
0
100
200
300
400
ROS production in aorta from WT and TG2-K/O mice
Arb
itra
ry U
nit
s
*
*
Conclusion and perspectives
• Despite the higher BP values, TG2-K/O presented improved vascular remodeling compared to WT.
• In TG2-K/O, Ang II failed to increase ROS production and M/L; moreover it failed to impair endothelial function in this group.
• Hence, TG2 may play a role in Ang II-induced vascular structural and functional alterations.
