Molecular Mechanisms of Contractile Dysfunction in Atrial Fibrillation
Derailment of proteostasis underlies structural and contractile dysfunction in AF
Bianca Brundel, PhD
Department of Physiology VUmc, and Clinical Pharmacy and Pharmacology UMCG
Atrial Fibrillation (AF)
• The most common cardiac arrhythmia
• Impact on patient • QoL • Mortality/morbidity • Stroke • Heart failure
• Causes of AF • Congenital heart disease (up to 30%) • Hypertension • Diabetes/Obesity
AF progression
Normal Paroxysmal Permanent
Ion-channel blockers
• Electrical remodeling: shortening APD • Changes in calcium handling Role of Calcium Handling in AF progression
Voigt, Europace 2012 AF progression
Normal Paroxysmal Permanent
Ion-channel blockers
Normal Permanent AF
Brundel, CVR 2002 Sustainable Structural damage (myolysis) Structural changes lead to contractile dysfunction and AF progression Brundel CVR 2002, De Groot and Allessie Circ 2010 Structural damage is rooted in derailment of proteostasis
Proteostasis: is homeostasis in protein expression, folding and function
Changes in gene expression
HDAC6 tubulin
Hartl, Nature 2011, Balch Science 2009 Tachypaced HL-1 mouse atrial cardiomyocytes
Control TP
Control 4 hrs TP
CaT
CS
Brundel CVR 2004, Circ Res 2006, JMCC 2006 Tachypaced Drosophila pupae
Zhang D, JMCC 2011 Den Hoed M, Nature Genetics, 2013 Zhang D, Circulation 2014 Tachypaced Drosophila pupae
Before
Before
During During
After
After
• overexpression/siRNA • Drug screen Role of Heat Shock Proteins in derailment of proteostasis and AF progression
HSP27 binds and stabilizes sarcomers HSP27 expression is induced in paroxysmal AF and gets exhausted in persistent AF
Hsp27
SR PAF
CAF
% structural damage
Brundel, JMCC 2006 HSP27 represents a druggable target in AF
• HSP overexpression
Control TP
CaT
CS
HSP27
140
120 NP 100 pSP64 pHSP27-wt 80 *** *** pHSP27-uP-A 60 *** pHSP27-P-D pHSP70
% CaT amplitude CaT % 40
20 0 2 3 4 Duration pacing (hrs) TP TP HSP27
12 Brundel Circ Res 2006, JMCC 2006, 2008, PLosOne 2011 HSP27 represents a druggable target in AF
• HSP27 was overexpressed only in the heart Control TP
• HSP27 levels are reduced in persistent AF patients
HSP27 • Overexpression of HSP27 protects against contractile dysfunction
• HSP27 represents a druggable target?
TP TP HSP27 Prevent derailment of proteostasis by HDAC inhibition
Deli Zhang
Histone deacetylases (HDAC) HDACs involved in pathological cardiac remodeling
deacetylate histones
(McKinsey TA, 2007,2012; Gupta et al. 2008; Liu F et al. 2008) HDACs involved in pathological cardiac remodeling
Bind MEF2
Nuclear export
(McKinsey TA, 2007,2012; Gupta et al. 2008; Liu F et al. 2008) HDACs involved in pathological cardiac remodeling
Cytosolic
Contractile proteins
(McKinsey TA, 2007,2012; Gupta et al. 2008; Liu F et al. 2008) HDACs involved in pathological cardiac remodeling
Metabolic homeostasis
(McKinsey TA, 2007,2012; Gupta et al. 2008; Liu F et al. 2008) HDACs involved in pathological cardiac remodeling
Cardiac stress
HDACs↑ HDACi HDACI/IIa HDACIIb
Histones/nuclear export Structural & contractile proteins
Pathological gene expression Contractile dysfunction
Cardiac remodeling
(McKinsey TA, 2007,2012; Gupta et al. 2008; Liu F et al. 2008) Tubacin and Nicotinamide protect against tachypacing-induced remodeling in HL-1 atrial cardiomyocytes
HDAC inhibitors HDACs Trichostatin A (TSA) I (1,2,3,8), IIa (4,5,7,9), IIb (6,10)& IV (11)
Sodium Butyrate (SoBu) I (1,2,3,8), IIa (4,5,7,9) Nicotinamide (Nic) III (Sirtuins1-7) Tubacin HDAC6 (IIb) HL-1 atrial cardiomyocytes
NP (normal pacing)
2.5 2.0 1.5
CaT (F1/F0) 1.0
TP (tachypacing ) 2.5 # # 2.0 1.5 *
CaT (F1/F0) 1.0 Control TSA SoBu Nic Tubacin
Zhang D. Circulation 2014 Tubacin and Nicotinamide protect against tachypacing-induced remodeling in Drosophila
After TP
# #
***
Zhang D. Circulation 2014 Overview effects of HDAC inhibitors in tachypaced HL-1 cardiomyocytes and Drosophila model
Zhang D. Circulation 2014 Inhibition of Tubulin DAC (TDAC) catalytic domain of HDAC6 rescues tachypacing-induced CaT reduction
HDAC TDAC A HDAC6 wt
HDAC6 m1 DN HDAC6 m2 DN HA tag HDAC6 m1-2 DN DN GADPH HDAC6 deacetylates tubulin and promotes microtubule depolymerization
HDAC6
(Matsuyama, Shimazu et al. 2002)
. Microtubules are important regulators of calcium signaling in the heart (F.C. Howarth, et al, 1999; B.G.Kerfant, 2001).
. Taxol, a microtubule stabilizer, prevents atrial fibrillation in rabbit (Xiao, Zhang et al, 2010).
Is the protective effect of HDAC6 inhibition in AF via modulation of microtubules? Tubacin protects against tachypacing-induced depolymerization of microtubules
NP AF (4Hz)
Tubulin Ac-Tub merge Tubulin Ac-tub merge
Con
Tubacin
Zhang D. Circulation 2014 Patients with AF reveal reduced amount of (acetyl) tubulin, especially in the LAA
SR PAF PeAF
R L R L R L R L R L R L R L R L R L R L Tub A-Tub GADPH
P<0.001 P<0.001 P=0.03 P=0.02
Tub Ac-tub
Zhang D. Circulation 2014 HDAC6 is activated in AF
SR PeAF HDAC6 GADPH 1.2 150 * * 1.0
• HDAC6 is activated in AF patients0.8 100 0.6 50 0.4
• HDAC6 causes contractileHDAC6/GADPH dysfuntion 0.2 HDAC6 activity ( a. u.) ( a. activity HDAC6 0 0.0 • Is HDAC6SR aAF druggable target in AF?SR PeAF
Zhang D. Circulation 2014 Prevent derailment of proteostasis by inhbition of ER stress- induced autophagy
Marit Wiersma Autophagy
‣ Autophagy in the heart • Cell survival • Cell death • Impaired cardiomyocyte function
‣ Basal activity in every cell: • Cellular quality control • Maintenance of cellular energetic balance
Tachypacing induces autophagy in cardiomyocytes Activation of autophagy: which pathway?
mTOR
mTORC1 mTORC2
mTOR-P2448S mTOR-P2481S
S6RP-P235-236S Akt-P473S
ER stress
Autophagy ER stress signaling activates autophagy in AF
IRE1 HSPA5 HSPA5
ATF6 HSPA5 HSPA5
P eIF2α PERK HSPA5 HSPA5
ER stress sensors
ATF4 ATG12 HSPA5
autophagy Inhibition of ER stress protects against contractile dysfunction in vitro
ER chaperone autophagy inhibitors ER stress inhibitor overexpression Inhibition of ER stress protects against contractile dysfunction in Drosophila
NP TP Control Control PepA BAF 4PBA
autophagy inhibitors ER stress inhibitor
NP TP 120
100 ### ### 80 *** 60
40
Heart rate (% basal) (% of rate Heart 20
0 C C PepA BAF 4PBA Is autophagy present in human AF? Patients with persistent AF reveal autophagy and ER stress
• Activation of autophagy by upstream ER stress in patients and experimental models
• ER stress as a druggable target in AF?
Conclusions
• Tachypaced HL1 cardiomyocytes and Drosophila are useful to • uncover mechanism underlying AF progression: key role derailment of proteostasis • Identify novel druggable targets, including • HSP27 • HDAC6 • ER stress
• Findings were confirmed in patients with paroxysmal and persistent AF
• Proof-Of-Concept for druggable targets: in dog model for AF
Next presentation: Molecular Intervention in Atrial Fibrillation Department of Physiology, VUmc Amsterdam and Pharmacy and Clinical Pharmacology, UMCG Groningen
Femke Hoogstra-Berends, Marit Wiersma, 2016 Denise van Marion, Xu Hu, Deli Zhang, Robert Henning, Bianca Brundel
MHI, Canada: Stanley Nattel and XiaoYan Qi
2013T096, 2013T144, 2013T088 고맙습니다 질문?