SNTA1 gene rescues ion channel function and is antiarrhythmic in cardiomyocytes derived from induced pluripotent stem cells from muscular dystrophy patients
- Department of Internal Medicine and Molecular and Integrative Physiology, Center for Arrhythmia Research, University of Michigan, United States
- Leviev Heart Center, Sheba Medical Center, Tel Hashomer, and Tel Aviv University, Israel
- Centro Nacional de Investigaciones Cardiovasculares (CNIC) Carlos III, Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Spain
- Department of Molecular and Integrative Physiology, University of Michigan Medical School, United States
- Department of Physiology, Biophysics and Systems Biology, Ruth and Bruce Rappaport Faculty of Medicine, Technion - Israel Institute of Technology, Israel
Figures
Figure 1
Altered ECG and arrhythmias in Duchenne Muscular Dystrophy (DMD) patients with cardiomyopathy.
(a) Abnormal ECG in a 34-year-old DMD male: PR interval, 116 ms; QRS, 120 ms; QT/QTc, 404/472 ms; and PRT axes, 18-16-90. (b) Holter recording from the same patient shows nonsustained monomorphic …
Figure 2
Duchenne Muscular Dystrophy (DMD) patient-specific iPSC-CMs do not express dystrophin.
(a) Top right, control and heterozygous female iPSC-CMs express dystrophin. iPSC-CMs from hemizygous dystrophic cell lines (Males 1 and 2) did not express the large dystrophin isoform. Top left, …
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Figure 2—source data 1
Full unedited gel.
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig2-data1-v1.zip
Figure 3 with 2 supplements
Action potential properties in control, Duchenne Muscular Dystrophy (DMD), and female iPSC-CMs.
(a) Representative action potentials of ventricular-like iPSC-cardiomyocytes from Control 1, heterozygous female, and DMD individuals. The respective dV/dt trace is shown below each action …
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Figure 3—source data 1
Action potential parameters.
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig3-data1-v1.zip
Figure 3—figure supplement 1
Electrophysiological analysis in Control 2 (human foreskin-derived BJ iPSC-CMs).
(a) Representative AP trace of ventricular-like control BJ iPSC-CMs obtained at 1 Hz of pacing. Inset. First derivative with respect to time (dV/dt). (b–f) Action potential properties. Recordings at …
Figure 3—figure supplement 2
Action potential properties of control, hemizygous, and heterozygous Duchenne Muscular Dystrophy (DMD) iPSC-CMs paced at 2 Hz.
(a) Maximal AP upstroke velocity was reduced in both hemizygous (****p < 0.0001), and the heterozygous female (***p = 0.0002) iPSC-CMs compared to control. (b–d) Overshoot values were similar among …
Figure 4 with 2 supplements
Conduction velocity (CV) is slower in iPSC-CM monolayers from Duchenne Muscular Dystrophy (DMD) hemizygous male and heterozygous female than control.
(a) Activation maps of action potential propagation at 1 Hz. Each color represents a different activation time with time zero appearing in green (¤ indicates the location of the stimuli for each …
Figure 4—figure supplement 1
Conduction velocity in control BJ iPSC-CM (Control 2) monolayer.
(a) Activation maps of action potential propagation at 1 Hz. Each color represents a different activation time with time zero appearing in green (¤ indicates the location of the stimuli for each …
Figure 4—figure supplement 2
Cx43 expression level in control, hemizygous, and heterozygous Duchenne Muscular Dystrophy (DMD) iPSC-CMs.
(a) Typical western blot for connexin43 expression. About 50 k cells were collected to quantify total dystrophin, connexin43, and actinin levels in iPSC-CMs. (b) Scatter plots of Cx43 detected in …
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Figure 4—figure supplement 2—source data 1
Full unedited gel showing Dystrophin, Actinin and Cx43 expression levels by WB technique.
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig4-figsupp2-data1-v1.zip
Figure 5 with 4 supplements
Sodium (INa) and Inward rectifier potassium (IK1) channel properties in control, Duchenne Muscular Dystrophy (DMD), and female iPSC-cardiomyocytes.
(a) Superimposed INa current traces for Control 1, hemizygous, and heterozygous iPSC-CMs elicited by the pulse protocol shown by the inset. (b) Left, normalized current–voltage (I/V) relationships. I…
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Figure 5—source data 1
Voltage clamp data.
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig5-data1-v1.zip
Figure 5—figure supplement 1
Cell capacitance.
No statistically significant differences were observed among control (32 ± 2 pF), female (39 ± 3 pF; p = 0.0715), and Duchenne Muscular Dystrophy (DMD) iPSC-CMs (38 ± 3 pF, p = 0.3257 for Male 1; …
Figure 5—figure supplement 2
Calcium channel properties in control, Duchenne Muscular Dystrophy (DMD), and female iPSC-CMs.
(a) Original calcium current traces obtained from all iPSC-CMs elicited by depolarizing potential as shown in the inset. (b) Current–voltage relationships showing no significant differences among …
Figure 5—figure supplement 3
SCN5A, KCNJ2, and CACNA1C mRNA expression in control, Duchenne Muscular Dystrophy (DMD), and female iPSC-CMs.
(a) SCN5A mRNA expression was increased in iPSC-CMs from hemizygous and heterozygous DMD individuals (top), as well as in the human left ventricle heart tissue from a Becker MD individual compared …
Figure 5—figure supplement 4
NaV1.5 protein level is significantly reduced in patient-specific Duchenne Muscular Dystrophy (DMD) iPSC-CMs.
(a) Representative western blot for each antibody used. The bands within the blue rectangles at ~250 KDa and below 50 KDa correspond to NaV1.5 and Kir2.1, respectively. About 50 k cells were …
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Figure 5—figure supplement 4—source data 1
Full unedited gel.
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig5-figsupp4-data1-v1.zip
Figure 6
Transfection of SNTA1 rescues membrane levels of Kir2.1 and NaV1.5 proteins in iPSC-CMs from Male 1 patient.
(a) Cartoon illustrating nonviral piggy-bac vector encoding SNTA1 for transfection in Male 1 iPSC-CMs. SNTA1 coding region (CDS) is driven by the CAG promoter and followed by green fluorescence …
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Figure 6—source data 1
SNTA1 quantification (Figure 6d).
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig6-data1-v1.zip
Figure 7
SNTA1 expression restores the electrophysiological deficiencies in Duchenne Muscular Dystrophy (DMD) iPSC-CMs.
(a, b) Normalized current–voltage (I/V) relationships for INa and IK1 in Male 1 before (black) and after (red) syntrophin expression at the specified voltages. Two-way analysis of variance (ANOVA) …
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Figure 7—source data 1
SNTA1 IV IK1 and IV INA (Figure 7A and B); and, Action potential properties (Figure 7C).
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig7-data1-v1.zip
Figure 8 with 1 supplement
SNTA1 expression prevents reentrant arrhythmias in Duchenne Muscular Dystrophy (DMD) iPSC-CM monolayers.
(a) Optical maps of action potential propagation during pacing at 1 Hz. Each color represents a different activation time with time zero appearing in blue (star below each monolayer indicates the …
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Figure 8—source data 1
Conduction Velocity and APD90 in the absence and in the presence of SNTA1 expression.
- https://cdn.elifesciences.org/articles/76576/elife-76576-fig8-data1-v1.zip
Figure 8—figure supplement 1
SNTA1 expression rescues contractility in Duchenne Muscular Dystrophy (DMD) iPSC-CM monolayers.
Contractility measured by cell shortening in spontaneously beating iPSC-CM monolayers from DMD hemizygous Male 1 is reduced with respect to control and rescued fully by SNTA1 expression. (a) …
Videos
Video 1
Duchenne Muscular Dystrophy (DMD) male arrhythmia patterns.
Movie # i Control iPSC-CM monolayer showing slow and organized spontaneous activity. Movies # ii–v. DMD hemizygous Male 1 iPSC-CM monolayers are inducible of high-frequency reentry maintained by one …
Additional files
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Supplementary file 1
Action potential parameters of iPSC-CMs paced at 1 and 2 Hz.
- https://cdn.elifesciences.org/articles/76576/elife-76576-supp1-v1.docx
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Supplementary file 2
Action potential parameters of iPSC-CMs paced at 1 and 2 Hz.
- https://cdn.elifesciences.org/articles/76576/elife-76576-supp2-v1.docx
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Supplementary file 3
Action potential parameters of iPSC-CMs at 1 and 2 Hz, Control 1 versus Control 2.
- https://cdn.elifesciences.org/articles/76576/elife-76576-supp3-v1.docx
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Supplementary file 4
Biophysical parameters of DMD, and female iPSC-CMs versus Control 1.
- https://cdn.elifesciences.org/articles/76576/elife-76576-supp4-v1.docx
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Supplementary file 5
Biophysical parameters of DMD, and female iPSC-CMs versus Control 2.
- https://cdn.elifesciences.org/articles/76576/elife-76576-supp5-v1.docx
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Supplementary file 6
Biophysical parameters of iPSC-CMs, Control 1 versus Control 2.
- https://cdn.elifesciences.org/articles/76576/elife-76576-supp6-v1.docx
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Supplementary file 7
Primers used in mRNA analysis.
- https://cdn.elifesciences.org/articles/76576/elife-76576-supp7-v1.docx
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Transparent reporting form
- https://cdn.elifesciences.org/articles/76576/elife-76576-transrepform1-v1.pdf
