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. 2018 Sep 11;11(3):626-634.
doi: 10.1016/j.stemcr.2018.07.012. Epub 2018 Aug 23.

Channelopathy as a SUDEP Biomarker in Dravet Syndrome Patient-Derived Cardiac Myocytes

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Channelopathy as a SUDEP Biomarker in Dravet Syndrome Patient-Derived Cardiac Myocytes

Chad R Frasier et al. Stem Cell Reports. .

Abstract

Dravet syndrome (DS) is a severe developmental and epileptic encephalopathy with a high incidence of sudden unexpected death in epilepsy (SUDEP). Most DS patients carry de novo variants in SCN1A, resulting in Nav1.1 haploinsufficiency. Because SCN1A is expressed in heart and in brain, we proposed that cardiac arrhythmia contributes to SUDEP in DS. We generated DS patient and control induced pluripotent stem cell-derived cardiac myocytes (iPSC-CMs). We observed increased sodium current (INa) and spontaneous contraction rates in DS patient iPSC-CMs versus controls. For the subject with the largest increase in INa, cardiac abnormalities were revealed upon clinical evaluation. Generation of a CRISPR gene-edited heterozygous SCN1A deletion in control iPSCs increased INa density in iPSC-CMs similar to that seen in patient cells. Thus, the high risk of SUDEP in DS may result from a predisposition to cardiac arrhythmias in addition to seizures, reflecting expression of SCN1A in heart and brain.

Keywords: SUDEP; cardiac arrhythmia; developmental and epileptic encephalopathy; epilepsy; induced pluripotent stem cell (iPSC); sodium channel.

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Figures

Figure 1
Figure 1
Immunofluorescence Staining and Spontaneous Contraction Rates of iPSC-CMs (A) Anti-α-actinin (red) and anti-connexin 43 (green) staining for DS5. (B) Anti-α-actinin (red) and anti-Nav1.5 (green) staining for DS5. (C) Anti-α-actinin (red) and anti-NKX 2.5 (green) staining for DS5. (D) Increased contraction rates in DS iPSC-CM lines compared with control. Data from two independent control lines were pooled. For DS4, DS10, and DS5, data from two independent clones were pooled. For all lines, data were collected from 4–6 wells of one differentiation. p < 0.05. (E) Representative MEA traces from control (left) and DS (right) iPSC-CM lines. (F) Quantification of contraction rates in MEA recordings. For control 3 (Con 3) and DS4, data were pooled from two different clones, with two independent differentiations per clone. p < 0.05. Immunofluorescence staining of DS2, DS4, and DS10 is shown in Figure S1. Data in (E) and (F) are presented as means ± SEM. Scale bar, 20 μm (A–C). See also Figure S2.
Figure 2
Figure 2
iPSC-CMs from DS Patients Have Increased Whole-Cell INa and Altered SCN5A Expression (A) Representative whole-cell INa traces from a control and three DS subjects. (B) Representative images of control 2 (Con 2) and DS5 iPSC-CMs used in electrophysiological recordings. (C) INa current-voltage relationships for control and three DS iPSC-CM lines. (D) Peak INa is increased in all four DS iPSC-CM lines compared with controls. p < 0.05 versus control; #p < 0.05 versus average of DS2, DS4, and DS10. (E) Current-voltage relationship comparing DS5 to control. Inset: representative INa trace from DS5. Peak INa data were pooled from two clones each for DS4, DS10, and DS5, respectively. Peak INa data for the control group were pooled from two independent control iPSC-CM lines (Con 2, 1 clone; Con 3, two clones). Data comparing the two control lines are shown in Figure S3. At least two independent differentiations were analyzed per line (Con 2, 3; Con 3-1, 2; Con 3-3, 2; DS2-3, 3; DS4-3, 3; DS4-4, 2; DS10-2, 4; DS10-7, 2; DS5-1, 2; DS5-2, 2). (F) qRT-PCR data from male and female control iPSC-CM lines, pooled DS patient iPSC-CMs, and adult human heart control tissue. Data in (C), (D), and (E) are presented as means ± SEM for all groups. For (F), data for DS samples are presented as means ± SEM (n = 4). See also Figures S3 and S4; Table S1.
Figure 3
Figure 3
Abnormal Electrocardiogram and Limited Heart-Rate Variability in DS Subject 5, Arrhythmogenic Substrates in DS5 iPSC-CMs, and Increased Sodium Current Density in a CRISPR-Generated Heterozygous SCN1A Deletion Line (A) Electrocardiogram recordings for DS subject 5 demonstrate normal sinus rhythm, T-wave inversions, and lateral T-wave flattening, abnormal for age. (B) Representative power-spectrum analysis for heart-rate variability from 24-hr ambulatory Holter monitoring demonstrates limited short-term variability as compared with normal values for pediatric patients. Inset: compiled data from annual Holters, n = 3. (C) Representative AP traces showing examples of delayed afterdepolarizations (DADs) in DS5 iPSC-CMs. (D) Quantification of the incidence of early afterdepolarizations (EADs) and DADs in control and DS5 iPSC-CM lines. AP data for Con 2 and DS5 are from two independent differentiations each. p < 0.05. (E) Representative whole-cell INa traces from SCN1A heterozygous deletion (SCN1A+/) and isogenic control (SCN1A+/+) iPSC-CMs. (F) INa current-voltage relationships for SCN1A+/ and SCN1A+/+ isogenic control iPSC-CM cells. (G) Peak INa is increased in SCN1A+/ iPSC-CMs compared with the SCN1A+/+ isogenic control line. Peak INa data for SCN1A+/+ and SCN1A+/ are from two independent differentiations each. p < 0.05 versus control. Data in (F) and (G) are presented as means ± SEM. See also Figure S4; Tables S2 and S3.

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References

    1. Ackerman M.J., Splawski I., Makielski J.C., Tester D.J., Will M.L., Timothy K.W., Keating M.T., Jones G., Chadha M., Burrow C.R. Spectrum and prevalence of cardiac sodium channel variants among black, white, Asian, and Hispanic individuals: implications for arrhythmogenic susceptibility and Brugada/long QT syndrome genetic testing. Heart Rhythm. 2004;1:600–607. - PubMed
    1. Auerbach D.S., Jones J., Clawson B.C., Offord J., Lenk G.M., Ogiwara I., Yamakawa K., Meisler M.H., Parent J.M., Isom L.L. Altered cardiac electrophysiology and SUDEP in a model of Dravet syndrome. PLoS One. 2013;8:e77843. - PMC - PubMed
    1. Bao Y., Isom L.L. NaV1.5 and regulatory β subunits in cardiac sodium channelopathies. Card. Electrophysiol. Clin. 2014;6:679–694.
    1. Cheng J., Tester D.J., Tan B.H., Valdivia C.R., Kroboth S., Ye B., January C.T., Ackerman M.J., Makielski J.C. The common African American polymorphism SCN5A-S1103Y interacts with mutation SCN5A-R680H to increase late Na current. Physiol. Genomics. 2011;43:461–466. - PMC - PubMed
    1. Claes L., Del-Favero J., Ceulemans B., Lagae L., Van Broeckhoven C., De Jonghe P. De novo mutations in the sodium-channel gene SCN1A cause severe myoclonic epilepsy of infancy. Am. J. Hum. Genet. 2001;68:1327–1332. - PMC - PubMed

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