Extended Data Fig. 5: The effect of dextromethorphan on potassium channels in Timothy syndrome iPSC-derived cardiomyocytes.
From: Sigma non-opioid receptor 1 is a potential therapeutic target for long QT syndrome
(a) Representative confocal fluorescent images of Timothy syndrome cardiomyocytes without treatment (left, TS) and with dextromethorphan (+Dxm). Scale bar, 5μm. (b) Quantification of fraction of green (hERG) over red (WGA) fluorescence signals in TS cardiomyocytes without treatment (n = 38) and with Dxm (n = 31). (c) Quantification of KCNH2 transcript in TS cardiomyocytes with and without Dxm (n = 4/group). (d) Representative immunoblots of human hERG and GAPDH protein using lysates from non-treated and Dxm-treated TS cardiomyocytes. (e) Quantification of hERG protein expressions (normalized to GAPDH) in non-treated and Dxm-treated TS cardiomyocytes (n = 3/group). The trend towards a decreased hERG protein expression in the treated group might be driven by one sample (#) with a reduced loading amount compared with others. (f) IKs current steady-state amplitudes were significantly reduced in TS cardiomyocytes (n = 10) compared to isogenic control (n = 10) at 20 and 50 mV steps from -40mV hold. (g) Representative traces of IKs currents (Chromanol 293B-sensitive) in isogenic Ctrl cardiomyocytes (black) and TS cardiomyocytes treated with Dxm (purple), Dxm and NE-100 (green) or without treatment (red, TS). The IKs traces are shown in Fig. 2l. (h) Representative confocal fluorescent images of Timothy syndrome cardiomyocyte without treatment (left, TS) and treated with dextromethorphan (+Dxm). Scale bar, 5μm. (i) Quantification of fraction of green (KV7.1) over red (WGA) fluorescence signals in TS cardiomyocytes without treatment (n = 21) and treated with Dxm (n = 31). (j) Quantification of KCNQ1 transcripts (isoform 1 and 2) in TS cardiomyocytes without treatment and treated with Dxm (n = 4/group). (k) Representative immunoblots of human KV7.1 and GAPDH protein using lysates from non-treated and Dxm-treated TS cardiomyocytes. (l-m) Quantification of KV7.1 isoform 1 (l) and isoform 2 protein expressions (m, normalized to GAPDH) in non-treated and Dxm-treated TS cardiomyocytes (n = 3/group). All data are mean ± s.d. The treatment of Dxm or NE-100 for all experiments was 5 μM, 2hrs. Unpaired two-tailed Student’s t-test was used for b, c, e, i, j, l, m between the groups and used for f at each voltage step. * P < 0.05. n.s., not significant. The cell samples were from at least two independent differentiations.
