Fig. 5: Genetic inhibition of SIRT1 in isogenic control iPSC-CMs activates ROS-CAMKII-RYR2 pathway and recapitulates pathogenic phenotypes of A388fs iPSC-CMs. | Nature Communications

Fig. 5: Genetic inhibition of SIRT1 in isogenic control iPSC-CMs activates ROS-CAMKII-RYR2 pathway and recapitulates pathogenic phenotypes of A388fs iPSC-CMs.

From: Lamin A/C deficiency-mediated ROS elevation contributes to pathogenic phenotypes of dilated cardiomyopathy in iPSC model

Fig. 5

a, b Western blot analysis of SIRT1, total CaMKII, ox-CaMKII, p-CaMKII, total RYR2, p-RYR2, and SUN1 expression in II-1-corr iPSC-CMs treated with scrambled siRNA (NC), SIRT1 siRNA (KD), or SIRT1 siRNA and SRT (KD + SRT), respectively. n = 4 independently biological repeats. c Cellular ROS levels in II−1-corr iPSC-CMs treated with scrambled siRNA, SIRT1 siRNA, or SIRT1 siRNA and SRT, respectively. n = 3 independently biological repeats. d Representative Ca2+ transient traces recorded from II−1-corr iPSC-CMs treated with scrambled siRNA, SIRT1 siRNA, or SIRT1 siRNA and SRT, respectively. e, f Bar graphs to compare the Ca2+ transient amplitude and diastolic [Ca2+]i among different groups in d. n = 19 (KD), 21 (NC, KD + SRT) cells. g Representative traces of cytosolic Ca2+ fluorescence in II−1-corr iPSC-CMs treated with scrambled siRNA, SIRT1 siRNA, or SIRT1 siRNA and SRT, respectively in NT solution and exposed to 0 Na+, 0 Ca2+ solution containing Tet and Caff. h, i Bar graphs to compare the RYR2-mediated SR Ca2+ leak and SR Ca2+ load among different groups in g. n = 24 (KD), 26 (KD + SRT), 30 (NC) cells. j Representative electrophysiological measurements of spontaneous action potentials in II-1-corr iPSC-CMs treated with scrambled siRNA, SIRT1 siRNA, or SIRT1 siRNA and SRT, respectively. k Bar graph to compare the occurrence of arrhythmias among different groups in j. l Representative graphs of staining of lamin A/C (green) and cardiac-specific marker TNNT2 (red) in II−1-corr iPSC-CMs treated with scrambled siRNA, SIRT1 siRNA, or SIRT1 siRNA and SRT, respectively. DAPI indicates nuclear staining (blue). Scale bar, 20 μm. m Bar graph to compare the occurrence of abnormal NE structure among different groups in l. n = 6 views from 3 independent experiments. Data are presented as mean ± SEM in b, c, e, f, h, i, m. p values were calculated by Dunnett’s multiple comparisons test in b, c, e, f, h, i, m. Source data are provided as a Source Data file.

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