SLM2-dependent effects in human iPSC-CMs
A. Diagram showing the generation of iPSC-CMs. Fibroblasts of human healthy donors were reprogrammed using non-integrating Sendai virus to generate iPSC clones. iPSC-CMs were then generated by Wnt modulation and metabolic selection. B. IB showing AAV6-mediated overexpression of SLM2 in iPSC-CMs. β-actin was used as a loading control. Mock indicates iPSC-CMs transfected with control AAV6 virus. C. Gene expression after overexpression of SLM2 by transduction with AAV6 virus (n = 4) analyzed by NGS. D. Immunofluorescence assay of the sarcomeric protein titin Z1/Z2. Scale bar, 50 µm. E. and F. Determination of sarcomere regularity using FFT. Four cardiac differentiation experiments for one cell line were used, and single iPSC-CMs were analyzed (Mock: n = 74 iPSC-CMs; SLM2: n = 85 iPSC-CMs). G. Change of Ca2+ transient rise time. H. Change of Ca2+ half decay time. I. and J. [Ca2+]i (F/F0) after overexpression of SLM2 in iPSC-CMs (Mock, n = 66; SLM2, n = 66). Data are presented as mean ± SEM. Statistical differences were calculated using Student’s t-test (*, P < 0.05). iPSC, induced pluripotent stem cell; iPSC-CM, iPSC-derived cardiomyocyte; AAV6, adeno-associated virus serotype 6; NGS, next-generation sequencing; FFT, Fast Fourier Transform.