Figure 3

iPSC can differentiate into functional CMs. (a) Reverse transcription-polymerase chain reaction (RT-PCR) for the expression of cardiac-specific genes in control- (WT) and CPVT-iPSC-derived beating explants (beating EBs); undifferentiated iPSCs and FHs were used as negative and positive controls, respectively. HGPRT: housekeeping gene; CACNA1C: calcium channel, voltage-dependent, α1A subunit; SCN5A: sodium channel, voltage-gated, type V, alpha subunit; KCNQ1: potassium, voltage-gated channel, KQT-like subfamily, member 1; MHCα: myosin heavy-chain α; MHC β: myosin heavy chain β. (b) Western blot of WT- and CPVT-IPSC-derived beating explants for RyR2. β-Actin was used as the loading control, and human FH was used as a positive control. (c) RyR2 expression quantification in WT- and CPVT- beating explants, calculated as densitometry RyR2/β-actin ratio (the diagram represents the mean of four independent experiments). (d) Immunostaining of CPVT-iPSC-derived CMs for α-actinin and RyR2. Nuclei stained in DAPI. Scale bar=20 μm. (e) Representative traces of spontaneous APs recorded in iPSC harvested from the healthy donor (WT-iPSC-CMs, left) and the patient carrying the CPVT mutation (CPVT-iPSC-CMs, right). A DAD is indicated by the arrow. (f) The main AP features measured in both iPSC-derived lines: overshoot, amplitude, MDP, maximal upstroke velocity, maximal repolarization velocity and AP duration at 30%, 50% or 90% of repolarization (respectively APD₃₀, APD₅₀ and APD₉₀). WT-iPSC-CMs, n=26; CPVT-iPSC-CMs, n=35. Values are mean±MSE