Fig. 4: Isolated murine aCMs can be genetically manipulated in vitro to functional effect.

a Myocytes isolated from a PLN-null mouse were transfected using lentiviral vectors expressing wildtype (WT) and human pathogenic R9C phospholamban (PLN) flag tagged cDNAs. Immunoblots to anti-flag and α-tubulin. b Confocal images of aCMs stained for FLAG expression post lentiviral transfection for PLN. Scale bars represent 20 µm. c Traction force microscopy of aCMs post transfection. d Myocytes isolated from a WT mouse were transfected using AAV9 vectors encoding Reep5 shRNA sequence and assessed by immunoblots against REEP5 and tubulin. Reep5 KD treatment were loaded at higher volume to produce quantifiable bands. e Confocal images of aCMs confirm shRNA knockdown of Reep5 and resulting disorganization of SR (RyR2) and ER (KDEL motif) compared to scrambled shRNA control; scale bars represent 20 µm. f Peak Ca²⁺ transient amplitude is not significantly affected by Reep5 KD (N = 6). g Time to peak Ca²⁺ transient amplitude is significantly higher after Reep5 KD, but transient decay time to 50% is not affected. All data expressed as mean ± SEM; N denotes biological replicates; significance indicated by *(p < 0.05), **(p < 0.01), and ***(p < 0.001).