Bulk RNA-sequencing from SVBP or TTL gene knock-out and WT engineered heart tissues

For this study, we used commercially available hiPSCs (Allen Institute, AICS-0031) and created knock-out lines for small vasohibin binding protein (SVBP) and tubulin tyrosine ligase (TTL), respectively. These hiPSCs were then differentiated into cardiomyocytes using a monolayer protocol and then further used for generation of engineered heart tissues (EHTs) to more closely model human physiology. EHTs were either cultured on control silicone posts or stiffer silicone posts to achieve enhanced afterload. Overall design: We casted EHTs composed of 1 Mio cells from freshly differentiated hiPSC-cardiomyocytes (cTnT+ cells >85%) in a fibrin-based matrix and cultured them in low-glucose DMEM. WT EHTs were exposed to control silicone posts (0.28 mN/mm) or stiffer silicone posts for afterload enhancement (0.8 mN/mm). SVBP-KO and TTL-KO EHTs were only cultured on stiff silicone posts. After 60 d, EHTs were harvested, snap-frozen and RNA extracted for bulk RNAseq. n=3 EHTs per group were included, however, for SVBP-KO no successful library preparation was achieved, reducing the n number in this group.