SummaryCardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66%–87%. Approximately 6.2–7.0 × 108 cells (4-layer) and 1.5–2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future.

To the best of our knowledge, this is the first report of heterotopic transplantation of a decellularized whole porcine heart with mesenchymal stem cells. The scaffolds endured surgical procedures. We detected short-term coronary artery perfusion in the transplanted scaffolds by angiography. Future studies should analyse the histological features of transplanted decellularized scaffolds and optimize the system for recellularization to apply this unique technology clinically.

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