Differentiation of Human Pluripotent Stem Cells Into Definitive Endoderm Cells in Various Flexible Three-Dimensional Cell Culture Systems: Possibilities and Limitations

The study investigated the differentiation of human pluripotent stem cells into definitive endoderm (DE) cells using various 3D cell culture systems, including suspension culture and nanofibrillar cellulose (NFC) hydrogel. It found that suspension culture was superior due to better mass transfer, leading to more effective DE differentiation. Spheroid size was crucial, with larger spheroids (1,000 cells) being more effective for DE differentiation, while smaller spheroids had low survival rates. The ROCK inhibitor Y-27632 improved spheroid survival and viability during differentiation. The study highlighted the importance of biomaterial properties, spheroid size, and culture conditions in optimizing stem cell differentiation, with implications for drug research, developmental biology, and regenerative medicine.