Pluripotent stem cells, such as embryonic stem cells and induced pluripotent stem (iPS) cells, are regarded as new sources for cell replacement therapy. These cells can unlimitedly expand under undifferentiated conditions and be differentiated into multiple cell types. Automated culture systems enable the large-scale production of cells. In addition to reducing the time and effort of researchers, an automated culture system improves the reproducibility of cell cultures. In the present study, we newly designed a fully automated cell culture system for human iPS maintenance. Using an automated culture system, hiPS cells maintained their undifferentiated state for 60 days. Automatically prepared hiPS cells had a potency of differentiation into three germ layer cells including dopaminergic neurons and pancreatic cells.

The Treg cells and islet cells were distributed randomly in the coaggregates. After intraportal transplantation of the coaggregates, Treg cells in the aggregates enabled the long-term survival of allogeneic islet cell grafts in the liver without the use of immunosuppressive drugs.

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Human pluripotent stem cells (hPSCs) are thought to be a promising cell-source solution for regenerative medicine due to their indefinite proliferative potential and ability to differentiate to functional somatic cells. However, issues remain with regard to achieving reproducible differentiation of cells with the required functionality for realizing human transplantation therapies and with regard to reducing the potential for bacterial or fungal contamination. To meet these needs, we have developed a closed-channel culture device and corresponding control system. Uniformly-sized spheroidal hPSCs aggregates were formed inside wells within a closed-channel and maintained continuously throughout the culture process. Functional islet-like endocrine cell aggregates were reproducibly induced following a 30-day differentiation protocol. Our system shows an easily scalable, novel method for inducing PSC differentiation with both purity and functionality.

The differentiation of pancreatic endocrine cells from human pluripotent stem cells has been thoroughly investigated for their application in cell therapy against diabetes. Although non-endocrine cells are inevitable contaminating by-products of the differentiation process, a comprehensive profile of such cells is lacking. Therefore, we characterized non-endocrine cells in iPSC-derived pancreatic islet cells (iPIC) using single-cell transcriptomic analysis. We found that non-endocrine cells consist of (1) heterogeneous proliferating cells, and (2) cells with not only pancreatic traits but also liver or intestinal traits marked by FGB or AGR2. Non-endocrine cells specifically expressed FGFR2, PLK1, and LDHB. We demonstrated that inhibition of pathways involving these genes selectively reduced the number of non-endocrine cells in the differentiation process. These findings provide useful insights into cell purification approaches and contribute to the improvement of the mass production of endocrine cells for stem cell-derived cell therapy for diabetes.