Single-Cell RNA-Seq Reveals that CD9 Is a Negative Marker of Glucose-Responsive Pancreatic β-like Cells Derived from Human Pluripotent Stem Cells

To date, it remains unclear if there are specific cell-surface markers for purifying glucose-responsive pancreatic β-like cells derived from human pluripotent stem cells (hPSCs). In searching for this, we generated an efficient protocol for differentiating β-like cells from human embryonic stem cells. We performed single-cell RNA sequencing and found that CD9 is a negative cell-surface marker of β-like cells, as most INS(+) cells are CD9(−). We purified β-like cells for spontaneous formation of islet-like organoids against CD9, and found significantly more NKX6.1(+)MAFA(+)C-PEPTIDE(+) β-like cells in the CD9(−) than in the CD9(+) population. CD9(−) cells also demonstrate better glucose responsiveness than CD9(+) cells. In humans, we observe more CD9(+)C-PEPTIDE(+) β cells in the fetal than in the adult cadaveric islets and more Ki67(+) proliferating cells among CD9(+) fetal β cells. Taken together, our experiments show that CD9 is a cell-surface marker for negative enrichment of glucose-responsive β-like cells differentiated from hPSCs.