Human pluripotent stem cell-derived β cells (hPSC-β cells) show the potential to restore euglycemia. However, the immature functionality of hPSC-β cells has limited their efficacy in application. Here, by deciphering the continuous maturation process of hPSC-β cells post transplantation via single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq), we show that functional maturation of hPSC-β cells is an orderly multistep process during which cells sequentially undergo metabolic adaption, removal of negative regulators of cell function, and establishment of a more specialized transcriptome and epigenome. Importantly, remodeling lipid metabolism, especially downregulating the metabolic activity of ceramides, the central hub of sphingolipid metabolism, is critical for β cell maturation. Limiting intracellular accumulation of ceramides in hPSC-β cells remarkably enhanced their function, as indicated by improvements in insulin processing and glucose-stimulated insulin secretion. In summary, our findings provide insights into the maturation of human pancreatic β cells and highlight the importance of ceramide homeostasis in function acquisition.

人类多能干细胞衍生的 β 细胞（hPSC-β 细胞）显示出恢复血糖正常的潜力。然而，hPSC-β细胞的功能不成熟限制了其应用功效。在这里，通过单细胞 RNA 测序 (scRNA-seq) 和转座酶可及染色质测序的单细胞测定 (scATAC-seq) 破译 hPSC-β 细胞移植后的持续成熟过程，我们表明 hPSC 的功能成熟-β细胞是一个有序的多步骤过程，在此过程中细胞依次经历代谢适应、去除细胞功能的负调节因子以及建立更专门的转录组和表观基因组。重要的是，重塑脂质代谢，特别是下调神经酰胺（鞘脂代谢的中枢）的代谢活性，对于 β 细胞成熟至关重要。限制 hPSC-β 细胞中神经酰胺的细胞内积累可显着增强其功能，胰岛素加工和葡萄糖刺激的胰岛素分泌的改善表明了这一点。总之，我们的研究结果提供了对人胰腺 β 细胞成熟的见解，并强调了神经酰胺稳态在功能获得中的重要性。