Restoration of PKM1 improves functional maturation of stem-cell derived-b by regulating PEP metabolism

In stem cell-derived β (SC-β) cells, metabolic abnormalities persist as obstacles to glucose responsiveness and functional maturation, with the primary metabolic bottlenecks yet to be identified. This study demonstrated that restoring pyruvate kinase 1 (PKM1) re-established pyruvate kinase activity, effectively reversing the SC-β-cell identity loss and functional impairment associated with phosphoenolpyruvate (PEP) accumulation. 13C-glucose labeling metabolomics revealed a disruption in glucose-stimulated metabolite production beginning at the glycolytic PEP stage in stem cell-derived islets (SC-islets). Uniquely, elevated PEP prevented glycolytic metabolite-dependent increase of exocytosis and significantly increased intracellular calcium levels through a KATP channel-independent pathway, even under basal glucose condition. Furthermore, exposure to PEP led to downregulated expression of genes involved in the TCA cycle and respiratory electron transport. This PEP accumulation in SC-islets was associated with markedly reduced pyruvate kinase activity. Overexpression of PKM1 rescued the transcriptional changes induced by PEP accumulation and improved both glucose-stimulated calcium responses and insulin secretion. These findings suggest that PKM1 plays a pivotal role in promoting SC-β cell differentiation and functional maturation by regulating PEP metabolism, offering potential improvements for SC-β cell replacement in diabetes treatment.