Functional, metabolic and transcriptional maturation of human pancreatic islets derived from stem cells

Abstract

Transplantation of pancreatic islet cells derived from human pluripotent stem cells is a promising treatment for diabetes. Despite progress in the generation of stem-cell-derived islets (SC-islets), no detailed characterization of their functional properties has been conducted. Here, we generated functionally mature SC-islets using an optimized protocol and benchmarked them comprehensively against primary adult islets. Biphasic glucose-stimulated insulin secretion developed during in vitro maturation, associated with cytoarchitectural reorganization and the increasing presence of alpha cells. Electrophysiology, signaling and exocytosis of SC-islets were similar to those of adult islets. Glucose-responsive insulin secretion was achieved despite differences in glycolytic and mitochondrial glucose metabolism. Single-cell transcriptomics of SC-islets in vitro and throughout 6 months of engraftment in mice revealed a continuous maturation trajectory culminating in a transcriptional landscape closely resembling that of primary islets. Our thorough evaluation of SC-islet maturation highlights their advanced degree of functionality and supports their use in further efforts to understand and combat diabetes.

Maturation of beta cells: lessons from in vivo and in vitro models

“…Stimulation indices, calcium fluxes and cyclical oxygen consumption were all increased following entrainment. In agreement with this, following SC-islet implantation and maturation in vivo, core clock components BMAL1, RORA and BHLHE41 were all upregulated, showing that enhanced beta cell maturation correlates with enhanced expression of core clock components [35]. Perhaps most intriguingly, the recent finding that circadian clock cycling may regulate the alternative splicing of subsets of target genes within beta cells adds a new dimension to the concept of circadian control of beta cell maturation and transcriptional regulation [51].…”

“…1). This acquisition of heightened OxPhos activity during maturation is mirrored in SC-islet studies that demonstrate increased abundance of OxPhos-related genes following enhanced in vitro culture conditions and maturation during murine engraftment [35,60,61]. Proteomics, transcriptomics and metabolomics studies within rat islets, as they transition from juvenile-to-adult states, also display enhanced OxPhos gene network signatures [62,63].…”

“…In line with this, a study reported that glycolytically derived dihydroxyacetone phosphate (DHAP) may signal glucose availability directly to the mTORC1 complex (albeit in human embryonic kidney cells) [33]. Metabolic tracing studies by us and others have shown that SC-islets show strong functional profiles despite limited mitochondrial metabolism of glucose; furthermore, a glycolytic bottleneck beyond the glyceraldehyde-3-phosphate (GA3P)/DHAP enzymatic step is present in SC-islets [34,35]. It is tempting to speculate that such a direct interplay of glycolytic DHAP generation and mTOR activity may in part be responsible for strong in vitro SC-islet function, without the canonical mitochondrial coupling seen in mature adult islets.…”

“…This pattern is also true of aspartate and glutamate, possibly demonstrating an underdeveloped function of malate-aspartate shuttle activity within immature SC-islets as well as limited intracellular glutamate signalling, which has also been implicated as a triggering and amplifying messenger within GSIS [83,84]. Interestingly, metabolic signatures of the pyruvate-isocitrate cycle are present within SC-islets without extended maturation periods and despite low glucose-responsive OxPhos metabolism [35].…”

“…1). Clues relating to this acquisition of glutathionebased redox signalling are seen in human SC-islet models of maturation whereby a multitude of glutathione-related genes, as well as de novo production of glutathione itself, are increased following extended maturation periods in engrafted mice and primary islets [35]. This pattern is also true of aspartate and glutamate, possibly demonstrating an underdeveloped function of malate-aspartate shuttle activity within immature SC-islets as well as limited intracellular glutamate signalling, which has also been implicated as a triggering and amplifying messenger within GSIS [83,84].…”