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Islet-enriched transcription factors (TFs) exert broad control over cellular processes in pancreatic α and β cells, and changes in their expression are associated with developmental state and diabetes. However, the implications of heterogeneity in TF expression across islet cell populations are not well understood. To define this TF heterogeneity and its consequences for cellular function, we profiled more than 40,000 cells from normal human islets by single-cell RNA-Seq and stratified α and β cells based on combinatorial TF expression. Subpopulations of islet cells coexpressing ARX/MAFB (α cells) and MAFA/MAFB (β cells) exhibited greater expression of key genes related to glucose sensing and hormone secretion relative to subpopulations expressing only one or neither TF. Moreover, all subpopulations were identified in native pancreatic tissue from multiple donors. By Patch-Seq, MAFA/MAFB-coexpressing β cells showed enhanced electrophysiological activity. Thus, these results indicate that combinatorial TF expression in islet α and β cells predicts highly functional, mature subpopulations.

Pancreatic islets secrete insulin from β cells and glucagon from α cells, and dysregulated secretion of these hormones is a central component of diabetes. Thus, an improved understanding of the pathways governing coordinated β and α cell hormone secretion will provide insight into islet dysfunction in diabetes. However, the 3D multicellular islet architecture, essential for coordinated islet function, presents experimental challenges for mechanistic studies of intracellular signaling pathways in primary islet cells. Here, we developed an integrated approach to study the function of primary human islet cells using genetically modified pseudoislets that resemble native islets across multiple parameters. Further, we developed a microperifusion system that allowed synchronous acquisition of GCaMP6f biosensor signal and hormone secretory profiles. We demonstrate the utility of this experimental approach by studying the effects of Gi and Gq GPCR pathways on insulin and glucagon secretion by expressing the designer receptors exclusively activated by designer drugs (DREADDs) hM4Di or hM3Dq. Activation of Gi signaling reduced insulin and glucagon secretion, while activation of Gq signaling stimulated glucagon secretion but had both stimulatory and inhibitory effects on insulin secretion, which occur through changes in intracellular Ca2+. The experimental approach of combining pseudoislets with a microfluidic system allowed the coregistration of intracellular signaling dynamics and hormone secretion and demonstrated differences in GPCR signaling pathways between human β and α cells.

Posttransplantation diabetes mellitus (PTDM) is a common and significant complication related to immunosuppressive agents required to prevent organ or cell transplant rejection. To elucidate the effects of 2 commonly used agents, the calcineurin inhibitor tacrolimus (TAC) and the mTOR inhibitor sirolimus (SIR), on islet function and test whether these effects could be reversed or prevented, we investigated human islets transplanted into immunodeficient mice treated with TAC or SIR at clinically relevant levels. Both TAC and SIR impaired insulin secretion in fasted and/or stimulated conditions. Treatment with TAC or SIR increased amyloid deposition and islet macrophages, disrupted insulin granule formation, and induced broad transcriptional dysregulation related to peptide processing, ion/calcium flux, and the extracellular matrix; however, it did not affect regulation of β cell mass. Interestingly, these β cell abnormalities reversed after withdrawal of drug treatment. Furthermore, cotreatment with a GLP-1 receptor agonist completely prevented TAC-induced β cell dysfunction and partially prevented SIR-induced β cell dysfunction. These results highlight the importance of both calcineurin and mTOR signaling in normal human β cell function in vivo and suggest that modulation of these pathways may prevent or ameliorate PTDM.

Cystic fibrosis-related (CF-related) diabetes (CFRD) is an increasingly common and devastating comorbidity of CF, affecting approximately 35% of adults with CF. However, the underlying causes of CFRD are unclear. Here, we examined cystic fibrosis transmembrane conductance regulator (CFTR) islet expression and whether the CFTR participates in islet endocrine cell function using murine models of β cell CFTR deletion and normal and CF human pancreas and islets. Specific deletion of CFTR from murine β cells did not affect β cell function. In human islets, CFTR mRNA was minimally expressed, and CFTR protein and electrical activity were not detected. Isolated CF/CFRD islets demonstrated appropriate insulin and glucagon secretion, with few changes in key islet-regulatory transcripts. Furthermore, approximately 65% of β cell area was lost in CF donors, compounded by pancreatic remodeling and immune infiltration of the islet. These results indicate that CFRD is caused by β cell loss and intraislet inflammation in the setting of a complex pleiotropic disease and not by intrinsic islet dysfunction from CFTR mutation.