Mature Β-cells Research Articles

177-OR: The Swi/Snf Chromatin Remodeling Complex Controls the Specification, Expansion, and Differentiation of Pancreatic Endocrine Progenitor Cells

A key feature of type 1 and type 2 diabetes is insufficient functional β-cell mass. Strategies to augment functional β-cell mass include directed differentiation of embryonic stem cells towards a β-cell fate, which requires extensive knowledge of the transcriptional programs governing endocrine cell differentiation in vivo. Pdx1, an essential transcription factor involved in pancreas formation, endocrine progenitor cell expansion and β-cell function, dynamically recruits coregulators to drive gene expression programs in those cells vital for its activity. Pdx1 binds to numerous coregulators in the β-cell, including the ATP-dependent Swi/Snf chromatin remodeling complex. Previous work by our group revealed a critical role for Pdx1:Swi/Snf in both the developing pancreas and mature β-cell. Here we interrogate the role of Swi/Snf on governing Pdx1-dependent gene expression programs and chromatin accessibility in pancreatic endocrine cell development. We developed a mouse model to eliminate the activity of the two core ATPase subunits of Swi/Snf, Brg1 and Brm from endocrine progenitor cells using Neurogenin3-Cre (Δendo). Conditional knockout of Brg1 (Brg1ΔendoBrm+/-) from endocrine progenitor cells leads to severe glucose intolerance, ad-libitum fed hyperglycemia, reduced serum insulin, compromised glucose stimulated insulin secretion from isolated islets and reduced islet mass in 4-week old animals. Whereas Brg1Δendo/+Brm-/- mice are normal, remarkably, no DKOΔendo (double knockout (DKO): targeted both Brg1 and Brm) mice have been found at weaning, suggesting loss of all Swi/Snf activity from the endocrine progenitor cell pool is postnatally lethal. Future experiments will be directed at determining how Swi/Snf controls Pdx1-dependent gene expression programs and chromatin accessibility in endocrine progenitor cells that are critical for islet cell expansion, differentiation and maturation. Disclosure J. Spaeth: None. R. K. Davidson: None. N. Casey: None. Funding National Institutes of Health (R03DK127129, K01DK115633)

Antagonistic epistasis of Hnf4α and FoxO1 metabolic networks through enhancer interactions in β-cell function

ObjectiveGenetic and acquired abnormalities contribute to pancreatic β-cell failure in diabetes. Transcription factors Hnf4α (MODY1) and FoxO1 are respective examples of these two components and act through β-cell-specific enhancers. However, their relationship is unclear. MethodsIn this report, we show by genome-wide interrogation of chromatin modifications that ablation of FoxO1 in mature β-cells enriches active Hnf4α enhancers according to a HOMER analysis. ResultsTo model the functional significance of this predicted unusual enhancer utilization, we generated single and compound knockouts of FoxO1 and Hnf4α in β-cells. Single knockout of either gene impaired insulin secretion in mechanistically distinct fashions as indicated by their responses to sulfonylurea and calcium fluxes. Surprisingly, the defective β-cell secretory function of either single mutant in hyperglycemic clamps and isolated islets treated with various secretagogues was completely reversed in double mutants lacking FoxO1 and Hnf4α. Gene expression analyses revealed distinct epistatic modalities by which the two transcription factors regulate networks associated with reversal of β-cell dysfunction. An antagonistic network regulating glycolysis, including β-cell “disallowed” genes, and a synergistic network regulating protocadherins emerged as likely mediators of the functional restoration of insulin secretion. ConclusionsThe findings provide evidence of antagonistic epistasis as a model of gene/environment interactions in the pathogenesis of β-cell dysfunction.

Metformin Perturbs Pancreatic Differentiation From Human Embryonic Stem Cells.

Metformin is becoming a popular treatment before and during pregnancy, but current literature on in utero exposure to metformin lacks long-term clinical trials and mechanistic studies. Current literature on the effects of metformin on mature pancreatic β-cells highlights its dual, opposing, protective, or inhibitory effects, depending on metabolic environment. However, the impact of metformin on developing human pancreatic β-cells remains unknown. In this study, we investigated the potential effects of metformin exposure on human pancreatic β-cell development and function in vitro. In the absence of metabolic challenges such as high levels of glucose and fatty acids, metformin exposure impaired the development and function of pancreatic β-cells, with downregulation of pancreatic genes and dysfunctional mitochondrial respiration. It also affected the insulin secretion function of pancreatic β-cells. These findings call for further in-depth evaluation of the exposure of human embryonic and fetal tissue during pregnancy to metformin and its implications for long-term offspring health.

Temporal Transcriptome Analysis Reveals Dynamic Gene Expression Patterns Driving β-Cell Maturation.

Newly differentiated pancreatic β cells lack proper insulin secretion profiles of mature functional β cells. The global gene expression differences between paired immature and mature β cells have been studied, but the dynamics of transcriptional events, correlating with temporal development of glucose-stimulated insulin secretion (GSIS), remain to be fully defined. This aspect is important to identify which genes and pathways are necessary for β-cell development or for maturation, as defective insulin secretion is linked with diseases such as diabetes. In this study, we assayed through RNA sequencing the global gene expression across six β-cell developmental stages in mice, spanning from β-cell progenitor to mature β cells. A computational pipeline then selected genes differentially expressed with respect to progenitors and clustered them into groups with distinct temporal patterns associated with biological functions and pathways. These patterns were finally correlated with experimental GSIS, calcium influx, and insulin granule formation data. Gene expression temporal profiling revealed the timing of important biological processes across β-cell maturation, such as the deregulation of β-cell developmental pathways and the activation of molecular machineries for vesicle biosynthesis and transport, signal transduction of transmembrane receptors, and glucose-induced Ca2+ influx, which were established over a week before β-cell maturation completes. In particular, β cells developed robust insulin secretion at high glucose several days after birth, coincident with the establishment of glucose-induced calcium influx. Yet the neonatal β cells displayed high basal insulin secretion, which decreased to the low levels found in mature β cells only a week later. Different genes associated with calcium-mediated processes, whose alterations are linked with insulin resistance and deregulation of glucose homeostasis, showed increased expression across β-cell stages, in accordance with the temporal acquisition of proper GSIS. Our temporal gene expression pattern analysis provided a comprehensive database of the underlying molecular components and biological mechanisms driving β-cell maturation at different temporal stages, which are fundamental for better control of the in vitro production of functional β cells from human embryonic stem/induced pluripotent cell for transplantation-based type 1 diabetes therapy.

Jazf1 acts as a regulator of insulin-producing β-cell differentiation in induced pluripotent stem cells and glucose homeostasis in mice.

Genetic susceptibility of type 2 diabetes and Juxtaposed with another zinc finger protein 1 (Jazf1) has been reported; however, the precise role of Jazf1 in metabolic processes remains elusive. In this study, using Jazf1-knockout (KO)-induced pluripotent stem cells (iPSC), pancreatic beta cell line MIN6 cells, and Jazf-1 heterozygous KO (Jazf1+/- ) mice, the effect of Jazf1 on gradual differentiation was investigated. We checked the alterations of the genes related with β-cell specification, maturation, and insulin release against glucose treatment by the gain and loss of the Jazf1 gene in the MIN6 cells. Because undifferentiated Jazf1-KO iPSC were not significantly different from wild-type (WT) iPSC, the size and endoderm marker expression after embryoid body (EB) and teratoma formation were investigated. Compared to EB and teratomas formed with WT iPSC, the EB and teratomas from with Jazf1-KO iPSC were smaller, and in teratomas, the expression of proliferation markers was reduced. Moreover, the expression of the gene sets for β-cell differentiation and the levels of insulin and C-peptide secreted by insulin precursor cells were notably reduced in β-cells differentiated from Jazf1-KO iPSC compared with those differentiated from WT iPSC. A comparison of Jazf1+/- and WT mice showed that Jazf1+/- mice had lower levels of serum insulin, pancreatic insulin expression, and decreased pancreatic β-cell size, which resulted in defects in the glucose homeostasis. These findings suggest that Jazf1 plays a pivotal role in the differentiation of β-cells and glucose homeostasis.

Scrt1, a transcriptional regulator of \u03b2-cell proliferation identified by differential chromatin accessibility during islet maturation

Glucose-induced insulin secretion, a hallmark of mature β-cells, is achieved after birth and is preceded by a phase of intense proliferation. These events occurring in the neonatal period are decisive for establishing an appropriate functional β-cell mass that provides the required insulin throughout life. However, key regulators of gene expression involved in functional maturation of β-cells remain to be elucidated. Here, we addressed this issue by mapping open chromatin regions in newborn versus adult rat islets using the ATAC-seq assay. We obtained a genome-wide picture of chromatin accessible sites (~ 100,000) among which 20% were differentially accessible during maturation. An enrichment analysis of transcription factor binding sites identified a group of transcription factors that could explain these changes. Among them, Scrt1 was found to act as a transcriptional repressor and to control β-cell proliferation. Interestingly, Scrt1 expression was controlled by the transcriptional repressor RE-1 silencing transcription factor (REST) and was increased in an in vitro reprogramming system of pancreatic exocrine cells to β-like cells. Overall, this study led to the identification of several known and unforeseen key transcriptional events occurring during β-cell maturation. These findings will help defining new strategies to induce the functional maturation of surrogate insulin-producing cells.

Redox Homeostasis in Pancreatic β-Cells: From Development to Failure.

Redox status is a key determinant in the fate of β-cell. These cells are not primarily detoxifying and thus do not possess extensive antioxidant defense machinery. However, they show a wide range of redox regulating proteins, such as peroxiredoxins, thioredoxins or thioredoxin reductases, etc., being functionally compartmentalized within the cells. They keep fragile redox homeostasis and serve as messengers and amplifiers of redox signaling. β-cells require proper redox signaling already in cell ontogenesis during the development of mature β-cells from their progenitors. We bring details about redox-regulated signaling pathways and transcription factors being essential for proper differentiation and maturation of functional β-cells and their proliferation and insulin expression/maturation. We briefly highlight the targets of redox signaling in the insulin secretory pathway and focus more on possible targets of extracellular redox signaling through secreted thioredoxin1 and thioredoxin reductase1. Tuned redox homeostasis can switch upon chronic pathological insults towards the dysfunction of β-cells and to glucose intolerance. These are characteristics of type 2 diabetes, which is often linked to chronic nutritional overload being nowadays a pandemic feature of lifestyle. Overcharged β-cell metabolism causes pressure on proteostasis in the endoplasmic reticulum, mainly due to increased demand on insulin synthesis, which establishes unfolded protein response and insulin misfolding along with excessive hydrogen peroxide production. This together with redox dysbalance in cytoplasm and mitochondria due to enhanced nutritional pressure impact β-cell redox homeostasis and establish prooxidative metabolism. This can further affect β-cell communication in pancreatic islets through gap junctions. In parallel, peripheral tissues losing insulin sensitivity and overall impairment of glucose tolerance and gut microbiota establish local proinflammatory signaling and later systemic metainflammation, i.e., low chronic inflammation prooxidative properties, which target β-cells leading to their dedifferentiation, dysfunction and eventually cell death.

A developmental lineage-based gene co-expression network for mouse pancreatic β-cells reveals a role for Zfp800 in pancreas development.

To gain a deeper understanding of pancreatic β-cell development, we used iterative weighted gene correlation network analysis to calculate a gene co-expression network (GCN) from 11 temporally and genetically defined murine cell populations. The GCN, which contained 91 distinct modules, was then used to gain three new biological insights. First, we found that the clustered protocadherin genes are differentially expressed during pancreas development. Pcdhγ genes are preferentially expressed in pancreatic endoderm, Pcdhβ genes in nascent islets, and Pcdhα genes in mature β-cells. Second, after extracting sub-networks of transcriptional regulators for each developmental stage, we identified 81 zinc finger protein (ZFP) genes that are preferentially expressed during endocrine specification and β-cell maturation. Third, we used the GCN to select three ZFPs for further analysis by CRISPR mutagenesis of mice. Zfp800 null mice exhibited early postnatal lethality, and at E18.5 their pancreata exhibited a reduced number of pancreatic endocrine cells, alterations in exocrine cell morphology, and marked changes in expression of genes involved in protein translation, hormone secretion and developmental pathways in the pancreas. Together, our results suggest that developmentally oriented GCNs have utility for gaining new insights into gene regulation during organogenesis.

Single Molecule-Based fliFISH Validates Radial and Heterogeneous Gene Expression Patterns in Pancreatic Islet β-Cells.

Single-cell RNA-sequencing (scRNA-Seq) technologies have greatly enhanced our understanding of islet cell transcriptomes and have revealed the existence of β-cell heterogeneity. However, comparison of scRNA-Seq data sets from different groups have highlighted inconsistencies in gene expression patterns, primarily due to variable detection of lower abundance transcripts. Furthermore, such analyses are unable to uncover the spatial organization of heterogeneous gene expression. In this study, we used fluctuation localization imaging–based fluorescence in situ hybridization (fliFISH) to quantify transcripts in single cells in mouse pancreatic islet sections. We compared the expression patterns of Insulin 2 (Ins2) with Mafa and Ucn3, two genes expressed in β-cells as they mature, as well as Rgs4, a factor with variably reported expression in the islet. This approach accurately quantified transcripts across a wide range of expression levels, from single copies to >100 copies/cell in one islet. Importantly, fliFISH allowed evaluation of transcript heterogeneity in the spatial context of an intact islet. These studies confirm the existence of a high degree of heterogeneous gene expression levels within the islet and highlight relative and radial expression patterns that likely reflect distinct β-cell maturation states along the radial axis of the islet.

Pancreatic β-cell heterogeneity in health and diabetes: classes, sources, and subtypes

Pancreatic β-cells perform glucose-stimulated insulin secretion, a process at the center of type 2 diabetes etiology. Efforts to understand how β-cells behave in healthy and stressful conditions have revealed a wide degree of morphological, functional, and transcriptional heterogeneity. Sources of heterogeneity include β-cell topography, developmental origin, maturation state, and stress response. Advances in sequencing and imaging technologies have led to the identification of β-cell subtypes, which play distinct roles in the islet niche. This review examines β-cell heterogeneity from morphological, functional, and transcriptional perspectives, and considers the relevance of topography, maturation, development, and stress response. It also discusses how these factors have been used to identify β-cell subtypes, and how heterogeneity is impacted by diabetes. We examine open questions in the field and discuss recent technological innovations that could advance understanding of β-cell heterogeneity in health and disease.

Virgin β-Cells at the Neogenic Niche Proliferate Normally and Mature Slowly.

Proliferation of pancreatic β-cells has long been known to reach its peak in the neonatal stages and decline during adulthood. However, β-cell proliferation has been studied under the assumption that all β-cells constitute a single, homogenous population. It is unknown whether a subpopulation of β-cells retains the capacity to proliferate at a higher rate and thus contributes disproportionately to the maintenance of mature β-cell mass in adults. We therefore assessed the proliferative capacity and turnover potential of virgin β-cells, a novel population of immature β-cells found at the islet periphery. We demonstrate that virgin β-cells can proliferate but do so at rates similar to those of mature β-cells from the same islet under normal and challenged conditions. Virgin β-cell proliferation rates also conform to the age-dependent decline previously reported for β-cells at large. We further show that virgin β-cells represent a long-lived, stable subpopulation of β-cells with low turnover into mature β-cells under healthy conditions. Our observations indicate that virgin β-cells at the islet periphery can divide but do not contribute disproportionately to the maintenance of adult β-cell mass.

The SNAG Domain of Insm1 Regulates Pancreatic Endocrine Cell Differentiation and Represses β- to δ-Cell Transdifferentiation.

The allocation and specification of pancreatic endocrine lineages are tightly regulated by transcription factors. Disturbances in differentiation of these lineages contribute to the development of various metabolic diseases, including diabetes. The insulinoma-associated protein 1 (Insm1), which encodes a protein containing one SNAG domain and five zinc fingers, plays essential roles in pancreatic endocrine cell differentiation and in mature β-cell function. In the current study, we compared the differentiation of pancreatic endocrine cells between Insm1 null and Insm1 SNAG domain mutants (Insm1delSNAG) to explore the specific function of the SNAG domain of Insm1. We show that the δ-cell number is increased in Insm1delSNAG but not in Insm1 null mutants as compared with the control mice. We also show a less severe reduction of the β-cell number in Insm1delSNAG as that in Insm1 null mutants. In addition, similar deficits are observed in α-, PP, and ε-cells in Insm1delSNAG and Insm1 null mutants. We further identified that the increased δ-cell number is due to β- to δ-cell transdifferentiation. Mechanistically, the SNAG domain of Insm1 interacts with Lsd1, the demethylase of H3K4me1/2. Mutation in the SNAG domain of Insm1 results in impaired recruitment of Lsd1 and increased H3K4me1/2 levels at hematopoietically expressed homeobox (Hhex) loci that are bound by Insm1, thereby promoting the transcriptional activity of the δ-cell-specific gene Hhex Our study has identified a novel function of the SNAG domain of Insm1 in the regulation of pancreatic endocrine cell differentiation, particularly in the repression of β- to δ-cell transdifferentiation.

In vitro Characterization of Insulin−Producing β-Cell Spheroids

Over the years, immortalized rodent β-cell lines such as RIN, HIT, MIN, βTC, and INS-1 have been used to investigate pancreatic β-cell physiology using conventional two-dimensional (2D) culture techniques. However, physical and physiological limitations inherent to 2D cell culture necessitates confirmatory follow up studies using sentient animals. Three-dimensional (3D) culture models are gaining popularity for their recapitulation of key features of in vivo organ physiology, and thus could pose as potential surrogates for animal experiments. In this study, we aimed to develop and characterize a rat insulinoma INS-1 3D spheroid model to compare with 2D monolayers of the same cell line. Ultrastructural verification was done by transmission electron microscopy and toluidine blue staining, which showed that both 2D monolayers and 3D spheroids contained highly granulated cells with ultrastructural features synonymous with mature pancreatic β-cells, with increased prominence of these features observed in 3D spheroids. Viability, as assessed by cellular ATP quantification, size profiling and glucose utilization, showed that our spheroids remained viable for the experimental period of 30 days, compared to the limiting 5-day passage period of INS-1 monolayers. In fact, increasing ATP content together with spheroid size was observed over time, without adverse changes in glucose utilization. Additionally, β-cell function, assessed by determining insulin and amylin secretion, showed that the 3D spheroids retained glucose sensing and insulin secretory capability, that was more acute when compared to 2D monolayer cultures. Thus, we were able to successfully demonstrate that our in vitro INS-1 β-cell 3D spheroid model exhibits in vivo tissue-like structural features with extended viability and lifespan. This offers enhanced predictive capacity of the model in the study of metabolic disease, β-cell pathophysiology and the potential treatment thereof.

Establishment of a long-term stable \u03b2-cell line and its application to analyze the effect of Gcg expression on insulin secretion

A pancreatic β-cell line MIN6 was previously established in our lab from an insulinoma developed in an IT6 transgenic mouse expressing the SV40 T antigen in β-cells. This cell line has been widely used for in vitro analysis of β-cell function, but tends to lose the mature β-cell features, including glucose-stimulated insulin secretion (GSIS), in long-term culture. The aim of this study was to develop a stable β-cell line that retains the characteristics of mature β-cells. Considering that mice derived from a cross between C3H and C57BL/6 strains are known to exhibit higher insulin secretory capacity than C57BL/6 mice, an IT6 male mouse of this hybrid background was used to isolate insulinomas, which were independently cultured. After 7 months of continuous culturing, we obtained the MIN6-CB4 β-cell line, which stably maintains its GSIS. It has been noted that β-cell lines express the glucagon (Gcg) gene at certain levels. MIN6-CB4 cells were utilized to assess the effects of differential Gcg expression on β-cell function. Our data show the functional importance of Gcg expression and resulting basal activation of the GLP-1 receptor in β-cells. MIN6-CB4 cells can serve as an invaluable tool for studying the regulatory mechanisms of insulin secretion, such as the GLP-1/cAMP signaling, in β-cells.

In vivo Imaging of Calcium Activities from Pancreatic β-cells in Zebrafish Embryos Using Spinning-disc Confocal and Two-photon Light-sheet Microscopy.

Visualizing the function of pancreatic β-cells in vivo has been a long-sought goal for β-cell researchers. Unlike imaging of β-cells in mammalian species with conventional positron emission tomography and single-photon emission computed tomography, which only provides limited spatial-temporal resolution, transparent zebrafish embryos are a unique model that allows high-resolution fluorescent imaging of β-cells in their native physiological microenvironment in vivo. Here, we detail a protocol for real-time visualization of individual β-cell function in vivo in a non-invasive manner, through combination of a novel transgenic zebrafish reporter line Tg (ins:Rcamp1.07) with both a commercial spinning-disc confocal microscope and an in-house developed super-resolution microscope (2P3A-DSLM). The protocol described here allows for the longitudinal monitoring of dynamic calcium activities from heterogeneous β-cells in early developing zebrafish embryos and is readily adaptable for use in imaging other important processes in islet biology, as well as screening new compounds that can promote β-cell function or maturation using a living whole organism system.

Recent Advances in the Generation of β-Cells from Induced Pluripotent Stem Cells as a Potential Cure for Diabetes Mellitus.

Diabetes mellitus (DM) is a group of metabolic disorders characterized by high blood glucose levels due to insufficient insulin secretion, insulin action, or both. The present-day solution to diabetes mellitus includes regular administration of insulin, which brings about many medical complications in diabetic patients. Although islet transplantation from cadaveric subjects was proposed to be a permanent cure, the increased risk of infections, the need for immunosuppressive drugs, and their unavailability had restricted its use. To overcome this, the generation of renewable and transplantable β-cells derived from autologous induced pluripotent stem cells (iPSCs) has gained enormous interest as a potential therapeutic strategy to treat diabetes mellitus permanently. To date, extensive research has been undertaken to derive transplantable insulin-producing β-cells (iβ-cells) from iPSCs in vitro by recapitulating the in vivo developmental process of the pancreas. This in vivo developmental process relies on transcription factors, signaling molecules, growth factors, and culture microenvironment. This review highlights the various factors facilitating the generation of mature β-cells from iPSCs. Moreover, this review also describes the generation of pancreatic progenitors and β-cells from diabetic patient-specific iPSCs, exploring the potential of the diabetes disease model and drug discovery. In addition, the applications of genome editing strategies have also been discussed to achieve patient-specific diabetes cell therapy. Last, we have discussed the current challenges and prospects of iPSC-derived β-cells to improve the relative efficacy of the available treatment of diabetes mellitus.

Generation of Functional Insulin-Producing Cells from Mouse Embryonic Stem Cells Through Protein Transduction of Transcription Factors.

In this chapter, we describe a simple and unique method for the differentiation of mouse embryonic stem cells into insulin-producing cells. In addition to cytokines and growth factors, key transcription factors for pancreatic development are applied in this method through protein transduction technology. Furthermore, a combination of nanofiber plates and laminin coatings improves the yield of differentiated cells. The insulin-producing cells derived through this method express marker genes of mature β-cells and have an ability to secrete insulin; therefore, these cells are useful for fundamental studies on pancreatic development, drug development, and regenerative medicine for diabetes.

Transcriptomic and Quantitative Proteomic Profiling Reveals Signaling Pathways Critical for Pancreatic Islet Maturation.

Pancreatic β-cell dysfunction and reduced insulin secretion play a key role in the pathogenesis of diabetes. Fetal and neonatal islets are functionally immature and have blunted glucose responsiveness and decreased insulin secretion in response to stimuli and are far more proliferative. However, the mechanisms underlying functional immaturity are not well understood. Pancreatic islets are composed of a mixture of different cell types, and the microenvironment of islets and interactions between these cell types are critical for β-cell development and maturation. RNA sequencing and quantitative proteomic data from intact islets isolated from fetal (embryonic day 19) and 2-week-old Sprague-Dawley rats were integrated to compare their gene and protein expression profiles. Ingenuity Pathway Analysis (IPA) was also applied to elucidate pathways and upstream regulators modulating functional maturation of islets. By integrating transcriptome and proteomic data, 917 differentially expressed genes/proteins were identified with a false discovery rate of less than 0.05. A total of 411 and 506 of them were upregulated and downregulated in the 2-week-old islets, respectively. IPA revealed novel critical pathways associated with functional maturation of islets, such as AMPK (adenosine monophosphate-activated protein kinase) and aryl hydrocarbon receptor signaling, as well as the importance of lipid homeostasis/signaling and neuronal function. Furthermore, we also identified many proteins enriched either in fetal or 2-week-old islets related to extracellular matrix and cell communication, suggesting that these pathways play critical roles in islet maturation. Our present study identified novel pathways for mature islet function in addition to confirming previously reported mechanisms, and provided new mechanistic insights for future research on diabetes prevention and treatment.

Topologically selective islet vulnerability and self-sustained downregulation of markers for \u03b2-cell maturity in streptozotocin-induced diabetes

Mouse models of Streptozotocin (STZ) induced diabetes represent the most widely used preclinical diabetes research systems. We applied state of the art optical imaging schemes, spanning from single islet resolution to the whole organ, providing a first longitudinal, 3D-spatial and quantitative account of β-cell mass (BCM) dynamics and islet longevity in STZ-treated mice. We demonstrate that STZ-induced β-cell destruction predominantly affects large islets in the pancreatic core. Further, we show that hyperglycemic STZ-treated mice still harbor a large pool of remaining β-cells but display pancreas-wide downregulation of glucose transporter type 2 (GLUT2). Islet gene expression studies confirmed this downregulation and revealed impaired β-cell maturity. Reversing hyperglycemia by islet transplantation partially restored the expression of markers for islet function, but not BCM. Jointly our results indicate that STZ-induced hyperglycemia results from β-cell dysfunction rather than β-cell ablation and that hyperglycemia in itself sustains a negative feedback loop restraining islet function recovery.

In Vivo Differentiation of Stem Cell-derived Human Pancreatic Progenitors to Treat Type 1 Diabetes.

Type 1 diabetes mellitus (T1DM) is an autoimmune disease that results from the loss of the pancreatic β-cells. The autoimmune destruction of the β-cells causes the loss of insulin production from the islets of the pancreas, resulting in the loss of blood glucose regulation. This loss of regulation, if not treated, can lead to a plethora of long-term complications in patients. Subsequently, T1DM patients rely on the administration of exogenous insulin sources to maintain their blood glucose levels. In this review, we summarize the history of T1DM therapy and current treatment options. Although treatments for T1DM have progressed substantially, none of the available treatment options allow the patient to live autonomously. Therefore, the challenge to develop a therapy that will fully reverse the disease still remains. A promising field of T1DM therapies is cell replacement therapies derived from human pluripotent stem cells. Here, we specifically review studies that employ stem-cell derived pancreatic progenitors transplanted for in vivo differentiation/maturation and discuss, in detail, the complications that arise post transplantation, including heterogeneity, graft immaturity, and host foreign bodyresponse. We also discuss efforts to induce human stem cell-derived mature β-cells in vitro and compare strategies regarding transplantation of pancreatic progenitors versus mature β-cells cells. Finally, we review key approaches that address critical limitations of in vivo progenitor differentiation including vascularization, oxygenation, and transplant location.The field of isletreplacement therapy has made tremendous progress in the last two decades. Ifthe strengths and limitations of the field continue to be identified andaddressed, future studies will lead to an idealtreatment for T1DM. Graphical abstract.