Pancreatic endocrine cells are categorized in to 5 types (alpha, beta, delta, pancreatic polypeptide cells and epsilon), which expresses glucagon, insulin, somatostatin, pancreatic polypeptide, and ghrelin, respectively. Several studies including lineage tracing in Ins2Akita diabetic mice have been done to investigate the identities of pancreatic endocrine cells which concludes, alpha cells have enormous plasticity, which enables them to be reprogrammed by specific transcription factors into insulin secreting beta like cells. Gene therapy has provided the beneficial outcome. Pdx1, MaFA and PAX4 (the transcription factors) in alpha cells can be over expressed which results in reprogramming the targeted alpha cells into beta cells. This trans-differentiation may be induced by infusing an adeno-associated virus (AAV) loaded with distinct transcription factors in the duct of pancreas. Several researches have demonstrated the successful restoration of enhanced insulin secretion in diabetes induced mice. Additionally ductal neurogenin3 (Ngn3), Sglt2 inhibitors, Igfbp1, GLP1 and several clinical and non-clinical agents has been postulated as a basis of beta cell neogenesis. Alpha cell owing to its high plasticity, on prolonged exposure to GABA reprogrammed into beta-like cell due to downregulation of Arx expression by GABA. The various approaches for beta cell neogenesis open a new window towards the establishment of novel gene therapy accession to treat diabetes. However, broad studies are still needed to improve and optimize this treatment methodology. The potentiality of endogenous pancreatic alpha cell to beta cell conversion methods and its outcomes are invigorating. This accomplishment is presently being under trial in non-human primates.

Pregnancy is associated with physiological adaptations that affect virtually all organs, enabling the mother to support the growing fetus and placenta while withstanding the demands of pregnancy. As a result, mammalian pregnancy is a unique state that exerts paradoxical effects on maternal health. On one hand, the metabolic stress induced by pregnancy can accelerate aging and functional decline in organs. On the other hand, pregnancy activates metabolic programming and tissue regenerative responses that can reverse age-related impairments. In this sense, the oocyte-to-blastocyst transition is not the only physiological reprogramming event in the mammalian body, as pregnancy-induced regeneration could constitute a second physiological reprogramming event. Here, we review findings on how pregnancy dualistically leads to aging and rejuvenation in the maternal body.

The adult pancreatic ducts have long been proposed to contain rare progenitors, some of which expressing Ngn3, that generate new beta cells in endocrine-islet homeostasis. Due to their postulated rarity and the lack of definitive markers, the existence or absence of ductal endocrine progenitors remains unsettled despite many studies. Genetic lineage tracing of ductal cells or Ngn3+ cells with currently available CreER drivers has been complicated by off-target labeling of pre-existing beta cells. Here, using dual-recombinase-mediated intersectional genetic strategy and newly-derived Ngn3-2A-CreER and Hnf1b-2A-CreER knock-in drivers, we succeeded in specifically labeling Ngn3-positive cells and Hnf1b-positive ductal cells without marking pre-existing beta cells. These data revealed no evidence of de novo generation of insulin-producing beta cells from ductal cells or endogenous Ngn3-positive cells in the adult pancreas during homeostasis.

Although originating from different germ layers, pancreatic islet cells and neurons share extensive similarities, both physiological (e.g., voltage-dependent release of a bioactive molecule) and molecular (e.g., highly similar composition of transcription factors and structural genes). Here we propose that two seemingly unrelated phenomena recognized in these cell types-neurotransmitter switching in neurons and the expression of two or more hormones in individual islet cells-share a deep resemblance, potentially reflecting an ancient molecular circuit of cell plasticity. Comparing and contrasting dynamic hormone expression in islet cells and transmitter switching in neurons may provide insights into the functions and underlying mechanisms of these phenomena.

The quest to find a progenitor cell in the adult pancreas has driven research in the field for decades. Many potential progenitor cell sources have been reported, but so far this is a matter of debate mainly due to reproducibility issues. The existence of adult Procr+ progenitor cells in mice islets has been recently reported. These were shown to comprise ~1% of islet cells, lack expression of Neurog3 and endocrine hormones, and to be capable of differentiating into all endocrine cell types. However, these findings had limited impact, as further evidence supporting the existence and function of Procr+ progenitors has not emerged.

We report here an unbiased comparison across mouse and human pancreatic samples, including adult islets and embryonic tissue, to track the existence of Procr+ progenitors originally described based on their global gene expression signature. We could not find Procr+ progenitors on other mouse or human adult pancreatic islet samples. Unexpectedly, our results revealed a transcriptionally close mesothelial cell population in the mouse and human embryonic pancreas. These Procr-like mesothelial cells of the embryonic pancreas share the salient transcriptional and epigenomic features of previously reported Procr+ progenitors found in adult pancreatic islets. Notably, we report here that Procr-like transcriptional signature is gradually established in mesothelial cells during mouse pancreas development from E12.5 to E17.5, which has its largest amount. Further supporting a developmentally relevant role in the human pancreas, we additionally report that a transcriptionally similar population is spontaneously differentiated from human pluripotent stem cells cultured in vitro along the pancreatic lineage.

Our results show that, although the previously reported Procr+ progenitor cell population could not be found in other adult pancreatic islet samples, a mesothelial cell population with a closely related transcriptional signature is present in both the mouse and human embryonic pancreas. Several lines of evidence presented in this work support a developmentally relevant function for these Procr-like mesothelial cells.

Diabetes is characterized by variable loss of insulin-producing beta cells, and new regenerative approaches to increasing the functional beta cell mass of patients hold promise for reversing disease progression. In this Review, we summarize recent chemical biology breakthroughs advancing our knowledge of beta cell regeneration. We present current chemical-based tools, sensors and mechanistic insights into pathways that can be targeted to enhance beta cell regeneration in model organisms. We group the pathways according to the cellular processes they affect, that is, proliferation, conversion of other mature cell types to beta cells and beta cell differentiation from progenitor-like populations. We also suggest assays for assessing the functionality of the regenerated beta cells. Although regeneration processes differ between animal models, such as zebrafish, mice and pigs, regenerative mechanisms identified in any one animal model may be translatable to humans. Overall, chemical biology-based approaches in beta cell regeneration give hope that specific molecular pathways can be targeted to enhance beta cell regeneration.

Recent advances in lineage-tracing technologies have significantly improved our understanding of pancreatic cell biology, particularly in elucidating the ontogeny and regenerative capacity of pancreatic cells. A deeper appreciation of the mechanisms underlying pancreatic cell identity and plasticity holds the potential to inform the development of new therapeutic modalities for conditions such as diabetes and pancreatitis. With this goal in mind, here we summarize advances, challenges, and future directions in tracing pancreatic cell origins and fates using lineage-tracing technologies. Given their essential role for blood glucose regulation, we pay particular attention on the insights gained from endocrine cells, especially β-cells.

Bi-hormonal islet endocrine cells have been proposed to represent an intermediate state of cellular transdifferentiation, enabling an increase in beta-cell mass in response to severe metabolic stress. Beta-cell plasticity and regenerative capacity are thought to decrease with age. We investigated the ontogeny of bi-hormonal islet endocrine cell populations throughout the human lifespan. Immunofluorescence microscopy was performed for insulin, glucagon, and somatostatin presence on paraffin-embedded sections of pancreata from 20 donors without diabetes aged between 11 days and 79 years of age. The mean proportional presence of glucagon-, insulin-, and somatostatin-immunoreactive cells within islets was 27.5%, 62.1%, and 12.1%, respectively. There was no change in the relative presence of alpha- or beta-cells with advancing age, but delta-cell presence showed a decline with age (R2 = 0.59, p < 0.001). The most abundant bi-hormonal cell phenotype observed co-stained for glucagon and insulin, representing 3.1 ± 0.3% of all islet cells. Glucagon/somatostatin and insulin/somatostatin bi-hormonal cells were also observed representing 2-3% abundance relative to islet cell number. Glucagon/insulin bi-hormonal cells increased with age (R2 = 0.30, p < 0.05) whilst insulin/somatostatin (R2 = 0.50, p < 0.01) and glucagon/somatostatin (R2 = 0.35, p < 0.05) cells decreased with age of donor. Findings show that bi-hormonal cells are present within human pancreatic islets throughout life, perhaps reflecting an ongoing potential for endocrine cell plasticity.

Pancreatic islet β-cell mass expands during pregnancy, but underlying mechanisms are not fully understood. This study examines the impact of pregnancy and cafeteria diet on islet morphology, associated cellular proliferation/apoptosis rates as well as β-cell lineage.

Non-pregnant and pregnant Ins1Cre/+;Rosa26-eYFP transgenic mice were maintained on either normal or high-fat cafeteria diet, with pancreatic tissue obtained at 18 days gestation. Immunohistochemical changes in islet morphology, β-/α-cell proliferation and apoptosis, as well as islet cell identity, neogenesis and ductal cell transdifferentiation were assessed.

Pregnant normal diet mice displayed an increase in body weight and glycaemia. Cafeteria feeding attenuated this weight gain while causing overt hyperglycaemia. Pregnant mice maintained on a normal diet exhibited typical expansion in islet and β-cell area, owing to increased β-cell proliferation and survival as well as ductal to β-cell transdifferentiation and β-cell neogenesis, alongside decreased β-cell dedifferentiation. Such pregnancy-induced islet adaptations were severely restricted by cafeteria diet. Accordingly, islets from these mice displayed high levels of β-cell apoptosis and dedifferentiation, together with diminished β-cell proliferation and lack of pregnancy-induced β-cell neogenesis and transdifferentiation, entirely opposing islet cell modifications observed in pregnant mice maintained on a normal diet.

Augmentation of β-cell mass during gestation arises through various mechanisms that include proliferation and survival of existing β-cells, transdifferentiation of ductal cells as well as β-cell neogenesis. Remarkably, cafeteria feeding almost entirely annuls pregnancy-induced islet adaptations, which may contribute to the development of gestational diabetes in the setting of dietary provoked metabolic stress.

Diabetes results from an inadequate number of insulin-producing human beta cells. There is currently no clinically available effective means to restore beta cell mass in millions of people with diabetes. Although the DYRK1A inhibitors, either alone or in combination with GLP-1 receptor agonists (GLP-1) or transforming growth factor β (TGF-β) superfamily inhibitors (LY), induce beta cell replication and increase beta cell mass, the precise mechanisms of action remain elusive. Here we perform single-cell RNA sequencing on human pancreatic islets treated with a DYRK1A inhibitor, either alone or with GLP-1 or LY. We identify cycling alpha cells as the most responsive cells to DYRK1A inhibition. Lineage trajectory analyses suggest that cycling alpha cells may serve as precursor cells that transdifferentiate into beta cells. Collectively, in addition to enhancing expression of beta cell phenotypic genes in beta cells, our findings suggest that regenerative drugs may be targeting cycling alpha cells in human islets.

Dysregulated glucagon secretion and inadequate functional beta cell mass are hallmark features of diabetes. While glucagon receptor (GCGR) antagonism ameliorates hyperglycemia and elicits beta cell regeneration in pre-clinical models of diabetes, it also promotes alpha and delta cell hyperplasia. We sought to investigate the mechanism by which loss of glucagon action impacts pancreatic islet non-alpha cells, and the relevance of these observations in a human islet context.

We used zebrafish, rodents, and transplanted human islets comprising six different models of interrupted glucagon signaling to examine their impact on delta and beta cell proliferation and mass. We also used models with global deficiency of the cationic amino acid transporter, SLC7A2, and mTORC1 inhibition via rapamycin, to determine whether amino acid-dependent nutrient sensing was required for islet non-alpha cell growth.

Inhibition of glucagon signaling stimulated delta cell proliferation in mouse and transplanted human islets, and in mouse islets. This was rapamycin-sensitive and required SLC7A2. Likewise, gcgr deficiency augmented beta cell proliferation via SLC7A2- and mTORC1-dependent mechanisms in zebrafish and promoted cell cycle engagement in rodent beta cells but was insufficient to drive a significant increase in beta cell mass in mice.

Our findings demonstrate that interruption of glucagon signaling augments islet non-alpha cell proliferation in zebrafish, rodents, and transplanted human islets in a manner requiring SLC7A2 and mTORC1 activation. An increase in delta cell mass may be leveraged for future beta cell regeneration therapies relying upon delta cell reprogramming.

Regeneration of insulin-producing β-cells is an alternative avenue to manage diabetes, and it is crucial to unravel this process in vivo during physiological responses to the lack of β-cells. Here, we aimed to characterize how hepatocytes can contribute to β-cell regeneration, either directly or indirectly via secreted proteins or metabolites, in a zebrafish model of β-cell loss. Using lineage tracing, we show that hepatocytes do not directly convert into β-cells even under extreme β-cell ablation conditions. A transcriptomic analysis of isolated hepatocytes after β-cell ablation displayed altered lipid- and glucose-related processes. Based on the transcriptomics, we performed a genetic screen that uncovers a potential role of the molybdenum cofactor (Moco) biosynthetic pathway in β-cell regeneration and glucose metabolism in zebrafish. Consistently, molybdenum cofactor synthesis 2 (Mocs2) haploinsufficiency in mice indicated dysregulated glucose metabolism and liver function. Together, our study sheds light on the liver-pancreas crosstalk and suggests that the molybdenum cofactor biosynthesis pathway should be further studied in relation to glucose metabolism and diabetes.

The pandemic scale of diabetes mellitus is alarming, its complications remain devastating, and current treatments still pose a major burden on those affected and on the healthcare system as a whole. As the disease emanates from the destruction or dysfunction of insulin-producing pancreatic β-cells, a real cure requires their restoration and protection. An attractive strategy is to regenerate β-cells directly within the pancreas; however, while several approaches for β-cell regeneration have been proposed in the past, clinical translation has proven challenging. This review scrutinizes recent findings in β-cell regeneration and discusses their potential clinical implementation. Hereby, we aim to delineate a path for innovative, targeted therapies to help shift from 'caring for' to 'curing' diabetes.

Following the near-total depletion of pancreatic beta-cells with streptozotocin (STZ), a partial recovery of beta-cell mass (BCM) can occur, in part due to the alpha- to beta-cell transdifferentiation with an intermediary insulin/glucagon bi-hormonal cell phenotype. However, human type 2 diabetes typically involves only a partial reduction in BCM and it is not known if recovery after therapeutic intervention involves islet cell transdifferentiation, or how this varies with age. Here, we used transgenic mouse models to examine if islet cell transdifferentiation contributes to BCM recovery following only a partial depletion of BCM. Cell lineage tracing was employed using Glucagon-Cre/yellow fluorescent protein (YFP) transgenic mice treated with STZ (25 mg/kg-neonates; 70 mg/kg-adults) or vehicle alone on 3 consecutive days. Mice were euthanized 2-30 days later with a prior glucose tolerance test on day 30, and immunofluorescence histology performed on the pancreata. Beta-cell abundance was reduced by 30-40% two days post STZ in both neonates and adults, and subsequently partially recovered in adult but not neonatal mice. Glucose tolerance recovered in adult females, but not in males or neonates. Bi-hormonal cell abundance increased 2-3-fold in STZ-treated mice vs. controls in both neonates and adults, as did transdifferentiated cells expressing insulin and the YFP lineage tag, but not glucagon. Transdifferentiated cell presence was an order of magnitude lower than that of bi-hormonal cells. We conclude that alpha- to beta-cell transdifferentiation occurs in mice following only a moderate depletion in BCM, and that this was accompanied by a partial recovery of BCM in adults.

Pancreatic neuroendocrine tumors (NETs) are rare well-differentiated neoplasms with limited therapeutic options and unknown cells of origin. The current classification of pancreatic neuroendocrine tumors is based on proliferative grading, and guides therapeutic strategies, however, tumors within grades exhibit profound heterogeneity in clinical manifestation and outcome. Manifold studies have highlighted intra-patient differences in tumors at the genetic and transcriptomic levels. Molecular classification might become an alternative or complementary basis for treatment decisions and reflect tumor biology, actionable cellular processes. Here, we provide a comprehensive review of genomic, transcriptomic, proteomic and epigenomic studies of pancreatic NETs to elucidate patterns shared between proposed subtypes that could form a foundation for new classification. We denote four NET subtypes with distinct molecular features, which were consistently reproduced using various omics technologies.

Stimulation of pancreatic beta cell regeneration could be a therapeutic lead to treat diabetes. Unlike humans, the zebrafish can efficiently regenerate beta cells, notably from ductal pancreatic progenitors. To gain insight into the molecular pathways involved in this process, we established the transcriptomic profile of the ductal cells after beta cell ablation in the adult zebrafish. These data highlighted the protein phosphatase calcineurin (CaN) as a new potential modulator of beta cell regeneration. We showed that CaN overexpression abolished the regenerative response, leading to glycemia dysregulation. On the opposite, CaN inhibition increased ductal cell proliferation and subsequent beta cell regeneration. Interestingly, the enhanced proliferation of the progenitors was paradoxically coupled with their exhaustion. This suggests that the proliferating progenitors are next entering in differentiation. CaN appears as a guardian which prevents an excessive progenitor proliferation to preserve the pool of progenitors. Altogether, our findings reveal CaN as a key player in the balance between proliferation and differentiation to enable a proper beta cell regeneration.

The zebrafish has become an outstanding model for studying organ development and tissue regeneration, which is prominently leveraged for studies of pancreatic development, insulin-producing β-cells, and diabetes. Although studied for more than two decades, many aspects remain elusive and it has only recently been possible to investigate these due to technical advances in transcriptomics, chemical-genetics, genome editing, drug screening, and in vivo imaging. Here, we review recent findings on zebrafish pancreas development, β-cell regeneration, and how zebrafish can be used to provide novel insights into gene functions, disease mechanisms, and therapeutic targets in diabetes, inspiring further use of zebrafish for the development of novel therapies for diabetes.

Metazoan organisms are heterocellular systems composed of hundreds of different cell types, which arise from an isogenic genome through differentiation. Cellular 'plasticity' further enables cells to alter their fate in response to exogenous cues and is involved in a variety of processes, such as wound healing, infection, and cancer. Recent advances in cellular model systems, high-dimensional single-cell technologies, and lineage tracing have sparked a renaissance in plasticity research. Here, we discuss the definition of cell plasticity, evaluate state-of-the-art model systems and techniques to study cell-fate dynamics, and explore the application of single-cell technologies to obtain functional insights into cell plasticity in healthy and diseased tissues. The integration of advanced biomimetic model systems, single-cell technologies, and high-throughput perturbation studies is enabling a new era of research into non-genetic plasticity in metazoan systems.

Diabetes mellitus is induced by quantitative and qualitative decline in pancreatic β cells. Although its radical therapy has not yet been established, β cell regeneration is a promising option. We investigate here two mouse models of β cell regeneration induced after ∼80% reduction in β cell number: Cre/loxP-mediated β cell ablation and partial pancreatectomy. Cre/loxP-mediated, mosaic-pattern of β cell ablation by diphtheria toxin (DT) prompted rapid β cell replenishment through repeated proliferation of rare, highly proliferative DT receptor-negative β cells along with increase in Hes1, Neurog3, Ascl1, and Aldh1a3 (immature/dedifferentiated β cell markers) and decrease in Mafa (a mature β cell marker) in the islets. In contrast, pancreatectomy also prompted active proliferation, but with no change in these immature/dedifferentiated or mature β cell markers. Our findings demonstrate that the mode of β cell regeneration differs between Cre/loxP-mediated β cell ablation and surgical β cell reduction, and the former involves β cell dedifferentiation followed by active repetitive cell proliferation of a small population of β cells.

Regeneration is a heroic biological process that restores tissue architecture and function in the face of day-to-day cell loss or the aftershock of injury. Capacities and mechanisms for regeneration can vary widely among species, organs, and injury contexts. Here, we describe "hallmarks" of regeneration found in diverse settings of the animal kingdom, including activation of a cell source, initiation of regenerative programs in the source, interplay with supporting cell types, and control of tissue size and function. We discuss these hallmarks with an eye toward major challenges and applications of regenerative biology.

Insulin-producing β-cells in pancreatic islets are regulated by systemic cues and, locally, by adjacent islet hormone-producing 'non-β-cells' (namely α-cells, δ-cells and γ-cells). Yet whether the non-β-cells are required for accurate insulin secretion is unclear. Here, we studied mice in which adult islets are exclusively composed of β-cells and human pseudoislets containing only primary β-cells. Mice lacking non-β-cells had optimal blood glucose regulation, enhanced glucose tolerance, insulin sensitivity and restricted body weight gain under a high-fat diet. The insulin secretion dynamics in islets composed of only β-cells was comparable to that in intact islets. Similarly, human β-cell pseudoislets retained the glucose-regulated mitochondrial respiration, insulin secretion and exendin-4 responses of entire islets. The findings indicate that non-β-cells are dispensable for blood glucose homeostasis and β-cell function. These results support efforts aimed at developing diabetes treatments by generating β-like clusters devoid of non-β-cells, such as from pluripotent stem cells differentiated in vitro or by reprograming non-β-cells into insulin producers in situ.

Dysregulated glucagon secretion and inadequate functional beta cell mass are hallmark features of diabetes. While glucagon receptor (GCGR) antagonism ameliorates hyperglycemia and elicits beta cell regeneration in pre-clinical models of diabetes, it also promotes alpha and delta cell hyperplasia. We sought to investigate the mechanism by which loss of glucagon action impacts pancreatic islet non-alpha cells, and the relevance of these observations in a human islet context.

We used zebrafish, rodents, and transplanted human islets comprising six different models of interrupted glucagon signaling to examine their impact on delta and beta cell proliferation and mass. We also used models with global deficiency of the cationic amino acid transporter, SLC7A2, and mTORC1 inhibition via rapamycin, to determine whether amino acid-dependent nutrient sensing was required for islet non-alpha cell growth.

Inhibition of glucagon signaling stimulated delta cell proliferation in mouse and transplanted human islets, and in mouse islets. This was rapamycin-sensitive and required SLC7A2. Likewise, gcgr deficiency augmented beta cell proliferation via SLC7A2- and mTORC1-dependent mechanisms in zebrafish and promoted cell cycle engagement in rodent beta cells but was insufficient to drive a significant increase in beta cell mass in mice.

Our findings demonstrate that interruption of glucagon signaling augments islet non-alpha cell proliferation in zebrafish, rodents, and transplanted human islets in a manner requiring SLC7A2 and mTORC1 activation. An increase in delta cell mass may be leveraged for future beta cell regeneration therapies relying upon delta cell reprogramming.

Animal longevity is a function of global vital organ functionality and, consequently, a complex polygenic trait. Yet, monogenic regulators controlling overall or organ-specific ageing exist, owing their conservation to their function in growth and development. Here, by using pathway analysis combined with wet-biology methods on several dynamic timelines, we identified Hnf1a as a novel master regulator of the maturation and ageing in the adult pancreatic islet during the first year of life. Conditional transgenic mice bearing suboptimal levels of this transcription factor in the pancreatic islets displayed age-dependent changes, with a profile echoing precocious maturation. Additionally, the comparative pathway analysis revealed a link between Hnf1a age-dependent regulation and immune signaling, which was confirmed in the ageing timeline of an overly immunodeficient mouse model. Last, the global proteome analysis of human islets spanning three decades of life largely backed the age-specific regulation observed in mice. Collectively, our results suggest a novel role of Hnf1a as a monogenic regulator of the maturation and ageing process in the pancreatic islet via a direct or indirect regulatory loop with immune signaling.

Diabetes Mellitus (DM), a chronic metabolic disease characterized by elevated blood glucose, is caused by various degrees of insulin resistance and dysfunctional insulin secretion, resulting in hyperglycemia. The loss and failure of functional β-cells are key mechanisms resulting in type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM).

Elucidating the underlying mechanisms of β-cell failure, and exploring approaches for β-cell neogenesis to reverse β-cell dysfunction may provide novel strategies for DM therapy.

Emerging studies reveal that genetic susceptibility, endoplasmic reticulum (ER) stress, oxidative stress, islet inflammation, and protein modification linked to multiple signaling pathways contribute to DM pathogenesis. Over the past few years, replenishing functional β-cell by β-cell neogenesis to restore the number and function of pancreatic β-cells has remarkably exhibited a promising therapeutic approach for DM therapy. In this review, we provide a comprehensive overview of the underlying mechanisms of β-cell failure in DM, highlight the effective approaches for β-cell neogenesis, as well as discuss the current clinical and preclinical agents research advances of β-cell neogenesis. Insights into the challenges of translating β-cell neogenesis into clinical application for DM treatment are also offered.

The pancreatic islet is the functional and structural unit of the pancreatic endocrine portion. Islet remodeling occurs in both normal development and pathogenesis of type 1 (T1D) and type 2 diabetes (T2D). However, accurately quantifying changes in islet cellular makeup and hormone expressions poses significant challenges due to large intra- and inter-donor heterogeneity and the limited scalability of traditional methods such as immunostaining. The cytometry by time-of-flight (CyTOF) technology enables simultaneous quantification of more than 30 protein markers at single-cell resolution in a high-throughput fashion. Moreover, with distinct DNA and viability markers, single live cells can be explicitly selected in CyTOF. Here, leveraging the CyTOF data generated by the Human Pancreas Analysis Program, we characterized more than 12 million islet cells from 71 donors. Our data revealed continued age-related changes in islet endocrine cell compositions, but the maturity of endocrine cells is reached by 3 years of age. We also observed significant changes in beta cell numbers and key protein expressions, along with a significant increase in bihormonal cells in T1D donors. In contrast, T2D donors exhibited minimal islet remodeling events. Our data shine a light on the islet dynamics during development and diabetes pathogenesis and suggest divergent pathogenesis processes of T1D and T2D. Our comprehensive approach not only elucidates islet plasticity but also establishes a foundation for integrated CyTOF analysis in islet biology and beyond.

The β-cells within the pancreas play a pivotal role in insulin production and secretion, responding to fluctuations in blood glucose levels. However, factors like obesity, dietary habits, and prolonged insulin resistance can compromise β-cell function, contributing to the development of Type 2 Diabetes (T2D). A critical aspect of this dysfunction involves β-cell dedifferentiation and transdifferentiation, wherein these cells lose their specialized characteristics and adopt different identities, notably transitioning towards progenitor or other pancreatic cell types like α-cells. This process significantly contributes to β-cell malfunction and the progression of T2D, often surpassing the impact of outright β-cell loss. Alterations in the expressions of specific genes and transcription factors unique to β-cells, along with epigenetic modifications and environmental factors such as inflammation, oxidative stress, and mitochondrial dysfunction, underpin the occurrence of β-cell dedifferentiation and the onset of T2D. Recent research underscores the potential therapeutic value for targeting β-cell dedifferentiation to manage T2D effectively. In this review, we aim to dissect the intricate mechanisms governing β-cell dedifferentiation and explore the therapeutic avenues stemming from these insights.

Regeneration, the ability to replace injured tissues and organs, is a phenomenon commonly associated with lower vertebrates but is also observed in mammals, in specific tissues. In this study, we investigated the regenerative potential of pancreatic islets following moderate beta-cell loss in mice. Using a rapid model of moderate ablation, we observed a compensatory response characterized by transient inflammation and proliferation signatures, ultimately leading to the recovery of beta-cell identity and function. Interestingly, this proliferative response occurred independently of inflammation, as demonstrated in ablated immunodeficient mice. Furthermore, exposure to high-fat diet stimulated beta-cell proliferation but negatively impacted beta-cell function. In contrast, an equivalent slower ablation model revealed a delayed but similar proliferative response, suggesting proliferation as a common regenerative response. However, high-fat diet failed to promote proliferation in this model, indicating a differential response to metabolic stressors. Overall, our findings shed light on the complex interplay between beta-cell loss, inflammation, and stress in modulating pancreatic islet regeneration. Understanding these mechanisms could pave the way for novel therapeutic strategies based on beta-cell proliferation.

Sleeve gastrectomy (SG) is one of the most commonly performed bariatric surgeries. SG treats type 2 diabetes mellitus better than several drugs. The mechanisms that underlie this phenomenon are not clear. This study proposed that somatostatin (SST) isoforms SST-14 and SST-28 are key in the carbohydrate after SG.

Surgeries were performed on 3 groups of Wistar rats: the fasting, surgery control, and SG groups. Plasma levels of glucose, insulin, SST-14, and SST-28 were measured at 2 survival periods after surgery. Islet SST receptor (SSTR) and cell populations were studied. We performed a pasireotide (SST-28 analogue) infusion assay in another group of rats to confirm the influence of SST-28 plasma levels on the delta-cell population.

This study found an elevation in the insulin response after SG in animals but a decrease in the insulin response over the long term with a loss of beta-cell mass. An increase in duodenal SST-28-producing cells in the duodenum and a loss of pancreatic SST-14-producing cells were observed after SG in animals but not in controls. The expression of SSTR type 5 in delta-cell populations from each group and the ability of the pasireotide infusion assay to decrease the delta-cell population indicated the effect of SST-28 plasma levels on delta-cell maintenance.

After SG initiates a compensatory response in the duodenum, beta-cell mass is depleted after loss of the brake that regulates SST-14 at the paracrine level in a nonobese, normoglycemic rat model. This was an experimental model, with no clinical translation to the human clinic, with a preliminary importance regarding new pathophysiologic perspectives or pathways.

Mesenchymal stem cells (MSCs) are known for their immunosuppressive properties. Based on the demonstrated anti-inflammatory effect of mouse MSCs from hair follicles (moMSCORS) in a murine wound closure model, this study evaluates their potential for preventing type 1 diabetes (T1D) in C57BL/6 mice. T1D was induced in C57BL/6 mice by repeated low doses of streptozotocin. moMSCORS were injected intravenously on weekly basis. moMSCORS reduced T1D incidence, the insulitis stage, and preserved insulin production in treated animals. moMSCORS primarily exerted immunomodulatory effects by inhibiting CD4+ T cell proliferation and activation. Ex vivo analysis indicated that moMSCORS modified the cellular immune profile within pancreatic lymph nodes and pancreatic infiltrates by reducing the numbers of M1 pro-inflammatory macrophages and T helper 17 cells and upscaling the immunosuppressive T regulatory cells. The proportion of pathogenic insulin-specific CD4+ T cells was down-scaled in the lymph nodes, likely via soluble factors. The moMSCORS detected in the pancreatic infiltrates of treated mice presumably exerted the observed suppressive effect on CD4+ through direct contact. moMSCORS alleviated T1D symptoms in the mouse, qualifying as a candidate for therapeutic products by multiple advantages: non-invasive sampling by epilation, easy access, permanent availability, scalability, and benefits of auto-transplantation.

Plasticity among cell lineages is a fundamental, but poorly understood, property of regenerative tissues. In the gut tube, the small intestine absorbs nutrients, whereas the colon absorbs electrolytes. In a striking display of inherent plasticity, adult colonic mucosa lacking the chromatin factor SATB2 is converted to small intestine. Using proteomics and CRISPR-Cas9 screening, we identify MTA2 as a crucial component of the molecular machinery that, together with SATB2, restrains colonic plasticity. MTA2 loss in the adult mouse colon activated lipid absorptive genes and functional lipid uptake. Mechanistically, MTA2 co-occupies DNA with HNF4A, an activating pan-intestinal transcription factor (TF), on colonic chromatin. MTA2 loss leads to HNF4A release from colonic chromatin, and accumulation on small intestinal chromatin. SATB2 similarly restrains colonic plasticity through an HNF4A-dependent mechanism. Our study provides a generalizable model of lineage plasticity in which broadly-expressed TFs are retained on tissue-specific enhancers to maintain cell identity and prevent activation of alternative lineages, and their release unleashes plasticity.

Islets of Langerhans are anatomically dispersed within the pancreas and exhibit regulatory coordination between islets in response to nutritional and inflammatory stimuli. However, within individual islets, there is also multi-faceted coordination of function between individual beta-cells, and between beta-cells and other endocrine and vascular cell types. This is mediated partly through circulatory feedback of the major secreted hormones, insulin and glucagon, but also by autocrine and paracrine actions within the islet by a range of other secreted products, including somatostatin, urocortin 3, serotonin, glucagon-like peptide-1, acetylcholine, and ghrelin. Their availability can be modulated within the islet by pericyte-mediated regulation of microvascular blood flow. Within the islet, both endocrine progenitor cells and the ability of endocrine cells to trans-differentiate between phenotypes can alter endocrine cell mass to adapt to changed metabolic circumstances, regulated by the within-islet trophic environment. Optimal islet function is precariously balanced due to the high metabolic rate required by beta-cells to synthesize and secrete insulin, and they are susceptible to oxidative and endoplasmic reticular stress in the face of high metabolic demand. Resulting changes in paracrine dynamics within the islets can contribute to the emergence of Types 1, 2 and gestational diabetes.

Pancreatic β-cell failure due to a reduction in function and mass has been defined as a primary contributor to the progression of type 2 diabetes (T2D). Reserving insulin-producing β-cells and hence restoring insulin production are gaining attention in translational diabetes research, and β-cell replenishment has been the main focus for diabetes treatment. Significant findings in β-cell proliferation, transdifferentiation, pluripotent stem cell differentiation, and associated small molecules have served as promising strategies to regenerate β-cells. In this review, we summarize current knowledge on the mechanisms implicated in β-cell dynamic processes under physiological and diabetic conditions, in which genetic factors, age-related alterations, metabolic stresses, and compromised identity are critical factors contributing to β-cell failure in T2D. The article also focuses on recent advances in therapeutic strategies for diabetes treatment by promoting β-cell proliferation, inducing non-β-cell transdifferentiation, and reprograming stem cell differentiation. Although a significant challenge remains for each of these strategies, the recognition of the mechanisms responsible for β-cell development and mature endocrine cell plasticity and remarkable advances in the generation of exogenous β-cells from stem cells and single-cell studies pave the way for developing potential approaches to cure diabetes.

The progression of type II diabetes (T2D) is characterized by a complex and highly variable loss of beta-cell mass, resulting in impaired insulin secretion. Many T2D drug discovery efforts aimed at discovering molecules that can protect or restore beta-cell mass and function have been developed using limited beta-cell lines and primary rodent/human pancreatic islets. Various high-throughput screening methods have been used in the context of drug discovery, including luciferase-based reporter assays, glucose-stimulated insulin secretion, and high-content screening. In this context, a cornerstone of small molecule discovery has been the use of immortalized rodent beta-cell lines. Although insightful, this usage has led to a more comprehensive understanding of rodent beta-cell proliferation pathways rather than their human counterparts. Advantages gained in enhanced physiological relevance are offered by three-dimensional (3D) primary islets and pseudoislets in contrast to monolayer cultures, but these approaches have been limited to use in low-throughput experiments. Emerging methods, such as high-throughput 3D islet imaging coupled with machine learning, aim to increase the feasibility of integrating 3D microtissue structures into high-throughput screening. This review explores the current methods used in high-throughput screening for small molecule modulators of beta-cell mass and function, a potentially pivotal strategy for diabetes drug discovery.

β-Cell replacement by in situ reprogramming of non-β-cells is a promising diabetes therapy. Following the observation that near-total β-cell ablation in adult mice triggers the reprogramming of pancreatic α-, δ-, and γ-cells into insulin (INS)-producing cells, recent studies are delving deep into the mechanisms controlling adult α-cell identity. Systematic analyses of the α-cell transcriptome and epigenome have started to pinpoint features that could be crucial for maintaining α-cell identity. Using different transgenic and chemical approaches, significant advances have been made in reprogramming α-cells in vivo into INS-secreting cells in mice. The recent reprogramming of human α-cells in vitro is an important step forward that must now be complemented with a comprehensive molecular dissection of the mechanisms controlling α-cell identity.

During the pathogenesis of type 1 diabetes (T1D) and type 2 diabetes (T2D), pancreatic islets, especially the β cells, face significant challenges. These insulin-producing cells adopt a regeneration strategy to compensate for the shortage of insulin, but the exact mechanism needs to be defined. High-fat diet (HFD) and streptozotocin (STZ) treatment are well-established models to study islet damage in T2D and T1D respectively. Therefore, we applied these two diabetic mouse models, triggered at different ages, to pursue the cell fate transition of islet β cells. Cre-LoxP systems were used to generate islet cell type-specific (α, β, or δ) green fluorescent protein (GFP)-labeled mice for genetic lineage tracing, thereinto β-cell GFP-labeled mice were tamoxifen induced. Single-cell RNA sequencing (scRNA-seq) was used to investigate the evolutionary trajectories and molecular mechanisms of the GFP-labeled β cells in STZ-treated mice. STZ-induced diabetes caused extensive dedifferentiation of β cells and some of which transdifferentiated into a or δ cells in both youth- and adulthood-initiated mice while this phenomenon was barely observed in HFD models. β cells in HFD mice were expanded via self-replication rather than via transdifferentiation from α or δ cells, in contrast, α or δ cells were induced to transdifferentiate into β cells in STZ-treated mice (both youth- and adulthood-initiated). In addition to the re-dedifferentiation of β cells, it is also highly likely that these "α or δ" cells transdifferentiated from pre-existing β cells could also re-trans-differentiate into insulin-producing β cells and be beneficial to islet recovery. The analysis of ScRNA-seq revealed that several pathways including mitochondrial function, chromatin modification, and remodeling are crucial in the dynamic transition of β cells. Our findings shed light on how islet β cells overcome the deficit of insulin and the molecular mechanism of islet recovery in T1D and T2D pathogenesis.

The consequences of mutations in genes associated with monogenic forms of diabetes on human pancreas development cannot be studied in a time-resolved fashion in vivo. More specifically, if recessive mutations in the insulin gene influence human pancreatic endocrine lineage formation is still an unresolved question.

To model the extremely reduced insulin levels in patients with recessive insulin gene mutations, we generated a novel knock-in H2B-Cherry reporter human induced pluripotent stem cell (iPSC) line expressing no insulin upon differentiation to stem cell-derived (SC-) β cells in vitro. Differentiation of iPSCs into the pancreatic and endocrine lineage, combined with immunostaining, Western blotting and proteomics analysis phenotypically characterized the insulin gene deficiency in SC-islets. Furthermore, we leveraged FACS analysis and confocal microscopy to explore the impact of insulin shortage on human endocrine cell induction, composition, differentiation and proliferation.

Interestingly, insulin-deficient SC-islets exhibited low insulin receptor (IR) signaling when stimulated with glucose but displayed increased IR sensitivity upon treatment with exogenous insulin. Furthermore, insulin shortage did not alter neurogenin-3 (NGN3)-mediated endocrine lineage induction. Nevertheless, lack of insulin skewed the SC-islet cell composition with an increased number in SC-β cell formation at the expense of SC-α cells. Finally, insulin deficiency reduced the rate of SC-β cell proliferation but had no impact on the expansion of SC-α cells.

Using iPSC disease modelling, we provide first evidence of insulin function in human pancreatic endocrine lineage formation. These findings help to better understand the phenotypic impact of recessive insulin gene mutations during pancreas development and shed light on insulin gene function beside its physiological role in blood glucose regulation.