Illuminating the complete ß-cell mass of the human pancreas- signifying a new view on the islets of Langerhans.

Pancreatic islets of Langerhans play a pivotal role in regulating blood glucose homeostasis, but critical information regarding their mass, distribution and composition is lacking within a whole organ context. Here, we apply a 3D imaging pipeline to generate a complete account of the insulin-producing islets throughout the human pancreas at a microscopic resolution and within a maintained spatial 3D context. These data show that human islets are far more heterogenous than previously accounted for with regards to their size distribution and cellular make up. By deep tissue 3D imaging, this in-depth study demonstrates that 50% of the human insulin-expressing islets are virtually devoid of glucagon-producing α-cells, an observation with significant implications for both experimental and clinical research.

Repressing miR-23a promotes the transdifferentiation of pancreatic α cells to ß cells via negatively regulating the expression of SDF-1α.

Pancreatic ß-cell failure is a pathological feature in type 1 diabetes. One promising approach involves inducing transdifferentiation of related pancreatic cell types, specifically α cells that produce glucagon. The chemokine stromal cell-derived factor-1 alpha (SDF-1α) is implicated in pancreatic α-to-ß like cell transition. Here, the serum level of SDF-1α was lower in T1D with C-peptide loss, the miR-23a was negatively correlated with SDF-1α. We discovered that exosomal miR-23a, secreted from ß cells, functionally downregulates the expression of SDF-1α, leading to increased Pax4 expression and decreased Arx expression in vivo. Adenovirus-vectored miR-23a sponge and mimic were constructed to further explored the miR-23a on pancreatic α-to-ß like cell transition in vitro, which yielded results consistent with our cell-based assays. Suppression of miR-23a upregulated insulin level and downregulated glucagon level in STZ-induced diabetes mice models, effectively promoting α-to-ß like cell transition. Our findings highlight miR-23a as a new therapeutic target for regenerating pancreatic ß cells from α cells.

GLP1R and GIPR expression and signaling in pancreatic alpha cells, beta cells and delta cells.

Glucagon-like peptide-1 receptor (GLP1R) and glucose-dependent insulinotropic polypeptide receptor (GIPR) are transmembrane receptors involved in insulin, glucagon and somatostatin secretion from the pancreatic islet. Therapeutic targeting of GLP1R and GIPR restores blood glucose levels in part by influencing beta cell, alpha cell and delta cell function. Despite the importance of the incretin-mimetics for diabetes therapy, our understanding of GLP1R and GIPR expression patterns and signaling within the islet remain incomplete. Here, we present the evidence for GLP1R and GIPR expression in the major islet cell types, before addressing signaling pathway(s) engaged, as well as their influence on cell survival and function. While GLP1R is largely a beta cell-specific marker within the islet, GIPR is expressed in alpha cells, beta cells, and (possibly) delta cells. GLP1R and GIPR engage Gs-coupled pathways in most settings, although the exact outcome on hormone release depends on paracrine communication and promiscuous signaling. Biased agonism away from beta-arrestin is an emerging concept for improving therapeutic efficacy, and is also relevant for GLP1R/GIPR dual agonism. Lastly, dual agonists exert multiple effects on islet function through GIPR > GLP1R imbalance, increased GLP1R surface expression and cAMP signaling, as well as beneficial alpha cell-beta cell-delta cell crosstalk.

DNA Methylation-Based Assessment of Cell Composition in Human Pancreas and Islets.

Assessment of pancreas cell type composition is crucial to the understanding of the genesis of diabetes. Current approaches use immunodetection of protein markers, for example, insulin as a marker of ß-cells. A major limitation of these methods is that protein content varies in physiological and pathological conditions, complicating the extrapolation to actual cell number. Here, we demonstrate the use of cell type-specific DNA methylation markers for determining the fraction of specific cell types in human islet and pancreas specimens. We identified genomic loci that are uniquely demethylated in specific pancreatic cell types and applied targeted PCR to assess the methylation status of these loci in tissue samples, enabling inference of cell type composition. In islet preparations, normalization of insulin secretion to ß-cell DNA revealed similar ß-cell function in pre-type 1 diabetes (T1D), T1D, and type 2 diabetes (T2D), which was significantly lower than in donors without diabetes. In histological pancreas specimens from recent-onset T1D, this assay showed ß-cell fraction within the normal range, suggesting a significant contribution of ß-cell dysfunction. In T2D pancreata, we observed increased α-cell fraction and normal ß-cell fraction. Methylation-based analysis provides an accurate molecular alternative to immune detection of cell types in the human pancreas, with utility in the interpretation of insulin secretion assays and the assessment of pancreas cell composition in health and disease.

Novel time-resolved reporter mouse reveals spatial and transcriptional heterogeneity during alpha cell differentiation.

AIMS/HYPOTHESIS: Glucagon-expressing pancreatic alpha cells have attracted much attention for their plasticity to transdifferentiate into insulin-producing beta cells; however, it remains unclear precisely when, and from where, alpha cells emerge and what regulates alpha cell fate. We therefore explored the spatial and transcriptional heterogeneity of alpha cell differentiation using a novel time-resolved reporter system. METHODS: We established the mouse model, 'Gcg-Timer', in which newly generated alpha cells can be distinguished from more-differentiated cells by their fluorescence. Fluorescence imaging and transcriptome analysis were performed with Gcg-Timer mice during the embryonic and postnatal stages. RESULTS: Fluorescence imaging and flow cytometry demonstrated that green fluorescence-dominant cells were present in Gcg-Timer mice at the embryonic and neonatal stages but not after 1 week of age, suggesting that alpha cell neogenesis occurs during embryogenesis and early neonatal stages under physiological conditions. Transcriptome analysis of Gcg-Timer embryos revealed that the mRNAs related to angiogenesis were enriched in newly generated alpha cells. Histological analysis revealed that some alpha cells arise close to the pancreatic ducts, whereas the others arise away from the ducts and adjacent to the blood vessels. Notably, when the glucagon signal was suppressed by genetic ablation or by chemicals, such as neutralising glucagon antibody, green-dominant cells emerged again in adult mice. CONCLUSIONS/INTERPRETATION: Novel time-resolved analysis with Gcg-Timer reporter mice uncovered spatiotemporal features of alpha cell neogenesis that will enhance our understanding of cellular identity and plasticity within the islets. DATA AVAILABILITY: Raw and processed RNA sequencing data for this study has been deposited in the Gene Expression Omnibus under accession number GSE229090.

A bright future for glucagon and alpha cell biology.

Long lagging behind insulin, glucagon research has caught up in large part, thanks to technological breakthroughs. Here we review how the field was propelled by the development of novel techniques and approaches. The glucagon radioimmunoassay and islet isolation are methods that now seem trivial, but for decades they were crucial in defining the biology of the pancreatic alpha cell and the role of glucagon in glucose homeostasis. More recently, mouse models have become the main workhorse of this research effort, if not of biomedical research in general. The mouse model allowed detailed mechanistic studies that are revealing alpha cell functions beyond its canonical glucoregulatory role. A recent profusion of gene expression and transcription regulation studies is providing new vistas into what constitutes alpha cell identity. In particular, the combination of transcriptomic techniques with functional recordings promises to move molecular guesswork into real-time physiology. The challenge right now is not to get enamored with these powerful techniques and to make sure that the research continues to be transformative and paradigm shifting. We should imagine a future in which the biology of the alpha cell will be studied at single-cell resolution, non-invasively, and in real time in the human body.

Conflicting Views About Interactions Between Pancreatic α-Cells and ß-Cells.

In type 1 diabetes, the reduced glucagon response to insulin-induced hypoglycemia has been used to argue that ß-cell secretion of insulin is required for the full glucagon counterregulatory response. For years, the concept has been that insulin from the ß-cell core flows downstream to suppress glucagon secretion from the α-cells in the islet mantle. This core-mantle relationship has been supported by perfused pancreas studies that show marked increases in glucagon secretion when insulin was neutralized with antisera. Additional support comes from a growing number of studies focused on vascular anatomy and blood flow. However, in recent years this core-mantle view has generated less interest than the argument that optimal insulin secretion is due to paracrine release of glucagon from α-cells stimulating adjacent ß-cells. This mechanism has been evaluated by knockout of ß-cell receptors and impairment of α-cell function by inhibition of Gi designer receptors exclusively activated by designer drugs. Other studies that support this mechanism have been obtained by pharmacological blocking of glucagon-like peptide 1 receptor in humans. While glucagon has potent effects on ß-cells, there are concerns with the suggested paracrine mechanism, since some of the supporting data are from isolated islets. The study of islets in static incubation or perifusion systems can be informative, but the normal paracrine relationships are disrupted by the isolation process. While this complicates interpretation of data, arguments supporting paracrine interactions between α-cells and ß-cells have growing appeal. We discuss these conflicting views of the relationship between pancreatic α-cells and ß-cells and seek to understand how communication depends on blood flow and/or paracrine mechanisms.

PAX4 loss of function increases diabetes risk by altering human pancreatic endocrine cell development.

The coding variant (p.Arg192His) in the transcription factor PAX4 is associated with an altered risk for type 2 diabetes (T2D) in East Asian populations. In mice, Pax4 is essential for beta cell formation but its role on human beta cell development and/or function is unknown. Participants carrying the PAX4 p.His192 allele exhibited decreased pancreatic beta cell function compared to homozygotes for the p.192Arg allele in a cross-sectional study in which we carried out an intravenous glucose tolerance test and an oral glucose tolerance test. In a pedigree of a patient with young onset diabetes, several members carry a newly identified p.Tyr186X allele. In the human beta cell model, EndoC-ßH1, PAX4 knockdown led to impaired insulin secretion, reduced total insulin content, and altered hormone gene expression. Deletion of PAX4 in human induced pluripotent stem cell (hiPSC)-derived islet-like cells resulted in derepression of alpha cell gene expression. In vitro differentiation of hiPSCs carrying PAX4 p.His192 and p.X186 risk alleles exhibited increased polyhormonal endocrine cell formation and reduced insulin content that can be reversed with gene correction. Together, we demonstrate the role of PAX4 in human endocrine cell development, beta cell function, and its contribution to T2D-risk.

Fluorescein-based sensors to purify human α-cells for functional and transcriptomic analyses.

Pancreatic α-cells secrete glucagon, an insulin counter-regulatory peptide hormone critical for the maintenance of glucose homeostasis. Investigation of the function of human α-cells remains a challenge due to the lack of cost-effective purification methods to isolate high-quality α-cells from islets. Here, we use the reaction-based probe diacetylated Zinpyr1 (DA-ZP1) to introduce a novel and simple method for enriching live α-cells from dissociated human islet cells with ~95% purity. The α-cells, confirmed by sorting and immunostaining for glucagon, were cultured up to 10 days to form α-pseudoislets. The α-pseudoislets could be maintained in culture without significant loss of viability, and responded to glucose challenge by secreting appropriate levels of glucagon. RNA-sequencing analyses (RNA-seq) revealed that expression levels of key α-cell identity genes were sustained in culture while some of the genes such as DLK1, GSN, SMIM24 were altered in α-pseudoislets in a time-dependent manner. In conclusion, we report a method to sort human primary α-cells with high purity that can be used for downstream analyses such as functional and transcriptional studies.

Alpha cell receptor for advanced glycation end products associate with glucagon expression in type 1 diabetes.

Hypoglycemia in type 1 diabetes associates with changes in the pancreatic islet α cells, where the receptor for advanced glycation end products (RAGE) is highly expressed. This study compared islet RAGE expression in donors without diabetes, those at risk of, and those with type 1 diabetes. Laser-dissected islets were subject to RNA bioinformatics and adjacent pancreatic tissue were assessed by confocal microscopy. We found that islets from type 1 diabetes donors had differential expression of the RAGE gene (AGER) and its correlated genes, based on glucagon expression. Random forest machine learning revealed that AGER was the most important predictor for islet glucagon levels. Conversely, a generalized linear model identified that glucagon expression could be predicted by expression of RAGE signaling molecules, its ligands and enzymes that create or clear RAGE ligands. Confocal imaging co-localized RAGE, its ligands and signaling molecules to the α cells. Half of the type 1 diabetes cohort comprised of adolescents and a patient with history of hypoglycemia-all showed an inverse relationship between glucagon and RAGE. These data confirm an association between glucagon and islet RAGE, its ligands and signaling pathways in type 1 diabetes, which warrants functional investigation into a role for RAGE in hypoglycemia.

Oleanolic acid and moderate drinking increase the pancreatic GLP-1R expression of the ß-cell mass deficiency induced hyperglycemia.

Background: Oleanolic acid (OA) and moderate drinking have been reported to attenuate diabetes. However, the underlying mechanism of OA and moderate drinking alone or in combination on the islet ß-cell deficiency induced diabetes is not fully elucidated. Methods: Male Sprague Dawley (SD) rats were intraperitoneally injected with 55 mg/kg streptozotocin (STZ) to induce ß-cell deficiency. OA, 5% ethanol (EtOH), or a mixture of OA in 5% ethanol (OA+EtOH) were applied to three treatment groups of hyperglycemia rats for 6 weeks. Results: STZ caused the increase of fast blood glucose (FBG) level.OA and EtOH treatment alone or in combination decreased the STZ increased FBG level during the 6 weeks of treatment. In addition, OA treatment also significantly increased the ß-cell to total islet cell ratio. Both EtOH and OA+EtOH treatments promoted the increase of total islet cell number and α-cell to ß-cell ratio when compared to OA group. STZ induced hyperglycemia dramatically reduced the glucagon-like peptide-1 receptor (GLP-1R) positive cells in islets, all the three treatments significantly increased the pancreatic GLP-1R positive cell number. In the meantime, STZ induced hyperglycemia suppressed the insulin mRNA expression and boosted the glucagon mRNA expression. EtOH and OA+EtOH treatments increased the insulin mRNA expression, but none of the 3 treatments altered the elevated glucagon level. Conclusion: GLP-1R positive cell ratio in islets is crucial for the blood glucose level of diabetes. OA and 5% ethanol alone or in combination suppresses the blood glucose level of ß-cell deficiency induced diabetes by increasing islet GLP-1R expression.

Metabolic regulation of glucagon secretion.

Since the discovery of glucagon 100 years ago, the hormone and the pancreatic islet alpha cells that produce it have remained enigmatic relative to insulin-producing beta cells. Canonically, alpha cells have been described in the context of glucagon's role in glucose metabolism in liver, with glucose as the primary nutrient signal regulating alpha cell function. However, current data reveal a more holistic model of metabolic signalling, involving glucagon-regulated metabolism of multiple nutrients by the liver and other tissues, including amino acids and lipids, providing reciprocal feedback to regulate glucagon secretion and even alpha cell mass. Here we describe how various nutrients are sensed, transported and metabolised in alpha cells, providing an integrative model for the metabolic regulation of glucagon secretion and action. Importantly, we discuss where these nutrient-sensing pathways intersect to regulate alpha cell function and highlight key areas for future research.

Glucose Controls Glucagon Secretion by Regulating Fatty Acid Oxidation in Pancreatic α-Cells.

Whole-body glucose homeostasis is coordinated through secretion of glucagon and insulin from pancreatic islets. When glucose is low, glucagon is released from α-cells to stimulate hepatic glucose production. However, the mechanisms that regulate glucagon secretion from pancreatic α-cells remain unclear. Here we show that in α-cells, the interaction between fatty acid oxidation and glucose metabolism controls glucagon secretion. The glucose-dependent inhibition of glucagon secretion relies on pyruvate dehydrogenase and carnitine palmitoyl transferase 1a activity and lowering of mitochondrial fatty acid oxidation by increases in glucose. This results in reduced intracellular ATP and leads to membrane repolarization and inhibition of glucagon secretion. These findings provide a new framework for the metabolic regulation of the α-cell, where regulation of fatty acid oxidation by glucose accounts for the stimulation and inhibition of glucagon secretion. ARTICLE HIGHLIGHTS: It has become clear that dysregulation of glucagon secretion and α-cell function plays an important role in the development of diabetes, but we do not know how glucagon secretion is regulated. Here we asked whether glucose inhibits fatty acid oxidation in α-cells to regulate glucagon secretion. We found that fatty acid oxidation is required for the inhibitory effects of glucose on glucagon secretion through reductions in ATP. These findings provide a new framework for the regulation of glucagon secretion by glucose.

Blocking IL-6 signaling improves glucose tolerance via SLC39A5-mediated suppression of glucagon secretion.

BACKGROUND AND AIMS: Hyperinsulinemia, hyperglucagonemia, and low-grade inflammation are frequently presented in obesity and type 2 diabetes (T2D). The pathogenic regulation between hyperinsulinemia/insulin resistance (IR) and low-grade inflammation is well documented in the development of diabetes. However, the cross-talk of hyperglucagonemia with low-grade inflammation during diabetes progression is poorly understood. In this study, we investigated the regulatory role of proinflammatory cytokine interleukin-6 (IL-6) on glucagon secretion. METHODS: The correlations between inflammatory cytokines and glucagon or insulin were analyzed in rhesus monkeys and humans. IL-6 signaling was blocked by IL-6 receptor-neutralizing antibody tocilizumab in obese or T2D rhesus monkeys, glucose tolerance was evaluated by intravenous glucose tolerance test (IVGTT). Glucagon and insulin secretion were measured in isolated islets from wild-type mouse, primary pancreatic α-cells and non-α-cells sorted from GluCre-ROSA26EYFP (GYY) mice, in which the enhanced yellow fluorescent protein (EYFP) was expressed under the proglucagon promoter, by fluorescence-activated cell sorting (FACS). Particularly, glucagon secretion in α-TC1 cells treated with IL-6 was measured, and RNA sequencing was used to screen the mediator underlying IL-6-induced glucagon secretion. SLC39A5 was knocking-down or overexpressed in α-TC1 cells to determine its impact in glucagon secretion and cytosolic zinc density. Dual luciferase and chromatin Immunoprecipitation were applied to analyze the signal transducer and activator of transcription 3 (STAT3) in the regulation of SLC39A5 transcription. RESULTS: Plasma IL-6 correlate positively with plasma glucagon levels, but not insulin, in rhesus monkeys and humans. Tocilizumab treatment reduced plasma glucagon, blood glucose and HbA1c in spontaneously obese or T2D rhesus monkeys. Tocilizumab treatment also decreased glucagon levels during IVGTT, and improved glucose tolerance. Moreover, IL-6 significantly increased glucagon secretion in isolated islets, primary pancreatic α-cells and α-TC1 cells. Mechanistically, we found that IL-6-activated STAT3 downregulated the zinc transporter SLC39A5, which in turn reduced cytosolic zinc concentration and ATP-sensitive potassium channel activity and augmented glucagon secretion. CONCLUSIONS: This study demonstrates that IL-6 increases glucagon secretion via the downregulation of zinc transporter SLC39A5. This result revealed the molecular mechanism underlying the pathogenesis of hyperglucagonemia and a previously unidentified function of IL-6 in the pathophysiology of T2D, providing a potential new therapeutic strategy of targeting IL-6/glucagon to preventing or treating T2D.

Glucagon cell hyperplasia and neoplasia: a recently recognized endocrine receptor disease.

Glucagon cell hyperplasia and neoplasia (GCHN) is the name of an endocrine receptor disease, whose morphology was first described in 2006. Three years later, this rare disease was found to be to be caused by an inactivating mutation of the glucagon receptor (GCGR) gene. Functionally, the genetic defect mainly affects glucagon signaling in the liver with changes in the metabolism of glycogen, fatty acids and amino acids. Recent results of several studies in GCGR knockout mice suggested that elevated serum amino acid levels probably stimulate glucagon cell hyperplasia with subsequent transformation into glucagon cell neoplasia. This process leads over time to numerous small and some large pancreatic neuroendocrine tumors which are potentially malignant. Despite high glucagon serum levels, the patients develop no glucagonoma syndrome. In 2015, GCHN was identified as an autosomal recessive hereditary disorder.

Amino acids and the changing face of the α-cell.

Glucagon has long been defined by its glucogenic action and as a result α-cells have been characterised based largely on their interaction with glucose. Recent findings have challenged this preconception, bringing to the fore the significant role glucagon plays in amino acid breakdown and underlining the importance of amino acids in glucagon secretion. The challenge that remains is defining the mechanism that underlie these effects - understanding which amino acids are most important, how they act on the α-cell and how their actions integrate with other fuels such as glucose and fatty acids. This review will describe the current relationship between amino acids and glucagon and how we can use this knowledge to redefine the α-cell.

Comparative transcriptomic analysis reveals the underlying molecular mechanism in high-fat diet-induced islet dysfunction.

Obesity, characterized by accumulation of adipose, is usually accompanied by hyperlipidemia and abnormal glucose metabolism, which destroys the function and structure of islet ß cells. However, the exact mechanism of islet deterioration caused by obesity has not yet been fully elucidated. Here, we fed C57BL/6 mice with a high-fat diet (HFD) for 2 (2M group) and 6 months (6M group) to construct obesity mouse models. Then, RNA-based sequencing was used to identify the molecular mechanisms in HFD-induced islet dysfunction. Compared with the control diet, a total of 262 and 428 differentially expressed genes (DEGs) were identified from islets of the 2M and 6M groups, respectively. GO and KEGG enrichment analysis revealed that the DEGs up-regulated in both the 2M and 6M groups are mainly enriched in response to endoplasmic reticulum stress and the pancreatic secretion pathway. DEGs down-regulated in both the 2M and 6M groups are mainly enriched in the neuronal cell body and protein digestion and absorption pathway. Notably, along with the HFD feeding, mRNA expression of islet cell markers was significantly down-regulated, such as Ins1, Pdx1, MafA (ß cell), Gcg, Arx (α cell), Sst (δcell), and Ppy (PP cell). In contrast, mRNA expression of acinar cell markers was remarkably up-regulated, such as Amy1, Prss2, and Pnlip. Besides, a large number of collagen genes were down-regulated, such as Col1a1, Col6a6, and Col9a2. Overall, our study provides a full-scale DEG map regarding HFD-induced islet dysfunction, which was helpful to understand the underlying molecular mechanism of islet deterioration further.

Small molecule glucagon release inhibitors with activity in human islets.

Purpose: Type 1 diabetes (T1D) accounts for an estimated 5% of all diabetes in the United States, afflicting over 1.25 million individuals. Maintaining long-term blood glucose control is the major goal for individuals with T1D. In T1D, insulin-secreting pancreatic islet ß-cells are destroyed by the immune system, but glucagon-secreting islet α-cells survive. These remaining α-cells no longer respond properly to fluctuating blood glucose concentrations. Dysregulated α-cell function contributes to hyper- and hypoglycemia which can lead to macrovascular and microvascular complications. To this end, we sought to discover small molecules that suppress α-cell function for their potential as preclinical candidate compounds. Prior high-throughput screening identified a set of glucagon-suppressing compounds using a rodent α-cell line model, but these compounds were not validated in human systems. Results: Here, we dissociated and replated primary human islet cells and exposed them to 24 h treatment with this set of candidate glucagon-suppressing compounds. Glucagon accumulation in the medium was measured and we determined that compounds SW049164 and SW088799 exhibited significant activity. Candidate compounds were also counter-screened in our InsGLuc-MIN6 ß-cell insulin secretion reporter assay. SW049164 and SW088799 had minimal impact on insulin release after a 24 h exposure. To further validate these hits, we treated intact human islets with a selection of the top candidates for 24 h. SW049164 and SW088799 significantly inhibited glucagon release into the medium without significantly altering whole islet glucagon or insulin content. In concentration-response curves SW088799 exhibited significant inhibition of glucagon release with an IC50 of 1.26 µM. Conclusion: Given the set of tested candidates were all top hits from the primary screen in rodent α-cells, this suggests some conservation of mechanism of action between human and rodents, at least for SW088799. Future structure-activity relationship studies of SW088799 may aid in elucidating its protein target(s) or enable its use as a tool compound to suppress α-cell activity in vitro.

FAP expression in alpha cells of Langherhans insulae-implications for FAPI radiopharmaceuticals' use.

PURPOSE: Radiopharmaceuticals targeting fibroblast activation protein (FAP) alpha are increasingly studied for diagnostic and therapeutic applications. We discovered FAP expression at immunohistochemistry (IHC) in the alpha cells of the Langerhans insulae of few patients. Therefore, we planned an investigation aimed at describing FAP expression in the pancreas and discussing the implications for radioligand applications. METHODS: We retrospectively included 40 patients from 2 institutions (20 pts each) according to the following inclusion/exclusion criteria: (i) pathology proven pancreatic ductal adenocarcinoma and neuroendocrine tumors (NET), 10 pts per each group at each center; (ii) and availability of paraffin-embedded tissue; and (iii) clinical-pathological records. We performed IHC analysis and applied a semiquantitative visual scoring system (0, negative staining; 1, present in less than 30%; 2, present in more than 30% of the area). FAP expression was assessed according to histology-NET (n = 20) vs ductal adenocarcinoma (n = 20)-and to previous treatments within the adenocarcinoma group. The local ethics committee approved the study (No. INT 21/16, 28 January 2016). RESULTS: The population consisted of 24 males and 16 females, with a median age of 68 and a range of 14-84 years; 8/20 adenocarcinoma patients received chemotherapy. In all the Langerhans insulae (40/40), pancreatic alpha cells were found to express FAP, with a score of 2. No difference was found among NET (20/20) and adenocarcinoma (20/20), nor according to neoadjuvant chemotherapy in the adenocarcinoma cohort (received or not received). CONCLUSION: Pancreatic Langerhans islet alpha cells normally express FAP. This is not expected to influence the diagnostic accuracy of FAP-targeting tracers. In the therapeutic setting, our results suggest the need to better elucidate FAPI radioligands' effects on the Langerhans insulae function.