Glucose-stimulated β-cells exhibit synchronized calcium dynamics across the islet that recruit β-cells to enhance insulin secretion. Compared with calcium dynamics, the formation and cell-to-cell propagation of electrical signals within the islet are poorly characterized. To determine factors that influence the propagation of electrical activity across the islet underlying calcium oscillations and β-cell synchronization, we used high-resolution complementary metal-oxide-semiconductor multielectrode arrays (CMOS-MEA) to measure voltage changes associated with the membrane potential of individual cells within intact C57BL6 mouse islets. We measured fast (milliseconds, spikes) and slow (seconds, waves) voltage dynamics. Single spike activity and wave signal velocity were both glucose-dependent, but only spike activity was influenced by N-methyl-d-aspartate receptor activation or inhibition. A repeated glucose stimulus revealed a highly responsive subset of cells in spike activity. When islets were pretreated for 72 h with glucolipotoxic medium, the wave velocity was significantly reduced. Network analysis confirmed that in response to glucolipotoxicity the synchrony of islet cells was affected due to slower propagating electrical waves and not due to altered spike activity. In summary, this approach provided novel insight regarding the propagation of electrical activity and the disruption of cell-to-cell communication due to excessive stimulation.

The gold standard for assessing the function of human islets or β-like cells derived from stem cells involves their engraftment under the kidney capsule of hyperglycemic, immunodeficient mice. Current models, such as streptozotocin treatment of severely immunodeficient mice or the NRG-Akita strain, are limited due to unstable and variable hyperglycemia and/or high morbidity. To address these limitations, we developed the IsletTester mouse via CRISPR/Cas9-mediated gene editing of glucokinase (Gck), the glucose sensor of the β-cells, directly in NSG zygotes. IsletTester mice are heterozygous for an Arg345→stop mutation in Gck and present with stable random hyperglycemia (∼250 mg/dL [14 mmol/L]), normal lifespan, and fertility. We demonstrate the utility of this model through functional engraftment of both human islets and human embryonic stem cell-derived β-like cells. The IsletTester mouse will enable the study of human islet biology over time and under different physiological conditions and can provide a useful preclinical platform to determine the functionality of stem cell-derived islet products.

Dysregulation and loss of immune tolerance toward pancreatic β-cell autoantigens are features of type 1 diabetes (T1D). Until recently, life-long insulin injection was the only approved treatment for T1D, but it does not address the underlying disease pathology. The aim for antigen-specific immunotherapy (ASI) is to restore tolerance. ASI holds potential as a new therapeutic strategy for treating autoimmune diseases with well-characterized antigens. Peptide ASI using processing-independent CD4+ T-cell epitopes (PIPs) shows promising results in several autoimmune diseases. Here, we successfully applied the principles of PIP design to the T1D autoantigen glutamate decarboxylase 65 (GAD65). Peptides spanning GAD65 predicted to be pan-HLA-DR binding were selected. Peptide 10 (P10) displayed enriched responses in peripheral blood mononuclear cells from people with T1D. The minimal epitope of the P10 peptide was fine mapped using T-cell hybridomas generated from HLA-DRB1*04:01 transgenic mice. This minimal epitope, P10Sol, was demonstrated, using a novel activation-induced marker assay, to induce tolerance to the parent peptide in the transgenic mice. Finally, we show that GAD65 P10Sol PIP is recognized by CD4+ T cells from people with T1D who possess a range of HLA-DR alleles and, therefore, can be defined as a pan-DR-binding peptide with therapeutic potential.

Differentiation protocols used to generate stem cell-derived islet cells yield heterogenous cell populations. We generated a human embryonic stem cell line that reports insulin- and glucagon-expressing cells in vitro and in vivo without altering their differentiation or function. We showed some insulin- and glucagon-expressing bihormonal cells are cell-autonomously fated to become α-like cells. This reporter cell line can be used to further study and improve stem cell-derived islet differentiation and transplantation.

It is unclear why some individuals with type 2 diabetes are unresponsive to treatment with glucagon-like peptide 1 receptor (GLP-1R) agonists, but hypothalamic-pituitary-adrenal (HPA) axis activation could play a role. We used [68Ga]Ga-NODAGA-exendin-4 positron emission tomography/computed tomography to compare pituitary GLP-1R expression between responders and nonresponders to treatment with GLP-1R agonists. Pituitary GLP-1R expression and HPA axis activation did not differ between responders and nonresponders to GLP-1R agonist treatment. In addition, pituitary radiolabeled exendin uptake was markedly higher in men than in women. Further study is required to explain treatment differences and understand sex differences in pituitary radiolabeled exendin uptake.

Identified genetic loci for C-peptide and type 1 diabetes (T1D) age at diagnosis (AAD) explain only a small proportion of their variation. We aimed to identify additional genetic loci associated with C-peptide and AAD. Some HLA allele/haplotypes associated with T1D also contributed to variability of C-peptide and AAD, whereas outside the HLA region, T1D loci were mostly not associated with C-peptide or AAD. Genetic variation within CTSH can affect AAD. There is still residual heritability of C-peptide and AAD outside of HLA that could benefit from larger meta-genome-wide association studies.

High-affinity islet autoantibodies predict type 1 diabetes in mice and humans and implicate germinal centers (GCs) in type 1 diabetes pathogenesis. T follicular helper (Tfh) cells are increased in type 1 diabetic individuals and alterations in Tfh-like cells in the peripheral blood predicted individual responses to abatacept. Tfh cells support GC responses and depend on the transcriptional repressor BCL6 for their maturation. We therefore hypothesized that CD4-driven deletion of Bcl6 would disrupt essential T-B lymphocyte interactions in GCs to prevent type 1 diabetes. To test this hypothesis, we generated Bcl6fl/fl-CD4.Cre.NOD mice and found they were completely protected against diabetes. Insulitis severity and tertiary lymphoid structure organization were preserved in the pancreas of Bcl6fl/fl-CD4.Cre.NOD mice, which did not show decreases in CD4+, CD8+, and B cell numbers in the pancreas and draining lymph nodes, relative to control Bcl6fl/fl.NOD mice. CD4-driven loss of functional BCL6 resulted in significantly reduced GC B cell and Tfh cell numbers in the pancreatic lymph nodes and pancreas at late prediabetic intervals. Spontaneous anti-insulin autoantibody was blunted in Bcl6fl/fl-CD4.Cre.NOD mice. These data highlight BCL6 as a novel therapeutic target in type 1 diabetes.

Maternally inherited diabetes and deafness (MIDD) is a mitochondrial disorder characterized primarily by hearing impairment and diabetes. m.3243A>G, the most common phenotypic variant, causes a complex rewiring of the cell with discontinuous remodeling of both mitochondrial and nuclear genome expressions. We propose that MIDD depends on a combination of insulin resistance and impaired β-cell function that occurs in the presence of high skeletal muscle heteroplasmy (approximately ≥60%) and more moderate cell heteroplasmy (∼25%-72%) for m.3243A>G. Understanding the complex mechanisms of MIDD is necessary to develop disease-specific management guidelines that are presently lacking.

Embedded in the developmental origins of health and disease (DOHaD) hypothesis, maternal hyperglycemia in utero, from preexisting diabetes or gestational diabetes mellitus, predisposes the offspring to excess adiposity and heightened risk of prediabetes and type 2 diabetes development. This transmission creates a vicious cycle increasing the presence of diabetes from one generation to another, leading to the question: How can we interrupt this vicious cycle? In this article, we present the current state of knowledge on the intergenerational transmission of diabetes from epidemiological life course studies. Then, we discuss the potential mechanisms implicated in the intergenerational transmission of diabetes with a focus on epigenetics. We present novel findings stemming from epigenome-wide association studies of offspring DNA methylation in blood and placental tissues, which shed light on potential molecular mechanisms implicated in the mother-offspring transmission of diabetes. Lastly, with a perspective on how to break the cycle, we consider interventions to prevent offspring obesity and diabetes development before puberty, as a critical period of the intergenerational cycle. This article is part of a series of perspectives that report on research funded by the American Diabetes Association Pathway to Stop Diabetes program.

Identifying patterns of metabolic heterogeneity in type 2 diabetes (T2D) can help in the development of optimal treatment strategies. We aimed to identify metabolic patterns in patients with T2D in the Republic of Korea and analyze the risk of developing diabetes-related complications according to patterns. We identified three distinct metabolic patterns and observed that each pattern was associated with a heightened risk of developing various cardiovascular diseases. These findings highlight the necessity of devising treatment strategies based on these patterns to prevent diabetes-related complications.

Cytochrome c oxidase subunit 6A2 (COX6A2) expression is increased in diabetic islets. Increased COX6A2 promotes β-cell apoptosis via modulation of cyclophilin D-mediated cytochrome c release from mitochondria to the cytoplasm. Carbohydrate-responsive element-binding protein epigenetically regulates COX6A2 expression in β-cells. Roux-en-Y gastric bypass reduces COX6A2 expression by regulating the glucagon-like peptide 1/cAMP-dependent protein kinase/carbohydrate-responsive element-binding protein signaling pathway.

MG53 is predominantly expressed in striated muscles. The role of MG53 in diabetes has gradually been elucidated but is still full of controversy. Some reports have indicated that MG53 is upregulated in animal models with metabolic disorders and that muscle-specific MG53 upregulation is sufficient to induce whole-body insulin resistance and metabolic syndrome through targeting both the insulin receptor (IR) and insulin receptor substrate 1 (IRS-1) for ubiquitin-dependent degradation. Additionally, MG53 has been identified as a myokine/cardiokine that is secreted from striated muscles into the bloodstream, and circulating MG53 has further been shown to trigger insulin resistance by binding to the extracellular domain of the IR, thereby allosterically inhibiting insulin signaling. Conversely, findings have been reported from other studies that contradict these results. Specifically, no significant change in MG53 expression in striated muscles or serum has been observed in diabetic models, and the MG53-mediated degradation of IRS-1 may be insufficient to induce insulin resistance due to the compensatory roles of other IRS subtypes. Furthermore, sustained elevation of MG53 levels in serum or systemic administration of recombinant human MG53 (rhMG53) has shown no impact on metabolic function. In this article, we will fully characterize these two disparate views, strive to provide critical insights into their contrasts, and propose several specific experimental approaches that may yield additional evidence. Our goal is to encourage the scientific community to elucidate the effects of MG53 on metabolic diseases and the molecular mechanisms involved, thereby providing the theoretical basis for the treatment of metabolic diseases and the applications of rhMG53.

There is variability in early-onset autoimmune diabetes presentation in individuals with monogenic autoimmunity; the mechanism(s) underlying this is unclear. We examined whether type 1 diabetes (T1D) polygenic risk contributes to clinical phenotype in monogenic autoimmune diabetes. Individuals with monogenic autoimmune diabetes had higher T1D genetic risk scores compared with control cohorts, driven largely by increased presence of T1D-risk DR3-DQ2 haplotype. Established T1D polygenic risk alleles, particularly class II HLA genes, contribute to clinical presentation in monogenic autoimmunity.

Shared medical appointments (SMAs) for diabetes and group prenatal care (GPC) for pregnant patients have emerged as innovative care delivery models. They have the potential to transform diabetes care by overcoming many of the time limitations of traditional one-on-one clinical visits. There is compelling evidence that SMAs improve glycemic control for nonpregnant patients with diabetes, GPC reduces Black and White health disparities in preterm birth, and diabetes GPC increases postpartum glucose tolerance test uptake among patients with gestational diabetes mellitus. GPC models stand out as one of few interventions that reduce racial health disparities, which we hypothesize occurs because their effect is inadvertently exerted on both the patient and clinician through an over 20-h meaningful shared experience. In this article I explore the evidence for SMAs and GPC in diabetes and pregnancy, theoretical underpinnings of the models, their potential to promote more equitable care, and future directions from my perspective as a physician in high-risk obstetrics and 2019 American Diabetes Association Pathway Accelerator Award recipient. This article is part of a series of perspectives that report on research funded by the American Diabetes Association Pathway to Stop Diabetes program.

Small glycemic increments (≤0.5 mmol/L) can exert suppressive actions on endogenous glucose production (EGP) however it is unclear if this is an insulin dependent or independent process. Here, we performed a low-rate glucose infusion in control participants without diabetes and in people with type 1 diabetes (T1D) to better understand this phenomenon. Glucose kinetics, hormones and metabolites were measured during a 1 mg/kg/min glucose infusion (90 min) which rapidly increased glucose by ∼0.3 mmol/L in control participants. Insulin concentrations and secretion quickly increased by ∼20%, resulting in a ∼40% suppression of EGP, while glucose disposal remained unchanged. Free fatty acids (FFA) and glucagon were gradually suppressed to ∼30% below baseline at 60 min. When repeated under constant basal insulin concentrations in participants with T1D, glucose infusion caused only partial and transient EGP suppression, hence glucose increased in a near-linear manner, reaching levels ∼2 mmol/L above baseline at 90 min. FFAs and glucagon remained unchanged, while glucose disposal modestly increased. This demonstrates that small glycemic increments exert subtle stimulatory effects on insulin secretion that have potent metabolic actions on the liver and adipose tissue. It is conceivable that subtle increases in glucose could potentially serve as a signal for β-cell adaptation.

Smoking is widely regarded as a risk factor for type 2 diabetes because nicotine contributes to insulin resistance by desensitizing the insulin receptors in muscle, liver, or fat. Little is known, however, about the immediate regulation of islet hormonal output by nicotine, an agonist of ionotropic cholinergic receptors. We investigated this by imaging cytosolic Ca2+ dynamics in mouse and human islets using confocal microscopy and measuring glucagon secretion in response to the alkaloid from isolated mouse islets. Nicotine acutely stimulated cytosolic Ca2+ in glucagon-secreting α-cells but not in insulin-secreting β-cells. The 2.8- ± 0.5-fold (P < 0.05) increase in Ca2+, observed in >70% of α-cells, correlated well with a 2.5- ± 0.3-fold stimulation of glucagon secretion. Nicotine-induced elevation of cytosolic Ca2+ relied on influx from the extracellular compartment rather than release of the cation from intracellular depots. Metabotropic cholinergic signaling, monitored at the level of intracellular diacylglycerol, was limited to 69% of α-cells versus 94% of β-cells. We conclude that parasympathetic regulation of pancreatic islet hormone release uses different signaling pathways in β-cells (metabotropic) and α-cells (metabotropic and ionotropic), resulting in the fine-tuning of acetylcholine-induced glucagon exocytosis. Sustained nicotinic stimulation is, therefore, likely to attenuate insulin sensitivity by increasing glucagon release.

Diabetic keratopathy (DK) is a common chronic metabolic disorder that causes ocular surface complications. Among various therapeutic approaches, local delivery of nerve growth factor (NGF) remains the most effective treatment of DK. However, achieving a sustained therapeutic effect with NGF and the frequent drug delivery burden remain challenging during clinical practice. Here, we developed a novel adeno-associated virus (AAV)-based NGF delivery system that achieved 1-year-long-lasting effects by a single injection. We refined the corneal stromal injection technique, resulting in reduced corneal edema and improved AAV distribution homogeneity. AAV serotype AAV.rh10 exhibited high tropism and specificity to corneal nerves. A dose of 2 × 109 vector genomes was determined to achieve efficient Ngf gene expression without inducing corneal immune responses. Moreover, NGF protein was highly expressed in trigeminal ganglion through a retrograde transport mechanism, indicating the capacity for repairing corneal nerve damage at both the root and corneal nerve endings. In a mouse DK model, a single injection of AAV-Ngf into the corneal stroma led to marked corneal nerve regeneration for over 5 months. Together, we provide a novel therapeutic paradigm for long-term effective treatment of DK, and this therapeutic approach is superior to current DK therapies.

Type 1 diabetes treatment stands at a crucial and exciting crossroad since the 2022 U.S. Food and Drug Administration approval of teplizumab to delay disease development. In this article, we discuss four major conceptual and practical issues that emerged as key to further advancement in type 1 diabetes research and therapies. First, collaborative networks leveraging the synergy between the type 1 diabetes research and care community members are key to fostering innovation, know-how, and translation into the clinical arena worldwide. Second, recent clinical trials in presymptomatic stage 2 and recent-onset stage 3 disease have shown the promise, and potential pitfalls, of using immunomodulatory and/or β-cell protective agents to achieve sustained remission or prevention. Third, the increasingly appreciated heterogeneity of clinical, immunological, and metabolic phenotypes and disease trajectories is of critical importance to advance the decision-making process for tailored type 1 diabetes care and therapy. Fourth, the clinical benefits of early diagnosis of β-cell autoimmunity warrant consideration of general population screening for islet autoantibodies, which requires further efforts to address the technical, organizational, and ethical challenges inherent to a sustainable program. Efforts are underway to integrate these four concepts into the future directions of type 1 diabetes research and therapy.