Glucagon-like peptide 1 receptor is a T cell-negative costimulatory molecule.

Glucagon-like peptide-1 receptor (GLP-1R) is a key regulator of glucose metabolism known to be expressed by pancreatic ß cells. We herein investigated the role of GLP-1R on T lymphocytes during immune response. Our data showed that a subset of T lymphocytes expresses GLP-1R, which is upregulated during alloimmune response, similarly to PD-1. When mice received islet or cardiac allotransplantation, an expansion of GLP-1Rpos T cells occurred in the spleen and was found to infiltrate the graft. Additional single-cell RNA sequencing (scRNA-seq) analysis conducted on GLP-1Rpos and GLP-1Rneg CD3+ T cells unveiled the existence of molecular and functional dissimilarities between both subpopulations, as the GLP-1Rpos are mainly composed of exhausted CD8 T cells. GLP-1R acts as a T cell-negative costimulatory molecule, and GLP-1R signaling prolongs allograft survival, mitigates alloimmune response, and reduces T lymphocyte graft infiltration. Notably, GLP-1R antagonism triggered anti-tumor immunity when tested in a preclinical mouse model of colorectal cancer.

TMEM219 regulates the transcription factor expression and proliferation of beta cells.

Pancreatic beta cells replenishment is considered the next therapeutic option for type 1 diabetes; while stimulating endogenous beta cells proliferation is the "holy grail" for those patients with exhausted beta cell mass. Here we are demonstrating that the pro-apoptotic receptor TMEM219 is expressed in fetal pancreas, in beta cell precursors and in in vitro embryonic-derived endocrine progenitors. TMEM219 signaling negatively regulates beta cells at early stages and induces Caspase 8-mediated cell death. Pharmacological blockade of TMEM219 further rescued beta cell precursor and proliferation markers, and decreased cell death, both in islets and in in vitro-derived endocrine progenitors, allowing for beta cell preservation. While addressing the upstream controlling TMEM219 expression, we determined the TMEM219 miRNet; indeed, one of those miRNAs, miR-129-2, is highly expressed in human islets, particularly in patients at risk or with established type 1 diabetes. miR-129-2 mimic downregulated TMEM219 expression in islets, in in vitro embryonic-derived endocrine progenitors and in highly proliferating insulinoma-derived cells. Moreover, miR-129-2 inhibitor induced a TMEM219 overexpression in insulinoma-derived cells, which restored cell proliferation and functional markers, thus acting as endogenous regulator of TMEM219 expression. The TMEM219 upstream regulator miR129-2 controls the fate of beta cell precursors and may unleash their regenerative potentials to replenish beta cells in type 1 diabetes.

Daytime hypoglycemic episodes during the use of an advanced hybrid closed loop system.

AIMS: The use of advanced hybrid closed loop systems is spreading due to the beneficial effects on glycometabolic control obtained in patients with type 1 diabetes. However, hypoglycemic episodes can be sometimes a matter of concern. We aim to compare the hypoglycemic risk of an advanced hybrid closed loop system and a predictive low glucose suspend sensor augmented pump. METHODS: In this retrospective three months observational study, we included 30 patients using Medtronic Minimed™ 780G advanced hybrid closed loop system and 30 patients using a Medtronic Minimed™ predictive low glucose suspend sensor augmented pump. RESULTS: The advanced hybrid closed loop system reduced the time spent above 180 mg/dL threshold and increased the time in range as compared to the predictive low glucose suspend. No severe hypoglycemia occurred in both groups and no differences were observed in the percentage of time spent below 70 mg/dl and 54 mg/dl glucose threshold. Nevertheless, more hypoglycemic episodes were recorded during daytime, but not in nighttime, with the use of the advanced hybrid closed loop system. CONCLUSIONS: Our results confirmed the general improvement of glycemic outcomes obtained with the advanced hybrid closed loop system; however more hypoglycemic episodes during daytime were evident.

Reversal of Experimental Autoimmune Diabetes With an sCD39/Anti-CD3 Treatment.

Extracellular (e)ATP, a potent proinflammatory molecule, is released by dying/damaged cells at the site of inflammation and is degraded by the membrane ectonucleotidases CD39 and CD73. In this study, we sought to unveil the role of eATP degradation in autoimmune diabetes. We then assessed the effect of soluble CD39 (sCD39) administration in prevention and reversal studies in NOD mice as well as in mechanistic studies. Our data showed that eATP levels were increased in hyperglycemic NOD mice compared with prediabetic NOD mice. CD39 and CD73 were found expressed by both α- and ß-cells and by different subsets of T cells. Importantly, prediabetic NOD mice displayed increased frequencies of CD3+CD73+CD39+ cells within their pancreata, pancreatic lymph nodes, and spleens. The administration of sCD39 into prediabetic NOD mice reduced their eATP levels, abrogated the proliferation of CD4+- and CD8+-autoreactive T cells, and increased the frequency of regulatory T cells, while delaying the onset of T1D. Notably, concomitant administration of sCD39 and anti-CD3 showed a strong synergism in restoring normoglycemia in newly hyperglycemic NOD mice compared with monotherapy with anti-CD3 or with sCD39. The eATP/CD39 pathway plays an important role in the onset of T1D, and its targeting might represent a potential therapeutic strategy in T1D.

Nanotargeted Delivery of Immune Therapeutics in Type 1 Diabetes.

Immune therapeutics holds great promise in the treatment of type 1 diabetes (T1D). Nonetheless, their progress is hampered by limited efficacy, equipoise, or issues of safety. To address this, a novel and specific nanodelivery platform for T1D that targets high endothelial venules (HEVs) presented in the pancreatic lymph nodes (PLNs) and pancreas is developed. Data indicate that the pancreata of nonobese diabetic (NOD) mice and patients with T1D are unique in their expression of newly formed HEVs. Anti-CD3 mAb is encapsulated in poly(lactic-co-glycolic acid)-poly(ethylene glycol) nanoparticles (NPs), the surfaces of which are conjugated with MECA79 mAb that recognizes HEVs. Targeted delivery of these NPs improves accumulation of anti-CD3 mAb in both the PLNs and pancreata of NOD mice. Treatment of hyperglycemic NOD mice with MECA79-anti-CD3-NPs results in significant reversal of T1D compared to those that are untreated, treated with empty NPs, or provided free anti-CD3. This effect is associated with a significant reduction of T effector cell populations in the PLNs and a decreased production of pro-inflammatory cytokine in the mice treated with MECA79-anti-CD3-NPs. In summary, HEV-targeted therapeutics may be used as a means by which immune therapeutics can be delivered to PLNs and pancreata to suppress autoimmune diabetes effectively.

Broadening horizons in mechanisms, management, and treatment of diabetic kidney disease.

Diabetic kidney disease (DKD) is the first cause of end-stage kidney disease in patients with diabetes and its prevalence is increasing worldwide. It encompasses histological alterations that mainly affect the glomerular filtration unit, which include thickening of the basement membrane, mesangial cell proliferation, endothelial alteration, and podocyte injury. These morphological abnormalities further result in a persistent increase of urinary albumin-to-creatinine ratio and in a reduction of the estimated glomerular filtration rate. Several molecular and cellular mechanisms have been recognized, up to date, as major players in mediating such clinical and histological features and many more are being under investigation. This review summarizes the most recent advances in understanding cell death mechanisms, intracellular signaling pathways and molecular effectors that play a role in the onset and progression of diabetic kidney damage. Some of those molecular and cellular mechanisms have been already successfully targeted in preclinical models of DKD and, in some cases, strategies have been tested in clinical trials. Finally, this report sheds light on the relevance of novel pathways that may become therapeutic targets for future applications in DKD.

eATP and autoimmune diabetes.

PURPOSE OF REVIEW: The purine nucleotide adenosine triphosphate (ATP) is released into extracellular spaces as extracellular ATP (eATP) as a consequence of cell injury or death and activates the purinergic receptors. Once released, eATP may facilitate T-lymphocyte activation and differentiation. The purpose of this review is to elucidate the role of ATP-mediated signaling in the immunological events related to type 1 diabetes (T1D). RECENT FINDINGS: T lymphocytes mediate immune response during the onset of T1D and promote pancreatic islet or whole pancreas rejection in transplantation. Recent data suggest a potential role for eATP in early steps of T1D onset and of allograft rejection. In different preclinical experimental models and clinical trials, several drugs targeting purinergic signaling have been employed to abrogate lymphocyte activation and differentiation, thus representing an achievable treatment to prevent/revert T1D or to induce long-term islet allograft function. SUMMARY: In preclinical and clinical settings, eATP-signaling inhibition induces immune tolerance in autoimmune disease and in allotransplantation. In this view, the purinergic system may represent a novel therapeutic target for auto- and allo-immunity.

The anti-inflammatory and immunological properties of GLP-1 Receptor Agonists.

In the last few years, a great interest has emerged in investigating the pleiotropic effects of Glucagon Like Peptide-1 Receptor Agonists (GLP-1RAs). While GLP-1RAs ability to lower plasma glucose and to induce weight loss has allowed them to be approved for the treatment of diabetes and obesity, consistent evidences from in vitro studies and preclinical models suggested that GLP-1RAs have anti-inflammatory properties and that may modulate the immune-system. Notably, such anti-inflammatory effects target different pathways in different tissues, underling the broad spectrum of GLP-1RAs actions. This review examines some of the currently proposed molecular mechanisms of GLP-1RAs actions and explores their potential benefits in reducing inflammatory responses, which may well suggest a future therapeutic use of GLP-1RAs in new indications.

Abnormalities of the oculomotor function in type 1 diabetes and diabetic neuropathy.

AIMS: Abnormalities in the oculomotor system may represent an early sign of diabetic neuropathy and are currently poorly studied. We designed an eye-tracking-based test to evaluate oculomotor function in patients with type 1 diabetes. METHODS: We used the SRLab-Tobii TX300 Eye tracker®, an eye-tracking device, coupled with software that we developed to test abnormalities in the oculomotor system. The software consists of a series of eye-tracking tasks divided into 4 classes of parameters (Resistance, Wideness, Pursuit and Velocity) to evaluate both smooth and saccadic movement in different directions. We analyzed the oculomotor system in 34 healthy volunteers and in 34 patients with long-standing type 1 diabetes. RESULTS: Among the 474 parameters analyzed with the eye-tracking-based system, 11% were significantly altered in patients with type 1 diabetes (p < 0.05), with a higher proportion of abnormalities observed in the Wideness (24%) and Resistance (10%) parameters. Patients with type 1 diabetes without diabetic neuropathy showed more frequently anomalous measurements in the Resistance class (p = 0.02). The classes of Velocity and Pursuit were less frequently altered in patients with type 1 diabetes as compared to healthy subjects, with anomalous measurements mainly observed in patients with diabetic neuropathy. CONCLUSIONS: Abnormalities in oculomotor system function can be detected in patients with type 1 diabetes using a novel eye-tracking-based test. A larger cohort study may further determine thresholds of normality and validate whether eye-tracking can be used to non-invasively characterize early signs of diabetic neuropathy. TRIAL: NCT04608890.

Indirect and Direct Effects of SARS-CoV-2 on Human Pancreatic Islets.

Recent studies have shown that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection may induce metabolic distress, leading to hyperglycemia in patients affected by coronavirus disease 19 (COVID-19). We investigated the potential indirect and direct effects of SARS-CoV-2 on human pancreatic islets in 10 patients who became hyperglycemic after COVID-19. Although there was no evidence of peripheral anti-islet autoimmunity, the serum of these patients displayed toxicity on human pancreatic islets, which could be abrogated by the use of anti-interleukin-1ß (IL-1ß), anti-IL-6, and anti-tumor necrosis factor α, cytokines known to be highly upregulated during COVID-19. Interestingly, the receptors of those aforementioned cytokines were highly expressed on human pancreatic islets. An increase in peripheral unmethylated INS DNA, a marker of cell death, was evident in several patients with COVID-19. Pathology of the pancreas from deceased hyperglycemic patients who had COVID-19 revealed mild lymphocytic infiltration of pancreatic islets and pancreatic lymph nodes. Moreover, SARS-CoV-2-specific viral RNA, along with the presence of several immature insulin granules or proinsulin, was detected in postmortem pancreatic tissues, suggestive of ß-cell-altered proinsulin processing, as well as ß-cell degeneration and hyperstimulation. These data demonstrate that SARS-CoV-2 may negatively affect human pancreatic islet function and survival by creating inflammatory conditions, possibly with a direct tropism, which may in turn lead to metabolic abnormalities observed in patients with COVID-19.

Immunogenicity and Safety of SARS-CoV-2 mRNA Vaccines in a Cohort of Patients With Type 1 Diabetes.

Patients with type 1 diabetes (T1D) may develop severe outcomes during coronavirus disease 2019 (COVID-19), but their ability to generate an immune response against the SARS-CoV-2 mRNA vaccines remains to be established. We evaluated the safety, immunogenicity, and glycometabolic effects of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mRNA vaccines in patients with T1D. A total of 375 patients (326 with T1D and 49 subjects without diabetes) who received two doses of the SARS-CoV-2 mRNA vaccines (mRNA-1273, BNT162b2) between March and April 2021 at ASST Fatebenefratelli Sacco were included in this monocentric observational study. Local and systemic adverse events were reported in both groups after SARS-CoV-2 mRNA vaccination, without statistical differences between them. While both patients with T1D and subjects without diabetes exhibited a parallel increase in anti-SARS-CoV-2 spike titers after vaccination, the majority of patients with T1D (70% and 78%, respectively) did not show any increase in the SARS-CoV-2-specific cytotoxic response compared with the robust increase observed in all subjects without diabetes. A reduced secretion of the T-cell-related cytokines interleukin-2 and tumor necrosis factor-α in vaccinated patients with T1D was also observed. No glycometabolic alterations were evident in patients with T1D using continuous glucose monitoring during follow-up. Administration of the SARS-CoV-2 mRNA vaccine is associated with an impaired cellular SARS-CoV-2-specific cytotoxic immune response in patients with T1D.

Pharmacologically Enhanced Regulatory Hematopoietic Stem Cells Revert Experimental Autoimmune Diabetes and Mitigate Other Autoimmune Disorders.

Type 1 diabetes (T1D) is characterized by the loss of immune self-tolerance, resulting in an aberrant immune responses against self-tissue. A few therapeutics have been partially successful in reverting or slowing down T1D progression in patients, and the infusion of autologous hematopoietic stem cells (HSCs) is emerging as an option to be explored. In this study, we proposed to pharmacologically enhance by ex vivo modulation with small molecules the immunoregulatory and trafficking properties of HSCs to provide a safer and more efficacious treatment option for patients with T1D and other autoimmune disorders. A high-throughput targeted RNA sequencing screening strategy was used to identify a combination of small molecules (16,16-dimethyl PGE2 and dexamethasone), which significantly upregulate key genes involved in trafficking (e.g., CXCR4) and immunoregulation (e.g., programmed death ligand 1). The pharmacologically enhanced, ex vivo-modulated HSCs (regulatory HSCs [HSC.Regs]) have strong trafficking properties to sites of inflammation in a mouse model of T1D, reverted autoimmune diabetes in NOD mice, and delayed experimental multiple sclerosis and rheumatoid arthritis in preclinical models. Mechanistically, HSC.Regs reduced lymphocytic infiltration of pancreatic ß cells and inhibited the activity of autoreactive T cells. Moreover, when tested in clinically relevant in vitro autoimmune assays, HSC.Regs abrogated the autoimmune response. Ex vivo pharmacological modulation enhances the immunoregulatory and trafficking properties of HSCs, thus generating HSC.Regs, which mitigated autoimmune diabetes and other autoimmune disorders.

The IGFBP3/TMEM219 pathway regulates beta cell homeostasis.

Loss of pancreatic beta cells is a central feature of type 1 (T1D) and type 2 (T2D) diabetes, but a therapeutic strategy to preserve beta cell mass remains to be established. Here we show that the death receptor TMEM219 is expressed on pancreatic beta cells and that signaling through its ligand insulin-like growth factor binding protein 3 (IGFBP3) leads to beta cell loss and dysfunction. Increased peripheral IGFBP3 was observed in established and at-risk T1D/T2D patients and was confirmed in T1D/T2D preclinical models, suggesting that dysfunctional IGFBP3/TMEM219 signaling is associated with abnormalities in beta cells homeostasis. In vitro and in vivo short-term IGFBP3/TMEM219 inhibition and TMEM219 genetic ablation preserved beta cells and prevented/delayed diabetes onset, while long-term IGFBP3/TMEM219 blockade allowed for beta cell expansion. Interestingly, in several patients' cohorts restoration of appropriate IGFBP3 levels was associated with improved beta cell function. The IGFBP3/TMEM219 pathway is thus shown to be a physiological regulator of beta cell homeostasis and is also demonstrated to be disrupted in T1D/T2D. IGFBP3/TMEM219 targeting may therefore serve as a therapeutic option in diabetes.

Novel Soluble Mediators of Innate Immune System Activation in Solid Allograft Rejection.

During the past years, solid allograft rejection has been considered the consequence of either cellular- or antibody-mediated reaction both being part of the adaptive immune response, whereas the role of innate immunity has been mostly considered less relevant. Recently, a large body of evidence suggested that the innate immune response and its soluble mediators may play a more important role during solid allograft rejection than originally thought. This review will highlight the role of novel soluble mediators that are involved in the activation of innate immunity during alloimmune response and solid allograft rejection. We will also discuss emerging strategies to alleviate the aforementioned events. Hence, novel, feasible, and safe clinical therapies are needed to prevent allograft loss in solid organ transplantation. Fully understanding the role of soluble mediators of innate immune system activation may help to mitigate solid allograft rejection and improve transplanted recipients' outcomes.

PD-1 blockade counteracts post-COVID-19 immune abnormalities and stimulates the anti-SARS-CoV-2 immune response.

A substantial proportion of patients who have recovered from coronavirus disease-2019 (COVID-19) experience COVID-19-related symptoms even months after hospital discharge. We extensively immunologically characterized patients who recovered from COVID-19. In these patients, T cells were exhausted, with increased PD-1+ T cells, as compared with healthy controls. Plasma levels of IL-1ß, IL-1RA, and IL-8, among others, were also increased in patients who recovered from COVID-19. This altered immunophenotype was mirrored by a reduced ex vivo T cell response to both nonspecific and specific stimulation, revealing a dysfunctional status of T cells, including a poor response to SARS-CoV-2 antigens. Altered levels of plasma soluble PD-L1, as well as of PD1 promoter methylation and PD1-targeting miR-15-5p, in CD8+ T cells were also observed, suggesting abnormal function of the PD-1/PD-L1 immune checkpoint axis. Notably, ex vivo blockade of PD-1 nearly normalized the aforementioned immunophenotype and restored T cell function, reverting the observed post-COVID-19 immune abnormalities; indeed, we also noted an increased T cell-mediated response to SARS-CoV-2 peptides. Finally, in a neutralization assay, PD-1 blockade did not alter the ability of T cells to neutralize SARS-CoV-2 spike pseudotyped lentivirus infection. Immune checkpoint blockade ameliorates post-COVID-19 immune abnormalities and stimulates an anti-SARS-CoV-2 immune response.

Next-gen therapeutics to spare and expand beta-cell mass.

The most effective and physiological way to treat hyperglycemia is to restore beta-cell function and to rescue production of endogenous insulin. Increasing evidence suggests that both type 1 and type 2 diabetes are characterized by a significant defect in beta-cell mass, leading to the manifestation of the disease. Novel alternative approaches are needed to spare and expand beta-cell mass in patients with diabetes. This review sets out to describe the latest findings on how to restore the beta-cell mass and function in both forms of diabetes to modulate their progression.

Hematopoietic Stem Cells in Type 1 Diabetes.

Despite the increasing knowledge of pathophysiological mechanisms underlying the onset of type 1 diabetes (T1D), the quest for therapeutic options capable of delaying/reverting the diseases is still ongoing. Among all strategies currently tested in T1D, the use of hematopoietic stem cell (HSC)-based approaches and of teplizumab, showed the most encouraging results. Few clinical trials have already demonstrated the beneficial effects of HSCs in T1D, while the durability of the effect is yet to be established. Investigators are also trying to understand whether the use of selected and better-characterized HSCs subsets may provide more benefits with less risks. Interestingly, ex vivo manipulated HSCs showed promising results in murine models and the recent introduction of the humanized mouse models accelerated the translational potentials of such studies and their final road to clinic. Indeed, immunomodulatory as well as trafficking abilities can be enhanced in genetically modulated HSCs and genetically engineered HSCs may be viewed as a novel "biologic" therapy, to be further tested and explored in T1D and in other autoimmune/immune-related disorders.

The IL-8-CXCR1/2 axis contributes to diabetic kidney disease.

AIMS/HYPOTHESIS: Inflammation has a major role in diabetic kidney disease. We thus investigated the role of the IL-8-CXCR1/2 axis in favoring kidney damage in diabetes. METHODS: Urinary IL-8 levels were measured in 1247 patients of the Joslin Kidney Study in type 2 diabetes (T2D). The expression of IL-8 and of its membrane receptors CXCR1/CXCR2 was quantified in kidney tissues in patients with T2D and in controls. The effect of CXCR1/2 blockade on diabetic kidney disease was evaluated in db/db mice. RESULTS: IL-8 urinary levels were increased in patients with T2D and diabetic kidney disease, with the highest urinary IL-8 levels found in the patients with the largest decline in glomerular filtration rate, with an increased albumin/creatine ratio and the worst renal outcome. Moreover, glomerular IL-8 renal expression was increased in patients with T2D, as compared to controls. High glucose elicits abundant IL-8 secretion in cultured human immortalized podocytes in vitro. Finally, in diabetic db/db mice and in podocytes in vitro, CXCR1/2 blockade mitigated albuminuria, reduced mesangial expansion, decreased podocyte apoptosis and reduced DNA damage. CONCLUSIONS/INTERPRETATION: The IL-8- CXCR1/2 axis may have a role in diabetic kidney disease by inducing podocyte damage. Indeed, targeting the IL-8-CXCR1/2 axis may reduce the burden of diabetic kidney disease.

Acute and long-term disruption of glycometabolic control after SARS-CoV-2 infection.

Patients with coronavirus disease 2019 (COVID-19) are reported to have a greater prevalence of hyperglycaemia. Cytokine release as a consequence of severe acute respiratory syndrome coronavirus 2 infection may precipitate the onset of metabolic alterations by affecting glucose homeostasis. Here we describe abnormalities in glycometabolic control, insulin resistance and beta cell function in patients with COVID-19 without any pre-existing history or diagnosis of diabetes, and document glycaemic abnormalities in recovered patients 2 months after onset of disease. In a cohort of 551 patients hospitalized for COVID-19 in Italy, we found that 46% of patients were hyperglycaemic, whereas 27% were normoglycaemic. Using clinical assays and continuous glucose monitoring in a subset of patients, we detected altered glycometabolic control, with insulin resistance and an abnormal cytokine profile, even in normoglycaemic patients. Glycaemic abnormalities can be detected for at least 2 months in patients who recovered from COVID-19. Our data demonstrate that COVID-19 is associated with aberrant glycometabolic control, which can persist even after recovery, suggesting that further investigation of metabolic abnormalities in the context of long COVID is warranted.

Regulatory B Cells in Autoimmune Diabetes.

Since they were discovered almost three decades ago, a subset of B cells denoted as regulatory B cells (Bregs) have elicited interest throughout the immunology community. Many investigators have sought to characterize their phenotype and to understand their function and immunosuppressive mechanisms. Indeed, studies in murine models have demonstrated that Bregs possess varied phenotypic markers and could be classified into different subsets whose action and pivotal role depend on the pathological condition or stimuli. Similar conclusions were drawn in clinical settings delineating an analogous Breg population phenotypically resembling the murine Bregs that ultimately may be associated with a state of tolerance. Recent studies suggested that Bregs may play a role in the onset of autoimmune diabetes. This review will focus on deciphering the different subclasses of Bregs, their emerging role in autoimmune diabetes, and their potential use as a cell-based therapeutic.