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The intricate etiology of type 1 diabetes mellitus (T1D), characterized by harmful interactions between the immune system and insulin-producing beta cells, has hindered the development of effective therapies including human islet transplantation, which requires strong immunosuppressants that impair beta cell survival and function. As such alternative immunomodulating therapies are required for successful transplantation. The discovery that pharmacological activation of the nuclear receptor LRH-1/NR5A2 can reverse hyperglycemia in mouse models of T1D by altering, and not suppressing the autoimmune attack, prompted us to investigate whether LRH-1/NR5A2 activation could improve human islet function/survival after xenotransplantation in immunocompetent mice. Human islets were transplanted under the kidney capsule of streptozotocin (STZ)-induced diabetic mice, and treatment with BL001 (LRH-1/NR5A2 agonist) or vehicle was administered one week post-transplant. Our study, encompassing 3 independent experiments with 3 different islet donors, revealed that mice treated for 8 weeks with BL001 exhibited lower blood glucose levels correlating with improved mouse survival rates as compared to vehicle-treated controls. Human C-peptide was detectable in BL001-treated mice at both 4 and 8 weeks indicating functional islet beta cells. Accordingly, in mice treated with BL001 for 8 weeks, the beta cell mass was preserved, while a significant decrease in alpha cells was observed compared to mice treated with BL001 for only 4 weeks. In contrast, vehicle-treated mice exhibited a reduction in insulin-expressing cells at 8 weeks compared to those at 4 weeks. These results suggest that BL001 significantly enhances the survival, engraftment, and functionality of human islets in a STZ-induced diabetic mouse model., Competing Interests: Two patents WO 2011 144725 A2 and WO 2016 156531 A1 related to BL001 have been published of which GB and C-VN are inventors. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Cobo-Vuilleumier, Lorenzo, Martin Vazquez, López Noriega, Nano, Piemonti, Martín and Gauthier.)

Autoantibodies to glutamate decarboxylase (GADA) are widely used in the prediction and classification of type 1 diabetes. GADA radiobinding assays (RBAs) using N-terminally truncated antigens offer improved specificity, but radioisotopes limit the high-throughput potential for population screening. Luciferase-based immunoprecipitation system (LIPS) assays are sensitive and specific alternatives to RBAs with the potential to improve risk stratification. The performance of assays using the Nanoluc luciferase (Nluc)-conjugated GAD65 constructs, Nluc-GAD65(96-585) and full length Nluc-GAD65(1-585), were evaluated in 434 well-characterized serum samples from patients with recent-onset type 1 diabetes and first-degree relatives. Nonradioactive, high-throughput LIPS assays are quicker and require less serum than RBAs. Of 171 relatives previously tested single autoantibody positive for autoantibodies to full-length GAD65 by RBA but had not progressed to diabetes, fewer retested positive by LIPS using either truncated (n = 72) or full-length (n = 111) antigen. The Nluc-GAD65(96-585) truncation demonstrated the highest specificity in LIPS assays overall, but in contrast to RBA, N-terminus truncations did not result in a significant increase in disease-specificity compared with the full-length antigen. This suggests that binding of nonspecific antibodies is affected by the conformational changes resulting from addition of the Nluc antigen. Nluc-GAD65(96-585) LIPS assays offer low-blood-volume, high-specificity GADA tests for screening and diagnostics., (© 2024 by the American Diabetes Association.)

Aim/hypothesis: We assessed whether HOMA-IR and the Matsuda Index are associated with transitions through stages of type 1 diabetes., Methods: Autoantibody (AAb)-positive relatives of individuals with type 1 diabetes (n=6256) from the TrialNet Pathway to Prevention were studied. Associations of indicators of insulin resistance (HOMA-IR) and insulin sensitivity (Matsuda Index) with BMI percentile (BMIp) and age were assessed with adjustments for measures of insulin secretion, Index60 and insulinogenic index (IGI). Cox regression was used to determine if tertiles of HOMA-IR and Matsuda Index predicted transitions from Not Staged (<2 AAbs) to Stage 1 (≥2 AAbs and normoglycaemia), from Stage 1 to Stage 2 (≥2 AAbs with dysglycaemia), and progression to Stage 3 (diabetes as defined by WHO/ADA criteria)., Results: There were strong associations of HOMA-IR (positive) and Matsuda Index (inverse) with baseline age and BMIp (p<0.0001). After adjustments for Index60, transitioning from Stage 1 to Stage 2 was associated with higher HOMA-IR and lower Matsuda Index (HOMA-IR: HR=1.71, p<0.0001; Matsuda Index, HR=0.40, p<0.0001), as with progressing from Stages 1 or 2 to Stage 3 (HOMA-IR: HR=1.98, p<0.0001; Matsuda Index: HR=0.46, p<0.0001). Without adjustments, associations of progression to Stage 3 were inverse for HOMA-IR and positive for Matsuda Index, opposite in directionality with adjustments. When IGI was used in place of Index60, the findings were similar., Conclusions/interpretation: Progression to Stages 2 and 3 of type 1 diabetes increases with HOMA-IR and decreases with the Matsuda Index after adjustments for insulin secretion. Indicators of insulin secretion appear helpful for interpreting associations of progression to type 1 diabetes with HOMA-IR or the Matsuda Index in AAb-positive relatives., (© 2023. The Author(s).)

Type 1 diabetes (T1D) presents a persistent medical challenge, demanding innovative strategies for sustained glycemic control and enhanced patient well-being. Beta cells are specialized cells in the pancreas that produce insulin, a hormone that regulates blood sugar levels. When beta cells are damaged or destroyed, insulin production decreases, which leads to T1D. Allo Beta Cell Transplantation has emerged as a promising therapeutic avenue, with the goal of reinstating glucose regulation and insulin production in T1D patients. However, the path to success in this approach is fraught with complex immunological hurdles that demand rigorous exploration and resolution for enduring therapeutic efficacy. This exploration focuses on the distinct immunological characteristics inherent to Allo Beta Cell Transplantation. An understanding of these unique challenges is pivotal for the development of effective therapeutic interventions. The critical role of glucose regulation and insulin in immune activation is emphasized, with an emphasis on the intricate interplay between beta cells and immune cells. The transplantation site, particularly the liver, is examined in depth, highlighting its relevance in the context of complex immunological issues. Scrutiny extends to recipient and donor matching, including the utilization of multiple islet donors, while also considering the potential risk of autoimmune recurrence. Moreover, unanswered questions and persistent gaps in knowledge within the field are identified. These include the absence of robust evidence supporting immunosuppression treatments, the need for reliable methods to assess rejection and treatment protocols, the lack of validated biomarkers for monitoring beta cell loss, and the imperative need for improved beta cell imaging techniques. In addition, attention is drawn to emerging directions and transformative strategies in the field. This encompasses alternative immunosuppressive regimens and calcineurin-free immunoprotocols, as well as a reevaluation of induction therapy and recipient preconditioning methods. Innovative approaches targeting autoimmune recurrence, such as CAR Tregs and TCR Tregs, are explored, along with the potential of stem stealth cells, tissue engineering, and encapsulation to overcome the risk of graft rejection. In summary, this review provides a comprehensive overview of the inherent immunological obstacles associated with Allo Beta Cell Transplantation. It offers valuable insights into emerging strategies and directions that hold great promise for advancing the field and ultimately improving outcomes for individuals living with diabetes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Caldara, Tomajer, Monti, Sordi, Citro, Chimienti, Gremizzi, Catarinella, Tentori, Paloschi, Melzi, Mercalli, Nano, Magistretti, Partelli and Piemonti.)

Aim: In a recent randomized, multicenter trial (NCT02814838) a short-term anti-inflammatory treatment with ladarixin (LDX; an inhibitor of the CXCR1/2 chemokine receptors) did not show benefit on preserving residual beta cell function in new-onset type 1 diabetes. We present a post hoc analysis of trial patients in the predefined subgroup analysis developed according to baseline daily insulin requirement (DIR) tertiles., Method: A double-blind, randomized (2:1), placebo-controlled study was conducted in 45 men and 31 women (aged 18-46 years) within 100 days of the first insulin administration. Patients received LDX (400 mg twice daily) for three cycles of 14 days on/14 days off, or placebo. The primary endpoint was the area under the curve for C-peptide [AUC (0-120 min)] in response to a 2-h mixed meal tolerance test (MMTT) at week 13 ± 1. Seventy-five patients completed the week 13 MMTT and were divided into three groups according to the DIR tertiles: lower, ≤ 0.23U/kg/die (n = 25); middle, 0.24-0.40 U/kg/die (n = 24); upper, ≥ 0.41 U/kg/die (n = 26)., Results: When considering the patients in the upper tertile (HIGH-DIR), C-peptide AUC (0-120 min) at 13 weeks was higher in the LDX group (n = 16) than in the placebo (n = 10) group [difference: 0.72 nmol/L (95% CI 0.9-1.34), p = 0.027]. This difference reduced over time (0.71 nmol/L at 26 weeks, p = 0.04; 0.42 nmol/L at 52 weeks, p = 0.29), while it has never been significant at any time in patients in the lower and/or middle tertile (LOW-DIR). We characterized at baseline the HIGH-DIR and found that endo-metabolic (HOMA-B, adiponectin, and glucagon-to-C-peptide ratio) and immunologic (chemokine (C-C motif) ligand 2 (CCL2)/monocyte chemoattractant protein 1 (MCP1) and Vascular Endothelial Growth Factor (VEGF)) features distinguished this group from LOW-DIR., Conclusion: While LDX did not prevent the progressive loss of beta-cell function in the majority of treated subjects, the post hoc analysis suggests that it could work in subjects with HIGH-DIR at baseline. As we found differences in endo-metabolic and immunologic parameters within this subgroup, this generates the hypothesis that the interactions between host factors and drug action can contribute to its efficacy. Further research is needed to evaluate this hypothesis., Competing Interests: LP served as a consultant for Dompé farmaceutici spa Milan, Italy. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The original clinical trial was funded by Dompé farmaceutici spa Milan, Italy. Each study center received research support and compensation from Dompé farmaceutici spa Milan, Italy to conduct the study and collect data., (Copyright © 2023 Sordi, Monti, Lampasona, Melzi, Pellegrini, Keymeulen, Gillard, Linn, Bosi, Rose, Pozzilli, Giorgino, Cossu and Piemonti.)

Aims: Wolfram Syndrome Spectrum Disorder (WFS1-SD), in its "classic" form, is a rare autosomal recessive disease with poor prognosis and wide phenotypic spectrum. Insulin dependent diabetes mellitus (DM), optic atrophy (OA) diabetes insipidus (DI) and sensorineural deafness (D) are the main features of WFS1-SD. Gonadal dysfunction (GD) has been described mainly in adults with variable prevalence and referred to as a minor clinical feature. This is the first case series investigating gonadal function in a small cohort of paediatric patients affected by WFS1-SD., Methods: Gonadal function was investigated in eight patients (3 male and 5 female) between 3 and 16 years of age. Seven patients have been diagnosed with classic WFS1-SD and one with non-classic WFS1-SD. Gonadotropin and sex hormone levels were monitored, as well as markers of gonadal reserve (inhibin-B and anti-Mullerian hormone). Pubertal progression was assessed according to Tanner staging., Results: Primary hypogonadism was diagnosed in 50% of patients (n=4), more specifically 67% (n=2) of males and 40% of females (n=2). Pubertal delay was observed in one female patient. These data confirm that gonadal dysfunction may be a frequent and underdiagnosed clinical feature in WFS1-SD., Conclusions: GD may represent a frequent and earlier than previously described feature in WFS1-SD with repercussions on morbidity and quality of life. Consequently, we suggest that GD should be included amongst clinical diagnostic criteria for WFS1-SD, as has already been proposed for urinary dysfunction. Considering the heterogeneous and elusive presentation of WFS1-SD, this clinical feature may assist in an earlier diagnosis and timely follow-up and care of treatable associated diseases (i.e. insulin and sex hormone replacement) in these young patients., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Frontino, Di Tonno, Stancampiano, Arrigoni, Rigamonti, Morotti, Canarutto, Bonfanti, Russo, Barera and Piemonti.)

Background: Allogeneic islet transplantation is a validated therapy in type 1 diabetes; however, there is decline of transplanted islet graft function over time and the mechanisms underlying this decline are unclear. We evaluated the distinct association between primary graft function (PGF) and 5-year islet transplantation outcomes., Methods: In this retrospective, multicentre, observational cohort study, we enrolled all patients from the Collaborative Islet Transplant Registry who received islet transplantation alone (ITA recipients) or islet-after-kidney transplantation (IAK recipients) between Jan 19, 1999, and July 17, 2020, with a calculable PGF (exposure of interest), measured 28 days after last islet infusion with a validated composite index of islet graft function (BETA-2 score). The primary outcome was cumulative incidence of unsuccessful islet transplantation, defined as an HbA 1c of 7·0% (53 mmol/mol) or higher, or severe hypoglycaemia (ie, requiring third-party intervention to correct), or a fasting C-peptide concentration of less than 0·2 ng/mL. Secondary outcomes were graft exhaustion (fasting C-peptide <0·3 ng/mL); inadequate glucose control (HbA 1c ≥7·0% [53 mmol/mol] or severe hypoglycaemia); and requirement for exogenous insulin therapy (≥14 consecutive days). Associations between PGF and islet transplantation outcomes were explored with a competing risk analysis adjusted for all covariates suspected or known to affect outcomes. A predictive model based on PGF was built and internally validated by using bootstraps resampling method., Findings: In 39 centres worldwide, we enrolled 1210 patients with a calculable PGF (of those without missing data, mean age 47 years [SD 10], 712 [59·5%] were female, and 865 (97·9%) were White), who received a median of 10·8 thousand islet-equivalents per kg of bodyweight (IQR 7·4-13·5). 986 (82·4%) were ITA recipients and 211 (17·6%) were IAK recipients. Of 1210 patients, 452 (37·4%) received a single islet infusion and 758 (62·6%) received multiple islet infusions. Mean PGF was 14·3 (SD 8·8). The 5-year cumulative incidence of unsuccessful islet transplantation was 70·7% (95% CI 67·2-73·9), and was inversely and linearly related to PGF, with an adjusted subhazard ratio (sHR) of 0·77 (95% CI 0·72-0·82) per 5-unit increase of BETA-2 score (p<0·0001). Secondary endpoints were similarly related to PGF. The model-adjusted median C-statistic values of PGF for predicting 5-year cumulative incidences of unsuccessful islet transplantation, graft exhaustion, inadequate glucose control, and exogenous insulin therapy were 0·70 (range 0·69-0·71), 0·76 (0·74-0·77), 0·65 (0·64-0·66), and 0·72 (0·71-0·73), respectively., Interpretation: This global multicentre study reports a linear and independent association between PGF and 5-year clinical outcomes of islet transplantation. The main study limitations are its retrospective design and the absence of analysis of complications., Funding: Public Health Service Research, National Institutes of Health, Juvenile Diabetes Research Foundation International, Agence National de la Recherche, Fondation de l'Avenir, and Fonds de Dotation Line Renaud-Loulou Gasté., Competing Interests: Declaration of interests CB, EP and FB were employed by The EMMES Company. All other authors declare no competing interests., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Aims/hypothesis: Islet autoantibodies (AAbs) are detected in >90% of individuals with clinically suspected type 1 diabetes at disease onset. A single AAb, sometimes at low titre, is often detected in some individuals, making their diagnosis uncertain. Type 1 diabetes genetic risk scores (GRS) are a useful tool for discriminating polygenic autoimmune type 1 diabetes from other types of diabetes, particularly the monogenic forms, but testing is not routinely performed in the clinic. Here, we used a type 1 diabetes GRS to screen for monogenic diabetes in individuals with weak evidence of autoimmunity, i.e. with a single AAb at disease onset., Methods: In a pilot study, we genetically screened 142 individuals with suspected type 1 diabetes, 42 of whom were AAb-negative, 27 of whom had a single AAb (single AAb-positive) and 73 of whom had multiple AAbs (multiple AAb-positive) at disease onset. Next-generation sequencing (NGS) was performed in 41 AAb-negative participants, 26 single AAb-positive participants and 60 multiple AAb-positive participants using an analysis pipeline of more than 200 diabetes-associated genes., Results: The type 1 diabetes GRS was significantly lower in AAb-negative individuals than in those with a single and multiple AAbs. Pathogenetic class 4/5 variants in MODY or monogenic diabetes genes were identified in 15/41 (36.6%) AAb-negative individuals, while class 3 variants of unknown significance were identified in 17/41 (41.5%). Residual C-peptide levels at diagnosis were higher in individuals with mutations compared to those without pathogenetic variants. Class 3 variants of unknown significance were found in 11/26 (42.3%) single AAb-positive individuals, and pathogenetic class 4/5 variants were present in 2/26 (7.7%) single AAb-positive individuals. No pathogenetic class 4/5 variants were identified in multiple AAb-positive individuals, but class 3 variants of unknown significance were identified in 19/60 (31.7%) patients. Several patients across the three groups had more than one class 3 variant., Conclusions/interpretation: These findings provide insights into the genetic makeup of patients who show weak evidence of autoimmunity at disease onset. Absence of islet AAbs or the presence of a single AAb together with a low type 1 diabetes GRS may be indicative of a monogenic form of diabetes, and use of NGS may improve the accuracy of diagnosis., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Intrahepatic islet transplantation is the standard cell therapy for β cell replacement. However, the shortage of organ donors and an unsatisfactory engraftment limit its application to a selected patients with type 1 diabetes. There is an urgent need to identify alternative strategies based on an unlimited source of insulin producing cells and innovative scaffolds to foster cell interaction and integration to orchestrate physiological endocrine function. We previously proposed the use of decellularized lung as a scaffold for β cell replacement with the final goal of engineering a vascularized endocrine organ. Here, we prototyped this technology with the integration of neonatal porcine islet and healthy subject-derived blood outgrowth endothelial cells to engineer a xenogeneic vascularized endocrine pancreas. We validated ex vivo cell integration and function, its engraftment and performance in a preclinical model of diabetes. Results showed that this technology not only is able to foster neonatal pig islet maturation in vitro, but also to perform in vivo immediately upon transplantation and for over 18 weeks, compared to normal performance within 8 weeks in various state of the art preclinical models. Given the recent progress in donor pig genetic engineering, this technology may enable the assembly of immune-protected functional endocrine organs., (© 2023. The Author(s).)

The field of cell therapy of type 1 diabetes is a particularly interesting example in the scenario of regenerative medicine. In fact, β-cell replacement has its roots in the experience of islet transplantation, which began 40 years ago and is currently a rapidly accelerating field, with several ongoing clinical trials using β cells derived from stem cells. Type 1 diabetes is particularly suitable for cell therapy as it is a disease due to the deficiency of only one cell type, the insulin-producing β cell, and this endocrine cell does not need to be positioned inside the pancreas to perform its function. On the other hand, the presence of a double immunological barrier, the allogeneic one and the autoimmune one, makes the protection of β cells from rejection a major challenge. Until today, islet transplantation has taught us a lot, pioneering immunosuppressive therapies, graft encapsulation, tissue engineering, and test of different implant sites and has stimulated a great variety of studies on β-cell function. This review starts from islet transplantation, presenting its current indications and the latest published trials, to arrive at the prospects of stem cell therapy, presenting the latest innovations in the field., (© 2022 S. Karger AG, Basel.)

An unbiased and replicable profiling of type 1 diabetes (T1D)-specific circulating immunome at disease onset has yet to be identified due to experimental and patient selection limitations. Multicolor flow cytometry was performed on whole blood from a pediatric cohort of 107 patients with new-onset T1D, 85 relatives of T1D patients with 0-1 islet autoantibodies (pre-T1D&#95;LR), 58 patients with celiac disease or autoimmune thyroiditis (CD&#95;THY) and 76 healthy controls (HC). Unsupervised clustering of flow cytometry data, validated by a semi-automated gating strategy, confirmed previous findings showing selective increase of naïve CD4 T cells and plasmacytoid DCs, and revealed a decrease in CD56 bright NK cells in T1D. Furthermore, a non-selective decrease of CD3 + CD56 + regulatory T cells was observed in T1D. The frequency of naïve CD4 T cells at disease onset was associated with partial remission, while it was found unaltered in the pre-symptomatic stages of the disease. Thanks to a broad cohort of pediatric individuals and the implementation of unbiased approaches for the analysis of flow cytometry data, here we determined the circulating immune fingerprint of newly diagnosed pediatric T1D and provide a reference dataset to be exploited for validation or discovery purposes to unravel the pathogenesis of T1D., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bechi Genzano, Bezzecchi, Carnovale, Mandelli, Morotti, Castorani, Favalli, Stabilini, Insalaco, Ragogna, Codazzi, Scotti, Del Rosso, Mazzi, De Pellegrin, Giustina, Piemonti, Bosi, Battaglia, Morelli, Bonfanti and Petrelli.)

Background: Non-coding genetic variants that influence gene transcription in pancreatic islets play a major role in the susceptibility to type 2 diabetes (T2D), and likely also contribute to type 1 diabetes (T1D) risk. For many loci, however, the mechanisms through which non-coding variants influence diabetes susceptibility are unknown., Results: We examine splicing QTLs (sQTLs) in pancreatic islets from 399 human donors and observe that common genetic variation has a widespread influence on the splicing of genes with established roles in islet biology and diabetes. In parallel, we profile expression QTLs (eQTLs) and use transcriptome-wide association as well as genetic co-localization studies to assign islet sQTLs or eQTLs to T2D and T1D susceptibility signals, many of which lack candidate effector genes. This analysis reveals biologically plausible mechanisms, including the association of T2D with an sQTL that creates a nonsense isoform in ERO1B, a regulator of ER-stress and proinsulin biosynthesis. The expanded list of T2D risk effector genes reveals overrepresented pathways, including regulators of G-protein-mediated cAMP production. The analysis of sQTLs also reveals candidate effector genes for T1D susceptibility such as DCLRE1B, a senescence regulator, and lncRNA MEG3., Conclusions: These data expose widespread effects of common genetic variants on RNA splicing in pancreatic islets. The results support a role for splicing variation in diabetes susceptibility, and offer a new set of genetic targets with potential therapeutic benefit., (© 2022. The Author(s).)

Aim: To evaluate the ability of ladarixin (LDX, 400 mg twice-daily for three cycles of 14 days on/14 days off), an inhibitor of the CXCR1/2 chemokine receptors, to maintain C-peptide production in adult patients with newly diagnosed type 1 diabetes., Materials and Methods: A double-blind, randomized (2:1), placebo-controlled study was conducted in 45 males and 31 females (aged 18-46 years) within 100 days of the first insulin administration. The primary endpoint was the area under the curve (AUC) for C-peptide in response to a 2-hour mixed meal tolerance test (AUC [0-120 min] ) at week 13 ± 1. Secondary endpoints included C-peptide AUC (15-120 min) , HbA1c, daily insulin requirement, severe hypoglycaemic events (SHE), the proportion of subjects achieving HbA1c less than 7.0% without SHE and maintaining a residual beta cell function. Follow-up assessments were scheduled at weeks 13 ± 1, 26 ± 2 and 52 ± 2., Results: In total, 26/26 (100%, placebo) and 49/50 (98%, LDX) patients completed week 13. The mean change from baseline to week 13 in C-peptide AUC (0-120 min) was -0.144 ± 0.449 nmol/L with placebo and 0.003 ± .322 nmol/L with LDX. The difference was not significant (0.149 nmol/L, 95% CI -0.04 to 0.33; P = .122). At week 26, the proportion of patients with HbA1c less than 7.0% without SHE was transiently higher in the LDX group (81% vs. 54%, P = .024). Otherwise, no significant secondary endpoint differences were noted. Transient metabolic benefit was seen at week 26 in favour of the LDX group in the prespecified subpopulation with fasting C-peptide less than the median value at screening., Conclusions: In newly diagnosed patients with type 1 diabetes, short-term LDX treatment had no appreciable effect on preserving residual beta cell function., (© 2022 The Authors. Diabetes, Obesity and Metabolism published by John Wiley & Sons Ltd.)

In the attempt to understand the origin of autoantibody (AAb) production in patients with and at risk for type 1 diabetes (T1D), multiple studies have analyzed and reported alterations in T follicular helper (Tfh) cells in presymptomatic AAb + subjects and patients with T1D. Yet, whether the regulatory counterpart of Tfh cells, represented by T follicular regulatory (Tfr) cells, is similarly altered is still unclear. To address this question, we performed analyses in peripheral blood, spleen, and pancreatic lymph nodes (PLN) of organ donor subjects with T1D. Blood analyses were also performed in living AAb - and AAb + subjects. While negligible differences in the frequency and phenotype of blood Tfr cells were observed among T1D, AAb - , and AAb + adult subjects, the frequency of Tfr cells was significantly reduced in spleen and PLN of T1D as compared with nondiabetic control subjects. Furthermore, adoptive transfer of Tfr cells delayed disease development in a mouse model of T1D, a finding that could indicate that Tfr cells play an important role in peripheral tolerance and regulation of autoreactive Tfh cells. Together, our findings provide evidence of Tfr cell alterations within disease-relevant tissues in patients with T1D, suggesting a role for Tfr cells in defective humoral tolerance and disease pathogenesis., (© 2021 by the American Diabetes Association.)

One hundred years after its discovery, insulin remains the life-saving therapy for many patients with diabetes. It has been a 100-years-old success story thanks to the fact that insulin therapy has continuously integrated the knowledge developed over a century. In 1982, insulin becomes the first therapeutic protein to be produced using recombinant DNA technology. The first "mini" insulin pump and the first insulin pen become available in 1983 and 1985, respectively. In 1996, the first generation of insulin analogues were produced. In 1999, the first continuous glucose-monitoring device for reading interstitial glucose was approved by the FDA. In 2010s, the ultra-long action insulins were introduced. An equally exciting story developed in parallel. In 1966. Kelly et al. performed the first clinical pancreas transplant at the University of Minnesota, and now it is a well-established clinical option. First successful islet transplantations in humans were obtained in the late 1980s and 1990s. Their ability to consistently re-establish the endogenous insulin secretion was obtained in 2000s. More recently, the possibility to generate large numbers of functional human β cells from pluripotent stem cells was demonstrated, and the first clinical trial using stem cell-derived insulin producing cell was started in 2014. This year, the discovery of this life-saving hormone turns 100 years. This provides a unique opportunity not only to celebrate this extraordinary success story, but also to reflect on the limits of insulin therapy and renew the commitment of the scientific community to an insulin free world for our patients., (© 2021. Springer-Verlag Italia S.r.l., part of Springer Nature.)

Ongoing beta cell death in type 1 diabetes (T1D) can be detected using biomarkers selectively discharged by dying beta cells into plasma. microRNA-375 (miR-375) ranks among the top biomarkers based on studies in animal models and human islet transplantation. Our objective was to identify additional microRNAs that are co-released with miR-375 proportionate to the amount of beta cell destruction. RT-PCR profiling of 733 microRNAs in a discovery cohort of T1D patients 1 h before/after islet transplantation indicated increased plasma levels of 22 microRNAs. Sub-selection for beta cell selectivity resulted in 15 microRNAs that were subjected to double-blinded multicenter analysis. This led to the identification of eight microRNAs that were consistently increased during early graft destruction: besides miR-375, these included miR-132/204/410/200a/429/125b, microRNAs with known function and enrichment in beta cells. Their potential clinical translation was investigated in a third independent cohort of 46 transplant patients by correlating post-transplant microRNA levels to C-peptide levels 2 months later. Only miR-375 and miR-132 had prognostic potential for graft outcome, and none of the newly identified microRNAs outperformed miR-375 in multiple regression. In conclusion, this study reveals multiple beta cell-enriched microRNAs that are co-released with miR-375 and can be used as complementary biomarkers of beta cell death.

Allogeneic islet transplantation is a standard of care treatment for patients with labile type 1 diabetes in many countries around the world, including Japan, the United Kingdom, Australia, much of continental Europe, and parts of Canada. The United States is now endorsing islet cell treatment for type 1 diabetes, but the FDA has chosen to consider islets as a biologic that requires licensure, making the universal implementation of the procedure in the clinic very challenging and opening the manufacture of islet grafts to private companies. The commercialization of human tissues raises significant legal and ethical issues and ironically leads to a situation where treatments developed as a result of the scientific and economic efforts of academia over several decades become exploited exclusively by for-profit entities., (© 2021 Steunstichting ESOT. Published by John Wiley & Sons Ltd.)

Background: Antibody responses to virus reflect exposure and potential protection., Methods: We developed a highly specific and sensitive approach to measuring antibodies against SARS-CoV-2 for population-scale immune surveillance. Antibody positivity was defined as a dual-positive response against both the receptor-binding domain and nucleocapsid proteins of SARS-CoV-2. Antibodies were measured by immunoprecipitation assays in capillary blood from 15,771 children aged 1 to 18 years living in Bavaria, Germany, and participating in a public health type 1 diabetes screening program (ClinicalTrials.gov: NCT04039945), in 1,916 dried blood spots from neonates in a Bavarian screening study (ClinicalTrials.gov: NCT03316261), and in 75 SARS-CoV-2-positive individuals. Virus positive incidence was obtained from the Bavarian health authority data., Findings: Dual-antibody positivity was detected in none of the 3,887 children in 2019 (100% specificity) and 73 of 75 SARS-CoV-2-positive individuals (97.3% sensitivity). Antibody surveillance in children during 2020 resulted in frequencies of 0.08% in January to March, 0.61% in April, 0.74% in May, 1.13% in June, and 0.91% in July. Antibody prevalence from April 2020 was 6-fold higher than the incidence of authority-reported cases (156 per 100,000 children), showed marked variation between the seven Bavarian regions (p < 0.0001), and was not associated with age or sex. Transmission in children with virus-positive family members was 35%. 47% of positive children were asymptomatic. No association with type 1 diabetes autoimmunity was observed. Antibody frequency in newborns was 0.47%., Conclusions: We demonstrate the value of population-based screening programs for pandemic monitoring., Funding: The work was supported by funding from the BMBF (FKZ01KX1818)., Competing Interests: The authors declare no competing interests., (© 2020 Elsevier Inc.)

Type 1 diabetes (T1D) is still considered a huge burden because the available treatments are not effective in preventing the onset or progression of the disease. Recently, the idea that diabetes is an autoimmune disease mediated exclusively by T cells has been reshaped. In fact, T cells are not the only players with an active role in beta cell destruction. Macrophages and neutrophils, which physiologically reside in pancreatic tissue, can also participate in tissue homeostasis and damage by promoting innate immune responses and modulating inflammation. During the development of the pancreatic islet inflammation there is a strong interplay of both adaptive and innate immune cells, and the presence of innate immune cells has been demonstrated both in exocrine and endocrine pancreatic compartments during the earliest stages of insulitis. Innate immune cell populations secrete cytokines, which must be considered both as physiological and pathological mediators. In fact, it has been demonstrated that cytokines could regulate directly and indirectly insulin secretion and, simultaneously, trigger inflammatory reaction. Indeed, cytokines pathways could represent targets both to improve glucose metabolism and to prevent autoimmune damage. Concordantly, the combination of immunomodulatory strategies against both innate and adaptive immunity should be tested in the next future, as they can be more efficient to prevent or delay islet damage and T1D onset., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Citro, Campo, Dugnani and Piemonti.)

Aim: The aim of this study was to investigate whether treatment with rapamycin plus vildagliptin restores β-cell function in patients with long-standing type 1 diabetes., Methods: A phase 2, single-center, randomized, double-blind, placebo-controlled study was conducted in long-standing type 1 diabetes patients randomly assigned (1:1:1) to 4 weeks of rapamycin (group 2), 4 weeks of rapamycin plus 12 weeks of vildagliptin (group 3), or double placebo (group 1). The primary outcome was the proportion of participants with a positive response to the Mixed-Meal Tolerance Test (C-peptide at 90 minutes > 0.2 nmol/L) at weeks 4 and 12. Secondary end points included insulin requirement, standard measures of glycemic control, and hormonal and immunological profile., Results: Fifty-five patients were randomly assigned to group 1 (n = 18), group 2 (n = 19), or group 3 (n = 18). No patient in any group showed a positive C-peptide response, and there was no significant difference at 4 and 12 weeks for the primary outcome. At 4 weeks, insulin requirement decreased from 0.54 to 0.48 U/kg/day in group 2 (P = .013), from 0.59 to 0.51 U/kg/day in group 3 (P < .001), whereas it did not change in group 1. At 12 weeks, glycated hemoglobin significantly decreased both in group 2 (from 7.3% [56 mmol/mol] to 7% [53 mmol/mol]; P = .045] and in group 3 (from 7.2% [55.5 mmol/mol] to 6.9% [52 mmol/mol]; P = .001]. Rapamycin treatment was associated with a decrease in insulin antibody titer and changes in hormonal/immunological profile., Conclusions: Rapamycin reduced insulin requirement, but did not restore β-cell function in patients with long-standing type 1 diabetes., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objective: Type 1 diabetes (T1D) is characterised by islet autoimmunity and beta cell destruction. A gut microbiota-immunological interplay is involved in the pathophysiology of T1D. We studied microbiota-mediated effects on disease progression in patients with type 1 diabetes using faecal microbiota transplantation (FMT)., Design: Patients with recent-onset (<6 weeks) T1D (18-30 years of age) were randomised into two groups to receive three autologous or allogenic (healthy donor) FMTs over a period of 4 months. Our primary endpoint was preservation of stimulated C peptide release assessed by mixed-meal tests during 12 months. Secondary outcome parameters were changes in glycaemic control, fasting plasma metabolites, T cell autoimmunity, small intestinal gene expression profile and intestinal microbiota composition., Results: Stimulated C peptide levels were significantly preserved in the autologous FMT group (n=10 subjects) compared with healthy donor FMT group (n=10 subjects) at 12 months. Small intestinal Prevotella was inversely related to residual beta cell function (r=-0.55, p=0.02), whereas plasma metabolites 1-arachidonoyl-GPC and 1-myristoyl-2-arachidonoyl-GPC levels linearly correlated with residual beta cell preservation (rho=0.56, p=0.01 and rho=0.46, p=0.042, respectively). Finally, baseline CD4 +CXCR3+T cell counts, levels of small intestinal Desulfovibrio piger and CCL22 and CCL5 gene expression in duodenal biopsies predicted preserved beta cell function following FMT irrespective of donor characteristics., Conclusion: FMT halts decline in endogenous insulin production in recently diagnosed patients with T1D in 12 months after disease onset. Several microbiota-derived plasma metabolites and bacterial strains were linked to preserved residual beta cell function. This study provides insight into the role of the intestinal gut microbiome in T1D., Trial Registration Number: NTR3697., Competing Interests: Competing interests: MN and WMDV are founders and in the Scientific Advisory Board of Caelus Health, the Netherlands. WMDV is Founder and in the Scientific Advisory Board of A-Mansia, Belgium. MN is in the Scientific Advisory Board of Kaleido Biosciences, USA., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY. Published by BMJ.)

Objective: Reparixin is an inhibitor of CXCR1/2 chemokine receptor shown to be an effective anti-inflammatory adjuvant in a pilot clinical trial in allotransplant recipients., Research Design and Methods: A phase 3, multicenter, randomized, double-blind, parallel-assignment study (NCT01817959) was conducted in recipients of islet allotransplants randomized (2:1) to reparixin or placebo in addition to immunosuppression. Primary outcome was the area under the curve (AUC) for C-peptide during the mixed-meal tolerance test at day 75 ± 5 after the first and day 365 ± 14 after the last transplant. Secondary end points included insulin independence and standard measures of glycemic control., Results: The intention-to-treat analysis did not show a significant difference in C-peptide AUC at both day 75 (27 on reparixin vs. 18 on placebo, P = 0.99) and day 365 (24 on reparixin vs. 15 on placebo, P = 0.71). There was no statistically significant difference between treatment groups at any time point for any secondary variable. Analysis of patient subsets showed a trend for a higher percentage of subjects retaining insulin independence for 1 year after a single islet infusion in patients receiving reparixin as compared with patients receiving placebo (26.7% vs. 0%, P = 0.09) when antithymocyte globulin was used as induction immunosuppression., Conclusions: In this first double-blind randomized trial, islet transplantation data obtained with reparixin do not support a role of CXCR1/2 inhibition in preventing islet inflammation-mediated damage., (© 2020 by the American Diabetes Association.)

Autoantibodies (AAbs) are a hallmark of Type 1 diabetes (T1D). Alterations in the frequency and phenotype of follicular helper (Tfh) T cells have been previously documented in patients with type 1 diabetes (T1D), but the contribution of follicular regulatory T (Treg) cells, which are responsible for suppressing AAb development, is less clear. Here, we investigated the frequency and activation status of follicular (CXCR5 + ) and conventional (CXCR5 - ) Treg cells in the blood of children with new-onset T1D, and children with risk for developing T1D (AAb-positive) and compared them to AAb-negative controls. Blood follicular and conventional Treg cells were higher in frequency in children with new onset T1D, but expressed reduced amounts of PD-1 as compared to AAb-negative children. Interestingly, the proportion of circulating FOXP3 + Tregs expressing PD-1 was also reduced in AAb-positive at-risk children as compared to AAb-negative controls, suggesting its potential use as a biomarker of disease progression. Follicular Treg cells were reduced in frequency in the spleens of prediabetic NOD mice as they became older and turned diabetic. Interestingly, PD-1 expression declined also on circulating follicular and conventional Treg cells in prediabetic NOD mice as they aged. Together, these findings show that the frequency of circulating follicular and conventional Treg cells and their levels of PD-1 change with disease progression in children at-risk for developing T1D and in NOD mice., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The early stages of type 1 diabetes (T1D) are characterized by local autoimmune inflammation and progressive loss of insulin-producing pancreatic β cells. Here we show that exposure to proinflammatory cytokines reveals a marked plasticity of the β-cell regulatory landscape. We expand the repertoire of human islet regulatory elements by mapping stimulus-responsive enhancers linked to changes in the β-cell transcriptome, proteome and three-dimensional chromatin structure. Our data indicate that the β-cell response to cytokines is mediated by the induction of new regulatory regions as well as the activation of primed regulatory elements prebound by islet-specific transcription factors. We find that T1D-associated loci are enriched with newly mapped cis-regulatory regions and identify T1D-associated variants disrupting cytokine-responsive enhancer activity in human β cells. Our study illustrates how β cells respond to a proinflammatory environment and implicate a role for stimulus response islet enhancers in T1D.

The blockade of pro-inflammatory mediators is a successful approach to improve the engraftment after islet transplantation. L-aptamers are chemically synthesized, nonimmunogenic bio-stable oligonucleotides that bind and inhibit target molecules conceptually similar to antibodies. We aimed to evaluate if blockade-aptamer-based inhibitors of C-C Motif Chemokine Ligand 2/monocyte chemoattractant protein-1 (CCL2/MCP-1) and C-X-C Motif Chemokine Ligand 12/stromal cell-derived factor-1 (CXCL12/SDF-1) are able to favor islet survival in mouse models for islet transplantation and for type 1 diabetes. We evaluated the efficacy of the CCL2-specific mNOX-E36 and the CXCL12-specific NOX-A12 on islet survival in a syngeneic mouse model of intraportal islet transplantation and in a multiple low doses of streptozotocin (MLD-STZ) diabetes induction model. Moreover, we characterized intrahepatic infiltrated leukocytes by flow cytometry before and 3 days after islet infusion in presence or absence of these inhibitors. The administration for 14 days of mNOX-E36 and NOX-A12 significantly improved islet engraftment, either compound alone or in combination. Intrahepatic islet transplantation recruited CD45 + leucocytes and more specifically CD45+/CD11b+ mono/macrophages; mNOX-E36 and NOX-A12 treatments significantly decreased the recruitment of inflammatory monocytes, CD11b + /Ly6C high /CCR2 + and CD11b + /Ly6C high /CXCR4 + cells, respectively. Additionally, both L-aptamers significantly attenuated diabetes progression in the MLD-STZ model. In conclusion, CCL2/MCP-1 and CXCL12/SDF-1 blockade by L-aptamers is an efficient strategy to improve islet engraftment and survival., (© 2019 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Low-carb and ketogenic diets are popular among clinicians and patients, but the appropriateness of reducing carbohydrates intake in obese patients and in patients with diabetes is still debated. Studies in the literature are indeed controversial, possibly because these diets are generally poorly defined; this, together with the intrinsic complexity of dietary interventions, makes it difficult to compare results from different studies. Despite the evidence that reducing carbohydrates intake lowers body weight and, in patients with type 2 diabetes, improves glucose control, few data are available about sustainability, safety and efficacy in the long-term. In this review we explored the possible role of low-carb and ketogenic diets in the pathogenesis and management of type 2 diabetes and obesity. Furthermore, we also reviewed evidence of carbohydrates restriction in both pathogenesis of type 1 diabetes, through gut microbiota modification, and treatment of type 1 diabetes, addressing the legitimate concerns about the use of such diets in patients who are ketosis-prone and often have not completed their growth., Competing Interests: The authors declare no conflict of interest.

Islet transplantation is superior to extrinsic insulin supplementation in the treating severe Type 1 diabetes. However, its efficiency and longevity are limited by substantial islet loss post-transplantation due to lack of engraftment and vascular supply. To overcome these limitations, we developed a novel approach to bio-fabricate functional, vascularized islet organs (VIOs) ex vivo. We endothelialized acellular lung matrixes to provide a biocompatible multicompartment scaffold with an intact hierarchical vascular tree as a backbone for islet engraftment. Over seven days of culture, islets anatomically and functionally integrated into the surrounding bio-engineered vasculature, generating a functional perfusable endocrine organ. When exposed to supra-physiologic arterial glucose levels in vivo and ex vivo, mature VIOs responded with a physiologic insulin release from the vein and provided more efficient reduction of hyperglycemia compared to intraportally transplanted fresh islets. In long-term transplants in diabetic mice, subcutaneously implanted VIOs achieved normoglycemia significantly faster and more efficiently compared to islets that were transplanted in deviceless fashion. We conclude that ex vivo bio-fabrication of VIOs enables islet engraftment and vascularization before transplantation, and thereby helps to overcome limited islet survival and function observed in conventional islet transplantation. Given recent progress in stem cells, this technology may enable assembly of functional personalized endocrine organs., (Copyright © 2019. Published by Elsevier Ltd.)

Background: Results in murine and nonhuman primate suggested that the bone marrow (BM) might be an alternative site for pancreatic islet transplantation., Methods: We report the results of 2 clinical studies in patients with type 1 diabetes receiving an intra-BM allogeneic islet transplantation: a feasibility study in patients with hepatic contraindications for liver islet allotransplantation receiving a single intra-BM islet infusion (n = 4) and a pilot randomized trial (1:1 allocation using blocks of size 6) in which patients were randomized to receive islets into either the liver (n = 6) or BM (n = 3) to evaluate islet transplant function and survival., Results: We observed no adverse events related to the intrabone injection procedure or the presence of islets in the BM. None of the recipient of an intra-BM allogeneic islet transplantation had a primary nonfunction, as shown by measurable posttransplantation C-peptide levels and histopathological evidence of insulin-producing cells or molecular markers of endocrine tissue in BM biopsy samples collected during follow-up. All patients receiving islets in the BM except 1 lost islet function during the first 4 months after infusion (2 with an early graft loss). Based on biopsies and immunomonitoring, we concluded that the islet loss was primarily caused by the recurrence of autoimmunity., Conclusions: Bone marrow is not a suitable alternative site for pancreatic islet allotransplantation in patients with type 1 diabetes.

Context: Recent studies have suggested that influenza A virus (IAV) might be involved in the etiology of diabetes., Objective and Methods: To address this question, we tested the ability of H1N1 pandemic IAV to infect, replicate, and damage human β cells/pancreatic islets in vitro and induce pancreatic damage and/or glucose metabolism alterations in chemical and autoimmune models of β cell damage in vivo. Moreover, we looked for direct and/or indirect evidence of correlation between IAV infection and autoimmunity/diabetes in humans., Results: Human H1N1 A/California/2009-derived viruses infected human pancreatic islets in vitro, inducing a proinflammatory response associated with substantial increases of CXCL9 and CXCL10 release. In vivo, infected mice showed a clear susceptibility to the virus, with its localization also found in extrapulmonary organs, including the pancreas. Infection was able to induce mild modifications of glycemia in C57B6 mice after chemical damage of islets but did not modulate the autoimmune damage of islets in NOD mice. One of 69 nasopharyngeal swabs collected from patients at the onset of type 1 diabetes yielded positive results for IAV. Pancreas sections from 17 organ donors available from the Network for Pancreatic Organ Donors With Diabetes showed the persistence of CXCL10-positive cells in islet autoimmunity-positive subjects; however, extremely rare cells stained for viral RNA and not preferentially in autoimmune subjects., Conclusion: Influenza H1N1 pdm strains are able to infect and replicate in mammalian pancreatic cells both in vitro and in vivo but did not cause any functional impairment consistent with diabetes.

Stem cells represent a potential candidate for β cell replacement in type 1 diabetes. Pluripotent stem cells are able to differentiate in vitro into functional insulin producing cells, that can restore normoglycemia in diabetic mice. Clinical trials with embryonic stem cell-derived pancreatic progenitors are ongoing. Besides, induced pluripotent stem cells offer the chance of personalized cell therapy. So far, transition to the clinic still needs to face critical issues, such as immunogenicity and safety of stem cell derived β cells. To this purpose, new strategies for immunoprotection, including micro and macro-encapsulation, but also gene editing approaches, are being explored., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Beta cell replacement has the potential to restore euglycemia in patients with insulin-dependent diabetes. Although great progress has been made in establishing allogeneic islet transplantation from deceased donors as the standard of care for those with the most labile diabetes, it is also clear that the deceased donor organ supply cannot possibly treat all those who could benefit from restoration of a normal beta cell mass, especially if immunosuppression were not required. Against this background, the International Pancreas and Islet Transplant Association in collaboration with the Harvard Stem Cell Institute, the Juvenile Diabetes Research Foundation (JDRF), and the Helmsley Foundation held a 2-day Key Opinion Leaders Meeting in Boston in 2016 to bring together experts in generating and transplanting beta cells derived from stem cells. The following summary highlights current technology, recent significant breakthroughs, unmet needs and roadblocks to stem cell-derived beta cell therapies, with the aim of spurring future preclinical collaborative investigations and progress toward the clinical application of stem cell-derived beta cells.

Background: The immunosuppressive drug rapamycin may influence insulin sensitivity in insulin-responsive tissues., Aims: This study aimed at evaluating the effectiveness of rapamycin pre-treatment before pancreatic islet allotransplantation (ITx) in patients with type 1 diabetes mellitus (T1DM)., Methods: Forty-one T1DM patients were studied. Thirteen patients with poor glycemic control underwent a short-term rapamycin treatment before ITx (Group 1), and they were compared to 28 patients undergoing ITx without rapamycin pre-treatment (Group 2). Outcomes were daily insulin requirement (DIR), fasting blood glucose, HbA1c, C-peptide and the SUITO index of beta-cell function. A subgroup of patients pre-treated with rapamycin before ITx underwent euglycemic hyperinsulinemic clamp with [6,6- 2 H 2 ] glucose before and after ITx to evaluate insulin sensitivity., Results: We found a significant reduction in DIR after rapamycin pre-treatment (- 8 ± 6 U/day, mean ± SD, p < 0.001) and 1 year after ITx. DIR reduction 1 year after ITx was greater in Group 1 as compared to Group 2 (- 37 ± 15 vs. - 19 ± 13 U/day, p = 0.005) and remained significant after adjusting for gender, age, glucose and baseline HbA1c (beta = 18.2 ± 5.9, p = 0.006). Fasting glucose and HbA1c significantly decreased 1 year after ITx in Group 1 (HbA1c: - 2.1 ± 1.4%, p = 0.002), while fasting C-peptide (+0.5 ± 0.3 nmol/l, p = 0.002) and SUITO index increased (+57.4 ± 39.7, p = 0.016), without differences between the two groups. Hepatic glucose production decreased after rapamycin pre-treatment (- 1.1 ± 1.1 mg/kg/min, p = 0.04) and after ITx (- 1.6 ± 0.6 mg/kg/min, p = 0.015), while no changes in peripheral glucose disposal were observed., Conclusions: Rapamycin pre-treatment before ITx succeeds in reducing insulin requirement, enhancing hepatic insulin sensitivity. This treatment may improve short-term ITx outcomes, possibly in selected patients with T1DM complicated by insulin resistance., Clinical Trial: Clinicaltrials.gov NCT01060605; NCT00014911.

Aims: Glutamate decarboxylase (GAD) antibodies are the most widely used predictive marker for Type 1 diabetes, but many individuals currently found to be GAD antibody-positive are unlikely to develop diabetes. We have shown previously that radioimmunoassays using N-terminally truncated 35 S-GAD 65 (96-585) offer better disease specificity with similar sensitivity to full-length 35 S-GAD 65 (1-585). To determine whether assay performance could be improved further, we evaluated a more radically truncated 35 S-GAD 65 (143-585) radiolabel., Methods: Samples from people with recent-onset Type 1 diabetes (n = 157) and their first-degree relatives (n = 745) from the Bart's-Oxford family study of childhood diabetes were measured for GAD antibodies using 35 S-labelled GAD 65 (143-585). These were screened previously using a local radioimmunoassay with 35 S-GAD 65 (1-585). A subset was also tested by enzyme-linked immunosorbent assay (ELISA), which performs well in international workshops, but requires 10 times more serum. Results were compared with GAD antibody measurements using 35 S-GAD 65 (1-585) and 35 S-GAD 65 (96-585)., Results: Sensitivity of GAD antibody measurement was maintained using 35 S-GAD 65 (143-585) compared with 35 S-GAD 65 (1-585) and 35 S-GAD 65 (96-585). Specificity for Type 1 diabetes was improved compared with 35 S-GAD 65 (1-585), but was similar to 35 S-GAD 65 (96-585). Relatives found to be GAD antibody-positive using these truncated labels were at increased risk of diabetes progression within 15 years, compared with those positive for GAD(1-585) antibody only, and at similar risk to those found GAD antibody-positive by ELISA., Conclusions: The first 142 amino acids of GAD 65 do not contribute to epitopes recognized by Type 1 diabetes-associated GAD antibodies. Low-volume radioimmunoassays using N-terminally truncated 35 S-GAD 65 are more specific than those using full-length GAD 65 and offer practical alternatives to the GAD antibody ELISA for identifying children at increased risk of Type 1 diabetes., (© 2018 Diabetes UK.)

Stem memory T cells (Tscm) constitute the earliest developmental stage of memory T cells, displaying stem cell-like properties, such as self-renewal capacity. Their superior immune reconstitution potential has sparked interest in cancer immune therapy, vaccine development, and immune reconstitution, whereas their role in autoimmunity is largely unexplored. Here we show that autoreactive CD8 + Tscm specific for β-cell antigens GAD65, insulin, and IGRP are present in patients with type 1 diabetes (T1D). In vitro, the generation of autoreactive Tscm from naive precursors required the presence of the homeostatic cytokine interleukin-7 (IL-7). IL-7 promotes glucose uptake via overexpression of GLUT1 and upregulation of the glycolytic enzyme hexokinase 2. Even though metabolism depends on glucose uptake, the subsequent oxidation of pyruvate in the mitochondria was necessary for Tscm generation from naive precursors. In patients with T1D, high expression of GLUT1 was a hallmark of circulating Tscm, and targeting glucose uptake via GLUT1 using the selective inhibitor WZB117 resulted in inhibition of Tscm generation and expansion. Our results suggest that autoreactive Tscm are present in patients with T1D and can be selectively targeted by inhibition of glucose metabolism., (© 2018 by the American Diabetes Association.)

Aims: Insulin autoantibodies (IAA) are often the first marker of autoimmunity detected in children in the preclinical phase of type 1 diabetes (T1D). Currently, the vast majority of laboratories adopt the radiobinding micro-assay (RBA) for measuring IAA. Our aim was to replace RBA with a novel non-radioactive IAA Luciferase Immuno Precipitation System (LIPS) assay with improved performance., Methods: We developed (pro)insulin antigens with alternative placements of a NanoLuc™ luciferase reporter (NLuc). Performance in LIPS was evaluated by testing sera from new onset T1D (n = 80), blood donors (n = 123), schoolchildren (n = 186), first-degree relatives (FDRs) from the Bart's Oxford family study (n = 53) and from the Belgian Diabetes Registry (n = 136), coded sera from the Islet Autoantibody Standardization Program (IASP) (T1D n = 50, blood donors n = 90)., Results: IAA LIPS based on B chain-NLuc proinsulin or B chain-NLuc insulin, in which NLuc was fused at the C-terminus of the insulin B chain, required only 2 μL of serum and a short incubation time, showed high concordance with RBA (Spearman r = 0.866 and 0.833, respectively), high assay performance (B chain-NLuc proinsulin ROC-AUC = 0.894 and B chain-NLuc insulin ROC-AUC = 0.916), and an adjusted sensitivity at 95% specificity ranking on par with the best assays submitted to the two most recent IASP workshops. In FDRs, the IAA LIPS showed improved discrimination of progressors to T1D compared to RBA., Conclusions: We established a novel high-performance non-radioactive IAA LIPS that might replace the current gold standard RBA and find wide application in the study of the IAA response in T1D.

Background: The advancement of knowledge in the field of regenerative medicine is increasing the therapeutic expectations of patients and clinicians on cell therapy approaches. Within these, stem cell therapies are often evoked as a possible therapeutic option for diabetes, already ongoing or possible in the near future., Aim: The purpose of this document is to make a point of the situation on existing knowledge and therapies with stem cells to treat patients with diabetes by focusing on some of the aspects that most frequently raise curiosity and discussion in clinical practice and in the interaction with the patient. In fact, at present there are no clinically approved treatments based on the use of stem cells for the treatment of diabetes, but several therapeutic approaches have already been evaluated or are being evaluated in clinical trials., Data Synthesis: It is possible to identify three large potential application fields: 1) the reconstruction of the β cell mass; 2) the immunomodulation in type 1 diabetes (T1D); 3) the treatment of complications. In this study we will limit the discussion to approaches that have the potential for clinical translation, deliberately omitting aspects of basic biology and preclinical data. Also, we intentionally omit the treatment of the complications that will be the subject of a future document. Finally, an overview of the Italian situation regarding the storage of cord blood cells for the therapy of diabetes will be given., (Copyright © 2017 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.)

Context: Increasing evidences suggest a correlation between gut and type 1 diabetes (T1D)., Objective: The objective of this study is to evaluate the gut inflammatory profile and microbiota in patients with T1D compared with healthy control (CTRL) subjects and patients with celiac disease (CD) as gut inflammatory disease controls., Design/setting/participants: The inflammatory status and microbiome composition were evaluated in biopsies of the duodenal mucosa of patients with T1D (n = 19), in patients with CD (n = 19), and CTRL subjects (n = 16) recruited at San Raffaele Scientific Institute, in Milan, Italy, between 2009 and 2015., Main Outcome Measures: Inflammation was evaluated by gene expression study and immunohistochemistry. Microbiome composition was analyzed by 16S ribosomal RNA gene sequencing., Results: An increased expression of CCL13, CCL19, CCL22, CCR2, COX2, IL4R, CD68, PTX3, TNFα, and VEGFA was observed in patients with T1D compared with CTRL subjects and patients with CD. Immunohistochemical analysis confirmed T1D-specific inflammatory status compared with healthy and CD control tissues, mainly characterized by the increase of the monocyte/macrophage lineage infiltration. The T1D duodenal mucosal microbiome results were different from the other groups, with an increase in Firmicutes and Firmicutes/Bacteroidetes ratio and a reduction in Proteobacteria and Bacteroidetes. The expression of genes specific for T1D inflammation was associated with the abundance of specific bacteria in the duodenum., Conclusions: This study shows that duodenal mucosa in T1D presents disease-specific abnormalities in the inflammatory profile and microbiota. Understanding the mechanisms underlying these features is critical to disentangle the complex pathogenesis of T1D and to gain new perspectives for future therapies targeting the intestine., (Copyright © 2017 by the Endocrine Society)

In patients with type 1 diabetes (T1D), pancreatic β cells are destroyed by a selective autoimmune attack and their replacement with functional insulin-producing cells is the only possible cure for this disease. The field of islet transplantation has evolved significantly from the breakthrough of the Edmonton Protocol in 2000, since significant advances in islet isolation and engraftment, together with improved immunosuppressive strategies, have been reported. The main limitations, however, remain the insufficient supply of human tissue and the need for lifelong immunosuppression therapy. Great effort is then invested in finding innovative sources of insulin-producing β cells. One old alternative with new recent perspectives is the use of non-human donor cells, in particular porcine β cells. Also the field of preexisting β cell expansion has advanced, with the development of new human β cell lines. Yet, large-scale production of human insulin-producing cells from stem cells is the most recent and promising alternative. In particular, the optimization of in vitro strategies to differentiate human embryonic stem cells into mature insulin-secreting β cells has made considerable progress and recently led to the first clinical trial of stem cell treatment for T1D. Finally, the discovery that it is possible to derive human induced pluripotent stem cells from somatic cells has raised the possibility that a sufficient amount of patient-specific β cells can be derived from patients through cell reprogramming and differentiation, suggesting that in the future there might be a cell therapy without immunosuppression.

Background: The maintenance or expansion of regulatory T (Treg) cells has a fundamental role in the achievement of immunological tolerance after transplantation. Here we aimed to determine mechanisms of human Treg cell depletion and reconstitution after anti-CD25 monoclonal antibody (mAb) treatment., Methods: Seventeen patients with type 1 diabetes who received pancreatic islet transplantation and anti-CD25 mAb as induction therapy were studied., Results: We observed an almost complete depletion of Treg cells after injection of anti-CD25 mAb. The kinetic of Treg cell depletion did not parallel the disappearance of CD25+ T cells as CD25 is also rapidly downregulated and internalized. Regulatory T cell reconstitution is completed within 6 months posttransplantation and appeared to be driven by IL-7-mediated homeostatic T cell proliferation. Anti-CD25 mAb treatment sensitizes Treg cell to the biological effect of IL-7, possibly rendering more common γc-chain available to interact with CD127. Homeostatic Treg cell proliferation is resistant to the inhibitory effect of rapamycin and FK506 but can be blocked by the presence of mycophenolate mofetil., Conclusions: Our data suggest that a compensatory mechanism of IL-7-mediated homeostatic proliferation can restore the inhibitory network of Treg cell after anti-CD25 induction therapy in islet allotransplantation.

Objective: Islet allotransplantation is a less invasive alternative to surgical pancreas transplantation for Type 1 diabetes, while percutaneous intraportal islet autotransplantation (PIPIAT) is usually performed after pancreatic surgery to prevent diabetes. Our aim was to assess the feasibility, radiological aspects, complications and clinical outcome of PIPIAT following pancreatic surgery for not only chronic pancreatitis but also benign and malignant nodules., Methods: From 2008 to 2012, 41 patients were enrolled for PIPIAT 12-48 h after pancreatic surgery (extended pancreatic surgery for chronic pancreatitis and benign/malignant neoplasms). PIPIAT was performed using a combined ultrasonography and fluoroscopy-guided technique (4-F catheter). PIPIAT feasibility, median follow-up and metabolic (insulin independence rate, graft function based on C-peptide levels) and oncologic outcomes were recorded., Results: PIPIAT was not performed in 7/41 patients (4 cases for an inadequate islet mass, 2 cases for haemodynamic instability and 1 case for islet culture contamination), while it was successfully performed in 34/34 patients. Procedure-related major complications occurred in four patients: two bleedings requiring transfusions, one patient with left portal vein thrombosis and one patient with sepsis. Median follow-up duration was 546 days. Insulin independence was achieved in 15/34 (44%) patients, partial graft function in 16/34 (47%) patients and no function in 3/34 (9%) patients. None of the 17 patients with malignant nodules developed liver metastases during follow-up., Conclusion: PIPIAT, performed under ultrasound and fluoroscopy combined guidance and not requiring immunosuppression, is feasible, with a relatively low complication rate and a better metabolic outcome than allotransplantation., Advances in Knowledge: PIPIAT can prevent pancreatogenic diabetes. Ultrasound is a useful tool for the guidance and monitoring of PIPIAT.

The presence of autoantibodies to multiple-islet autoantigens confers high risk for the development of type 1 diabetes. Four major autoantigens are established (insulin, glutamate decarboxylase, IA2, and zinc transporter-8), but the molecular identity of a fifth, a 38-kDa membrane glycoprotein (Glima), is unknown. Glima antibodies have been detectable only by immunoprecipitation from extracts of radiolabeled islet or neuronal cells. We sought to identify Glima to enable efficient assay of these autoantibodies. Mouse brain and lung were shown to express Glima. Membrane glycoproteins from extracts of these organs were enriched by detergent phase separation, lectin affinity chromatography, and SDS-PAGE. Proteins were also immunoaffinity purified from brain extracts using autoantibodies from the sera of patients with diabetes before SDS-PAGE. Eluates from gel regions equivalent to 38 kDa were analyzed by liquid chromatography-tandem mass spectrometry for protein identification. Three proteins were detected in samples from the brain and lung extracts, and in the immunoaffinity-purified sample, but not in the negative control. Only tetraspanin-7, a multipass transmembrane glycoprotein with neuroendocrine expression, had physical characteristics expected of Glima. Tetraspanin-7 was confirmed as an autoantigen by demonstrating binding to autoantibodies in type 1 diabetes. We identify tetraspanin-7 as a target of autoimmunity in diabetes, allowing its exploitation for diabetes prediction and immunotherapy., (© 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

Although numerous chemokine/chemokine receptor pathways have been described to be implicated in the pathogenesis of type 1 diabetes (T1D), the CXCR1/2 axis has recently been proved to be crucial for leucocyte recruitment involved in insulitis and β cell damage. Multiple strategies blocking the CXCR1/2 pathway are available such as neutralizing antibodies, small molecules and peptide-derived inhibitors. They were firstly and widely used in cancer thanks to their anti-tumorigenic activity and only recently they were tested as a new interventional approach for T1D. As well, CXCR1/2 inhibition has been demonstrated to prevent inflammation- and autoimmunity-mediated damage of the pancreatic islets through inhibiting the migration of CXCR1/2-expressing cells. Among them, neutrophils, macrophages, and, although to a smaller extent, lymphoid cells are the main CXCR1/2-expressing cells. These results supported the active role of the innate immunity in the autoimmune process and opened new interventional approaches for the management of T1D.

In the developed world, pancreatic surgery is becoming more common, with an increasing number of patients developing diabetes because of either partial or total pancreatectomy, with a significant impact on quality of life and survival. Although these patients are expected to consume increasing health care resources in the near future, many aspects of diabetes after pancreatectomy are still not well defined. The treatment of diabetes in these patients takes advantage of the therapies used in type 1 and 2 diabetes; however, no specific guidelines for its management, both immediately after pancreatic surgery or in the long term, have been developed. In this article, on the basis of both the literature and our clinical experience, we address the open issues and discuss the most appropriate therapeutic options for patients with diabetes after pancreatectomy.

Chemokines and their receptors have been associated with or implicated in the pathogenesis of type 1 diabetes (T1D), but the identification of a single specific chemokine/receptor pathway that may constitute a suitable target for the development of therapeutic interventions is still lacking. Here, we used multiple low-dose (MLD) streptozotocin (STZ) injections and the NOD mouse model to investigate the potency of CXCR1/2 inhibition to prevent inflammation- and autoimmunity-mediated damage of pancreatic islets. Reparixin and ladarixin, noncompetitive allosteric inhibitors, were used to pharmacologically blockade CXCR1/2. Transient blockade of said receptors was effective in preventing inflammation-mediated damage in MLD-STZ and in preventing and reversing diabetes in NOD mice. Blockade of CXCR1/2 was associated with inhibition of insulitis and modification of leukocytes distribution in blood, spleen, bone marrow, and lymph nodes. Among leukocytes, CXCR2(+) myeloid cells were the most decreased subpopulations. Together these results identify CXCR1/2 chemokine receptors as "master regulators" of diabetes pathogenesis. The demonstration that this strategy may be successful in preserving residual β-cells holds the potential to make a significant change in the approach to management of human T1D., (© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

Islet transplantation is one of the most promising and effective therapies for restoring normoglycemia in type 1 diabetes (T1D) patients, but islet engraftment is one of the main obstacles hampering long-term success. Monitoring graft loss, caused either by immunological or nonimmunological events, occurring in the first phase after transplantation and at later stages of a patient's life is a very important issue. Among the imaging approaches previously applied, magnetic resonance imaging (MRI) monitoring of islet fate following labeling with superparamagnetic iron oxide agents yielded promising results. The aim of this study was to translate into patients the method of islet labeling and MRI monitoring developed in our preclinical setting and to compare imaging results with graft clinical outcome.Three T1D patients and one nondiabetic patient undergoing autotransplantation following subtotal pancreatectomy received Endorem(®)-labeled islets. Patients were monitored by MRI and metabolically (HbA1c, exogenous insulin requirement, and C-peptide, TEF) at 1, 3, and 7 days following transplantation and once a month up to 10 months. Labeled transplanted islets appeared as hypointense areas scattered within the liver parenchyma, whose absolute number at 24 h after transplantation reflected the labeling efficiency. In patients #1 and #3 with good midterm graft function, MRI follow-up showed an important early loss of hypointense spots followed by a slow and progressive disappearance at later timepoints. Graft loss of function in patient #2 4 weeks after transplantation was associated with the complete disappearance of all hypointense signals. The autotransplanted patient, stably insulin free, showed no significant signal reduction during the first 3 days, followed by loss of spots similar to a patient with good midterm graft function. These results suggest that MRI monitoring of islet transplantation at early time points could represent a meaningful readout for helping in predicting transplant failure or success, but its relevance for mid/long-term islet function assessment appears evanescent.

Type 1 diabetes (T1D) is caused by the chronic autoimmune destruction of insulin producing beta cells. Beta cell replacement therapy through whole pancreas or islet transplantation is a therapeutic option for patients in which a stable glucose control is not achievable with exogenous insulin therapy. Long-term insulin independence is, however, hampered by the recipient immune response that includes activation of inflammatory pathways and specific allo- and autoimmunity. The identification and monitoring of soluble and cellular biomarkers are of critical relevance for the prediction of graft damage, for the evaluation of responses to immune-modulating therapy, and for target pathways identification to generate novel drugs or therapeutic approaches. The final objective of immune monitoring is to find ways to improve the outcome of pancreas and islet transplantation. In this review, we discuss the available tools to monitor the innate, humoral and cellular responses after islet and pancreas transplantation, and the most relevant findings generated by these measurements.

Background: Our final objective is to develop an adoptive therapy with tolerogenic donor-specific type 1 T regulatory cells for patients with type 1 diabetes undergoing islet transplantation. The achievement of this objective depends on the availability of an immunosuppressive treatment compatible with the survival, function, and expansion of type 1 T regulatory cells., Methods: For this purpose, we designed a single-group, phase 1 to 2 trial with an immunosuppression protocol including: (i) rapamycin treatment before the first islet infusion (starting ≥ 30 days before transplantation); (ii) induction therapy with anti-thymocyte globulin (ATG) instead of anti-interleukin-2Ra monoclonal antibody (after the first islet infusion only); (iii) short-term treatment with steroids and interleukin-1Ra (right before and for 2 weeks after each infusion); rapamycin+mycophenolate mofetil treatment as maintenance therapy. The target enrollment was 10 patients., Results: Ten of 15 patients who started the pretransplant rapamycin treatment completed it. Nine of 10 patients did not complete the induction therapy with ATG, and three of 10 required adaptation of maintenance immunosuppression caused by side effects. Four of 10 patients acquired insulin independence which can be maintained up to year 3 after last infusion. All six other patients have lost their graft, and the early graft loss was associated with lower dose of ATG during induction., Conclusion: This protocol resulted feasible, safe but less efficient in maintaining graft survival during the time than other T-cell depletion-based protocols. An adequate induction at the first infusion should be considered to improve the overall clinical outcome.

Type 1 diabetes is due to destruction of pancreatic β-cells. Lysine deacetylase inhibitors (KDACi) protect β-cells from inflammatory destruction in vitro and are promising immunomodulators. Here we demonstrate that the clinically well-tolerated KDACi vorinostat and givinostat revert diabetes in the nonobese diabetic (NOD) mouse model of type 1 diabetes and counteract inflammatory target cell damage by a mechanism of action consistent with transcription factor--rather than global chromatin--hyperacetylation. Weaning NOD mice received low doses of vorinostat and givinostat in their drinking water until 100-120 d of age. Diabetes incidence was reduced by 38% and 45%, respectively, there was a 15% increase in the percentage of islets without infiltration, and pancreatic insulin content increased by 200%. Vorinostat treatment increased the frequency of functional regulatory T-cell subsets and their transcription factors Gata3 and FoxP3 in parallel to a decrease in inflammatory dendritic cell subsets and their cytokines IL-6, IL-12, and TNF-α. KDACi also inhibited LPS-induced Cox-2 expression in peritoneal macrophages from C57BL/6 and NOD mice. In insulin-producing β-cells, givinostat did not upregulate expression of the anti-inflammatory genes Socs1-3 or sirtuin-1 but reduced levels of IL-1β + IFN-γ-induced proinflammatory Il1a, Il1b, Tnfα, Fas, Cxcl2, and reduced cytokine-induced ERK phosphorylation. Further, NF-κB genomic iNos promoter binding was reduced by 50%, and NF-κB-dependent mRNA expression was blocked. These effects were associated with NF-κB subunit p65 hyperacetylation. Taken together, these data provide a rationale for clinical trials of safety and efficacy of KDACi in patients with autoimmune disease such as type 1 diabetes.