Proinflammatory cytokines suppress nonsense-mediated RNA decay to impair regulated transcript isoform processing in pancreatic ß cells.

Introduction: Proinflammatory cytokines are implicated in pancreatic ß cell failure in type 1 and type 2 diabetes and are known to stimulate alternative RNA splicing and the expression of nonsense-mediated RNA decay (NMD) components. Here, we investigate whether cytokines regulate NMD activity and identify transcript isoforms targeted in ß cells. Methods: A luciferase-based NMD reporter transiently expressed in rat INS1(832/13), human-derived EndoC-ßH3, or dispersed human islet cells is used to examine the effect of proinflammatory cytokines (Cyt) on NMD activity. The gain- or loss-of-function of two key NMD components, UPF3B and UPF2, is used to reveal the effect of cytokines on cell viability and function. RNA-sequencing and siRNA-mediated silencing are deployed using standard techniques. Results: Cyt attenuate NMD activity in insulin-producing cell lines and primary human ß cells. These effects are found to involve ER stress and are associated with the downregulation of UPF3B. Increases or decreases in NMD activity achieved by UPF3B overexpression (OE) or UPF2 silencing raise or lower Cyt-induced cell death, respectively, in EndoC-ßH3 cells and are associated with decreased or increased insulin content, respectively. No effects of these manipulations are observed on glucose-stimulated insulin secretion. Transcriptomic analysis reveals that Cyt increases alternative splicing (AS)-induced exon skipping in the transcript isoforms, and this is potentiated by UPF2 silencing. Gene enrichment analysis identifies transcripts regulated by UPF2 silencing whose proteins are localized and/or functional in the extracellular matrix (ECM), including the serine protease inhibitor SERPINA1/α-1-antitrypsin, whose silencing sensitizes ß-cells to Cyt cytotoxicity. Cytokines suppress NMD activity via UPR signaling, potentially serving as a protective response against Cyt-induced NMD component expression. Conclusion: Our findings highlight the central importance of RNA turnover in ß cell responses to inflammatory stress.

Autologous Pancreatic Islet Cell Transplantation Following Pancreatectomy for Pancreas Diseases Other Than Chronic Pancreatitis: A 15-Y Study of the Milan Protocol.

BACKGROUND: Pancreatogenic diabetes, a consequence of pancreatic tissue loss following pancreatectomy, poses a significant challenge for patients undergoing pancreatic surgery. Islet autotransplantation (IAT) offers a promising approach to prevent or alleviate pancreatogenic diabetes, but its application has been limited to individuals with painful chronic pancreatitis. METHODS: This study presents a 15-y clinical experience with the Milan Protocol, which expands IAT after pancreatectomy to a broader spectrum of patients with malignant and nonmalignant pancreatic diseases. The analysis evaluates feasibility, efficacy, and safety of IAT. Modified Igls criteria validated through the arginine test and mixed meal tolerance tests were used to assess long-term metabolic outcomes. RESULTS: Between November 2008 and June 2023, IAT procedures were performed on 114 of 147 candidates. IAT-related complications occurred in 19 of 114 patients (16.7%), with 5 being potentially serious. Patients exhibited sustained C-peptide secretion over the 10-y follow-up period, demonstrating a prevalence of optimal and good beta-cell function. Individuals who underwent partial pancreatectomy demonstrated superior metabolic outcomes, including sustained C-peptide secretion and a reduced risk of developing diabetes or insulin dependence compared with those who underwent total pancreatectomy. For patients who had total pancreatectomy, the quantity of infused islets and tissue volume were identified as critical factors influencing metabolic outcomes. An increased risk of recurrence or progression of baseline diseases was not observed in subjects with neoplasms. CONCLUSIONS: These findings provide valuable insights into the benefits and applications of IAT as a therapeutic option for pancreatogenic diabetes after pancreatic surgery, expanding its potential beyond painful chronic pancreatitis.

LDHB contributes to the regulation of lactate levels and basal insulin secretion in human pancreatic ß cells.

Using 13C6 glucose labeling coupled to gas chromatography-mass spectrometry and 2D 1H-13C heteronuclear single quantum coherence NMR spectroscopy, we have obtained a comparative high-resolution map of glucose fate underpinning ß cell function. In both mouse and human islets, the contribution of glucose to the tricarboxylic acid (TCA) cycle is similar. Pyruvate fueling of the TCA cycle is primarily mediated by the activity of pyruvate dehydrogenase, with lower flux through pyruvate carboxylase. While the conversion of pyruvate to lactate by lactate dehydrogenase (LDH) can be detected in islets of both species, lactate accumulation is 6-fold higher in human islets. Human islets express LDH, with low-moderate LDHA expression and ß cell-specific LDHB expression. LDHB inhibition amplifies LDHA-dependent lactate generation in mouse and human ß cells and increases basal insulin release. Lastly, cis-instrument Mendelian randomization shows that low LDHB expression levels correlate with elevated fasting insulin in humans. Thus, LDHB limits lactate generation in ß cells to maintain appropriate insulin release.

Fat-to-blood recirculation of partially dysfunctional PD-1+CD4 Tconv cells is associated with dysglycemia in human obesity.

Obesity is characterized by the accumulation of T cells in insulin-sensitive tissues, including the visceral adipose tissue (VAT), that can interfere with the insulin signaling pathway eventually leading to insulin resistance (IR) and type 2 diabetes. Here, we found that PD-1+CD4 conventional T (Tconv) cells, endowed with a transcriptomic and functional profile of partially dysfunctional cells, are diminished in VAT of obese patients with dysglycemia (OB-Dys), without a concomitant increase in apoptosis. These cells showed enhanced capacity to recirculate into the bloodstream and had a non-restricted TCRß repertoire divergent from that of normoglycemic obese and lean individuals. PD-1+CD4 Tconv were reduced in the circulation of OB-Dys, exhibited an altered migration potential, and were detected in the liver of patients with non-alcoholic steatohepatitis. The findings suggest a potential role for partially dysfunctional PD-1+CD4 Tconv cells as inter-organ mediators of IR in obese patients with dysglycemic.

Improved Specificity of Glutamate Decarboxylase 65 Autoantibody Measurement Using Luciferase-Based Immunoprecipitation System Assays.

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.

Proinflammatory Cytokines Suppress Nonsense-Mediated RNA Decay to Impair Regulated Transcript Isoform Processing in Pancreatic ß-Cells.

Proinflammatory cytokines are implicated in pancreatic ß-cell failure in type 1 and type 2 diabetes and are known to stimulate alternative RNA splicing and the expression of Nonsense-Mediated RNA Decay (NMD) components. Here, we investigate whether cytokines regulate NMD activity and identify transcript isoforms targeted in ß-cells. A luciferase-based NMD reporter transiently expressed in rat INS1(832/13), human-derived EndoC-ßH3 or dispersed human islet cells is used to examine the effect of proinflammatory cytokines (Cyt) on NMD activity. Gain- or loss-of function of two key NMD components UPF3B and UPF2 is used to reveal the effect of cytokines on cell viability and function. RNA-sequencing and siRNA-mediated silencing are deployed using standard techniques. Cyt attenuate NMD activity in insulin-producing cell lines and primary human ß-cells. These effects are found to involve ER stress and are associated with downregulation of UPF3B. Increases or decreases in NMD activity achieved by UPF3B overexpression (OE) or UPF2 silencing, raises or lowers Cyt-induced cell death, respectively, in EndoC-ßH3 cells, and are associated with decreased or increased insulin content, respectively. No effects of these manipulations are observed on glucose-stimulated insulin secretion. Transcriptomic analysis reveals that Cyt increase alternative splicing (AS)-induced exon skipping in the transcript isoforms, and this is potentiated by UPF2 silencing. Gene enrichment analysis identifies transcripts regulated by UPF2 silencing whose proteins are localized and/or functional in extracellular matrix (ECM) including the serine protease inhibitor SERPINA1/α-1-antitrypsin, whose silencing sensitises ß-cells to Cyt cytotoxicity. Cytokines suppress NMD activity via UPR signalling, potentially serving as a protective response against Cyt-induced NMD component expression. Our findings highlight the central importance of RNA turnover in ß-cell responses to inflammatory stress.

HOMA-IR and the Matsuda Index as predictors of progression to type 1 diabetes in autoantibody-positive relatives.

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.

Allo Beta Cell transplantation: specific features, unanswered questions, and immunological challenge.

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.

Glucose and lactate levels are lower in EUS-aspirated cyst fluid of mucinous vs non-mucinous pancreatic cystic lesions.

BACKGROUND: Distinguishing mucinous (M) pancreatic cystic neoplasms (PCNs) from non-mucinous (NM) is challenging but crucial. Low intracystic glucose level has shown diagnostic tool promise, however further investigation is needed to understand metabolic processes. AIMS: To compare the diagnostic accuracy of intracystic glucose and CEA levels in a large cohort and explore lactate levels as potential marker. METHODS: PCNs≥15 mm which underwent EUS-fine needle aspiration were prospectively enrolled. Glucose, CEA and lactate levels were measured. Diagnostic accuracy for M-PCN diagnosis was evaluated using surgical/cytology reports or multidisciplinary evaluations. RESULTS: 169 PCNs were included (64 % M-PCNs). Median intracystic glucose was significantly lower in M-PCNs (1 mg/dL) compared to NM-PCNs (101 mg/dL); mean intracystic CEA was significantly higher in M-PCNs (152.5 ng/mL) compared to NM-PCNs (0.3 ng/mL). ROC curve analysis revealed best glucose cut-off ≤58 mg/dL (accuracy 93.5 %) and CEA cut-off >2.5 ng/mL (accuracy 90.5 %) for M-PCNs. Intracystic lactates were significantly lower in M-PCNs correlating directly with glucose. Single glucose dosage evidenced best diagnostic accuracy respect markers combination. CONCLUSION: Intracystic glucose demonstrated high diagnostic utility for M-PCNs differentiation, surpassing CEA. Lactate levels correlated with glucose, suggesting their uptake by M-PCNs cells. These findings contribute to a better metabolic landscape understanding glucose use as diagnostic marker.

IL-1ß+ macrophages fuel pathogenic inflammation in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is a lethal disease with high resistance to therapies1. Inflammatory and immunomodulatory signals co-exist in the pancreatic tumour microenvironment, leading to dysregulated repair and cytotoxic responses. Tumour-associated macrophages (TAMs) have key roles in PDAC2, but their diversity has prevented therapeutic exploitation. Here we combined single-cell and spatial genomics with functional experiments to unravel macrophage functions in pancreatic cancer. We uncovered an inflammatory loop between tumour cells and interleukin-1ß (IL-1ß)-expressing TAMs, a subset of macrophages elicited by a local synergy between prostaglandin E2 (PGE2) and tumour necrosis factor (TNF). Physical proximity with IL-1ß+ TAMs was associated with inflammatory reprogramming and acquisition of pathogenic properties by a subset of PDAC cells. This occurrence was an early event in pancreatic tumorigenesis and led to persistent transcriptional changes associated with disease progression and poor outcomes for patients. Blocking PGE2 or IL-1ß activity elicited TAM reprogramming and antagonized tumour cell-intrinsic and -extrinsic inflammation, leading to PDAC control in vivo. Targeting the PGE2-IL-1ß axis may enable preventive or therapeutic strategies for reprogramming of immune dynamics in pancreatic cancer.

The Relevance of Advanced Therapy Medicinal Products in the Field of Transplantation and the Need for Academic Research Access: Overcoming Bottlenecks and Claiming a New Time.

The field of transplantation has witnessed the emergence of Advanced Therapy Medicinal Products (ATMPs) as highly promising solutions to address the challenges associated with organ and tissue transplantation. ATMPs encompass gene therapy, cell therapy, and tissue-engineered products, hold immense potential for breakthroughs in overcoming the obstacles of rejection and the limited availability of donor organs. However, the development and academic research access to ATMPs face significant bottlenecks that hinder progress. This opinion paper emphasizes the importance of addressing bottlenecks in the development and academic research access to ATMPs by implementing several key strategies. These include the establishment of streamlined regulatory processes, securing increased funding for ATMP research, fostering collaborations and partnerships, setting up centralized ATMP facilities, and actively engaging with patient groups. Advocacy at the policy level is essential to provide support for the development and accessibility of ATMPs, thereby driving advancements in transplantation and enhancing patient outcomes. By adopting these strategies, the field of transplantation can pave the way for the introduction of innovative and efficacious ATMP therapies, while simultaneously fostering a nurturing environment for academic research.

Efficacy and Safety of Reparixin in Patients with Severe COVID-19 Pneumonia: A Phase 3, Randomized, Double-Blind Placebo-Controlled Study.

INTRODUCTION: Polymorphonuclear cell influx into the interstitial and bronchoalveolar spaces is a cardinal feature of severe coronavirus disease 2019 (COVID-19), principally mediated by interleukin-8 (IL-8). We sought to determine whether reparixin, a novel IL-8 pathway inhibitor, could reduce disease progression in patients hospitalized with severe COVID-19 pneumonia. METHODS: In this Phase 3, randomized, double-blind, placebo-controlled, multicenter study, hospitalized adult patients with severe COVID-19 pneumonia were randomized 2:1 to receive oral reparixin 1200 mg three times daily or placebo for up to 21 days or until hospital discharge. The primary endpoint was the proportion of patients alive and free of respiratory failure at Day 28, with key secondary endpoints being the proportion of patients free of respiratory failure at Day 60, incidence of intensive care unit (ICU) admission by Day 28 and time to recovery by Day 28. RESULTS: Of 279 patients randomized, 182 received at least one dose of reparixin and 88 received placebo. The proportion of patients alive and free of respiratory failure at Day 28 was similar in the two groups {83.5% versus 80.7%; odds ratio 1.63 [95% confidence interval (CI) 0.75, 3.51]; p = 0.216}. There were no statistically significant differences in the key secondary endpoints, but a numerically higher proportion of patients in the reparixin group were alive and free of respiratory failure at Day 60 (88.7% versus 84.6%; p = 0.195), fewer required ICU admissions by Day 28 (15.8% versus 21.7%; p = 0.168), and a higher proportion recovered by Day 28 compared with placebo (81.6% versus 74.9%; p = 0.167). Fewer patients experienced adverse events with reparixin than placebo (45.6% versus 54.5%), most mild or moderate intensity and not related to study treatment. CONCLUSIONS: This trial did not meet the primary efficacy endpoints, yet reparixin showed a trend toward limiting disease progression as an add-on therapy in COVID-19 severe pneumonia and was well tolerated. TRIAL REGISTRATION: ClinicalTrials.gov: NCT04878055, EudraCT: 2020-005919-51.

Interleukin-7 improves the fitness of regulatory T cells for adoptive transfer.

Adoptive regulatory T-cell (Treg) transfer has emerged as a promising therapeutic strategy for regulating immune responses in organ transplantation, graft versus host disease, and autoimmunity, including Type 1 diabetes. Traditionally, Treg for adoptive therapy have been sorted and expanded in vitro using high doses of IL-2, demonstrating stability and suppressive capabilities. However, limitations in their long-term survival post-infusion into patients have been observed. To address this challenge, we investigated a novel expansion protocol incorporating interleukin-7 (IL-7) alongside the traditional method utilizing IL-2 (referred to as IL-7 method, IL-7M). Our study revealed that naïve Treg express significant levels of CD127 and display robust responsiveness to IL-7, characterized by STAT-5 phosphorylation. Expanding naïve Treg with the IL-7M protocol led to a substantial enrichment of CD45RA+ CD62L+ CD95+ Treg but showing a reduction in the final cell yield and suppressive function. Moreover, Treg expanded with the IL-7M exhibited preserved telomere length and demonstrated enhanced resistance to cytokine withdrawal and fas-mediated apoptosis. When transferred into NSG mice IL-7M-Treg persisted longer and reduced the expansion of T cells, but did not significantly reduce the severity of xenoGvHD. In conclusion, our data demonstrate the feasibility of expanding naïve Treg in the presence of IL-7 to generate a Treg product enriched in poorly differentiated CD45RA+ cells with enhanced survival capabilities.

Fibronectin fragments generated by pancreatic trypsin act as endogenous inhibitors of pancreatic tumor growth.

BACKGROUND: The pancreatic microenvironment has a defensive role against cancer but it can acquire tumor-promoting properties triggered by multiple mechanisms including alterations in the equilibrium between proteases and their inhibitors. The identification of proteolytic events, targets and pathways would set the basis for the design of new therapeutic approaches. METHODS AND RESULTS: Here we demonstrate that spheroids isolated from human and murine healthy pancreas and co-transplanted orthotopically with pancreatic ductal adenocarcinoma (PDAC) in mouse pancreas inhibited tumor growth. The effect was mediated by trypsin-generated fibronectin (FN) fragments released by pancreatic spheroids. Tumor inhibition was observed also in a model of acute pancreatitis associated with trypsin activation. Mass spectrometry proteomic analysis of fragments and mAb against different FN epitopes identified the FN type III domain as responsible for the activity. By inhibiting integrin α5ß1, FAK and FGFR1 signaling, the fragments induced tumor cell detachment and reduced cell proliferation. Consistent with the mutual relationship between the two pathways, FGF2 restored both FGFR1 and FAK signaling and promoted PDAC cell adhesion and proliferation. FAK and FGFR inhibitors additively inhibited PDAC growth in vitro and in orthotopic in vivo models. CONCLUSIONS: This study identifies a novel role for pancreatic trypsin and fibronectin cleavage as a mechanism of protection against cancer by the pancreatic microenvironment. The finding of a FAK-FGFR cross-talk in PDAC support the combination of FAK and FGFR inhibitors for PDAC treatment to emulate the protective effect of the normal pancreas against cancer.

Post hoc analysis of a randomized, double-blind, prospective trial evaluating a CXCR1/2 inhibitor in new-onset type 1 diabetes: endo-metabolic features at baseline identify a subgroup of responders.

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.

Paediatric Wolfram syndrome Type 1: should gonadal dysfunction be part of the diagnostic criteria?

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.