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Background: Closed-loop control of type 1 diabetes is receiving increasing attention due to advancement in glucose sensor and insulin pump technology. Here the function and structure of a class of control algorithms designed to exert control to range, defined as insulin treatment optimizing glycemia within a predefined target range by preventing extreme glucose fluctuations, are studied. Methods: The main contribution of the article is definition of a modular architecture for control to range. Emphasis is on system specifications rather than algorithmic realization. The key system architecture elements are two interacting modules: range correction module, which assesses the risk for incipient hyper- or hypoglycemia and adjusts insulin rate accordingly, and safety supervision module, which assesses the risk for hypoglycemia and attenuates or discontinues insulin delivery when necessary. The novel engineering concept of range correction module is that algorithm action is relative to a nominal open-loop strategy—a predefined combination of basal rate and boluses believed to be optimal under nominal conditions. Results: A proof of concept of the feasibility of our control-to-range strategy is illustrated by using a prototypal implementation tested in silico on patient use cases. These functional and architectural distinctions provide several advantages, including (i) significant insulin delivery corrections are only made if relevant risks are detected; (ii) drawbacks of integral action are avoided, e.g., undershoots with consequent hypoglycemic risks; (iii) a simple linear model is sufficient and complex algorithmic constraints are replaced by safety supervision; and (iv) the nominal profile provides straightforward individualization for each patient. Conclusions: We believe that the modular control-to-range system is the best approach to incremental development, regulatory approval, industrial deployment, and clinical acceptance of closed-loop control for diabetes.

Rationale: We recently demonstrated that the Metabesity factor HMG20A regulates islet beta-cell functional maturity and adaptation to physiological stress such as pregnancy and pre-diabetes. HMG20A also dictates central nervous system (CNS) development via inhibition of the LSD1-CoREST complex but its expression pattern and function in adult brain remains unknown. Herein we sought to determine whether HMG20A is expressed in the adult CNS, specifically in hypothalamic astrocytes that are key in glucose homeostasis and whether similar to islets, HMG20A potentiates astrocyte function in response to environmental cues. Methods: HMG20A expression profile was assessed by quantitative PCR (QT-PCR), Western blotting and/or immunofluorescence in: 1) the hypothalamus of mice exposed or not to either a high-fat diet or a high-fat high-sucrose regimen, 2) human blood leukocytes and adipose tissue obtained from healthy or diabetic individuals and 3) primary mouse hypothalamic astrocytes exposed to either high glucose or palmitate. RNA-seq and cell metabolic parameters were performed on astrocytes treated or not with a siHMG20A. Astrocyte-mediated neuronal survival was evaluated using conditioned media from siHMG20A-Treated astrocytes. The impact of ORY100. An Inhibitor Of The LSD1-CoREST Complex, On Hmg20a Expression, Reactive Astrogliosis And Glucose Metabolism Was Evaluated In Vitro And In Vivo In High-Fat High-Sucrose Fed Mice. Results: We Show That Hmg20a Is Predominantly Expressed In Hypothalamic Astrocytes, The Main Nutrient-Sensing Cell Type Of The Brain. Hmg20a Expression Was Upregulated In Diet-Induced Obesity And Glucose Intolerant Mice, Correlating With Increased Transcript Levels Of Gfap And Il1b Indicative Of Inflammation And Reactive Astrogliosis. Hmg20a Transcript Levels Were Also Increased In Adipose Tissue Of Obese Non-Diabetic Individuals As Compared To Obese Diabetic Patients. Hmg20a Silencing In Astrocytes Resulted In Repression Of Inflammatory, Cholesterol Biogenesis And Epithelial-To-Mesenchymal Transition Pathways Which Are Hallmarks Of Reactive Astrogliosis. Accordingly, Hmg20a Depleted Astrocytes Exhibited Reduced Mitochondrial Bioenergetics And Increased Susceptibility To Apoptosis. Neuron Viability Was Also Hindered In HMG20A-Depleted Astrocyte-Derived Conditioned Media. Ory1001 Treatment Rescued Expression Of Reactive Astrogliosis-Linked Genes In Hmg20a Ablated Astrocytes While Enhancing Cell Surface Area, Gfap Intensity And Stat3 Expression In Healthy Astrocytes, Mimicking The Effect Of Hmg20a. Furthermore, Ory1001 Treatment Protected Against Obesity-Associated Glucose Intolerance In Mice Correlating With A Regression Of Hypothalamic Hmg20a Expression, Indicative Of Reactive Astrogliosis Attenuation With Improved Health Status. Conclusion: Hmg20a Coordinates The Astrocyte Polarization State. Under Physiological Pressure Such As Obesity And Insulin Resistance That Induces Low Grade Inflammation, Hmg20a Expression Is Increased To Induce Reactive Astrogliosis In An Attempt To Preserve The Neuronal Network And Re-Establish Glucose Homeostasis. Nonetheless, A Chronic Metabesity State Or Functional Mutations Will Result In Lower Levels Of Hmg20a, Failure To Promote Reactive Astrogliosis And Increase Susceptibility Of Neurons To Stress-Induced Apoptosis. Such Effects Could Be Reversed By Ory1001 Treatment Both In Vitro And In Vivo, Paving The Way For A New Therapeutic Approach For Type 2 Diabetes Mellitus., The authors are supported by grants from the Consejería de Salud, Fundación Pública Andaluza Progreso y Salud, Junta de Andalucía (PI-0727-2010 and PI-0001-2020 to B.R.G.; PI-0085-2013 to P.I.L.; PI-0006-2016 to E.F.M.; PI-0574-2012 to S.Y.R.Z; PI-0247-2016 to F.J.B.S.), the Consejería de Economía, Innovación y Ciencia (P10.CTS.6359 to B.R.G.; CTS.8081 to E.G.F.), the Ministerio de Economía y Competitividad co-funded by Fondos FEDER (PI10/00871, PI13/00593 and BFU2017-83588-P to B.R.G.; PRE2018-084907 to M.E.M.V.G.; PI13/00309; PI17/01004 to F.J.B.S.; BFU2014-5343-P to J.C.R.; and AGL2017-86927-R to F.M.), Vencer el Cancer (B.R.G), DiabetesCero (B.R.G.) and the Juvenile Diabetes Research Foundation (17-2013-372 and 2-SRA-2019-837-S-B to B.R.G.). E.F.M. was a recipient of a Juan de la Cierva Incorporación Fellowship from the Ministerio de Economía y Competitividad (IJCI-2015-26238). S.Y.R.Z is a recipient of a postdoctoral fellowship from Consejería de Salud, Junta de Andalucía (RH-0070-2013). L.L.N. is supported by a Consejeria de Economia, Conocimiento, Empresas y Universidad postdoctoral fellowship (DOC_00652). F.J.B.S. and E.G.F. are recipients of "Nicolás Monardes" research contracts from Consejería de Salud Junta de Andalucía, (C-0070-2012 and C-0031-2016). A.M.M. is supported by CPII19/00023 and PI18/01590 from the Instituto de Salud Carlos III co-funded by Fondos FEDER. CIBERDEM is an initiative of the Instituto de Salud Carlos III. V.C. is supported by a AECC investigator award.

ObjectiveType 1 diabetes (T1D) is an autoimmune disease caused by the destruction of pancreatic β-cells producing insulin. Both T1D patients and animal models exhibit gut microbiota and mucosa alterations, although the exact cause for these remains poorly understood. We investigated the production of key cytokines controlling gut integrity, the abundance of segmented filamentous bacteria (SFB) involved in the production of these cytokines, and the respective role of autoimmune inflammation and hyperglycaemia.DesignWe used several mouse models of autoimmune T1D as well as mice rendered hyperglycaemic without inflammation to study gut mucosa and microbiota dysbiosis. We analysed cytokine expression in immune cells, epithelial cell function, SFB abundance and microbiota composition by 16S sequencing. We assessed the role of anti-tumour necrosis factor α on gut mucosa inflammation and T1D onset.ResultsWe show in models of autoimmune T1D a conserved loss of interleukin (IL)-17A, IL-22 and IL-23A in gut mucosa. Intestinal epithelial cell function was altered and gut integrity was impaired. These defects were associated with dysbiosis including progressive loss of SFB. Transfer of diabetogenic T-cells recapitulated these gut alterations, whereas induction of hyperglycaemia with no inflammation failed to do so. Moreover, anti-inflammatory treatment restored gut mucosa and immune cell function and dampened diabetes incidence.ConclusionOur results demonstrate that gut mucosa alterations and dysbiosis in T1D are primarily linked to inflammation rather than hyperglycaemia. Anti-inflammatory treatment preserves gut homeostasis and protective commensal flora reducing T1D incidence.

Introduction Type 1 diabetes is heterogeneous in its presentation and progression. Variations in clinical presentation between children and adults, and with ethnic group warrant further study in the UK to improve understanding of this heterogeneity. Early interventions to limit beta cell damage in type 1 diabetes are undergoing evaluation, but recruitment is challenging. The protocol presented describes recruitment of people with clinician-assigned, new-onset type 1 diabetes to understand the variation in their manner of clinical presentation, to facilitate recruitment into intervention studies and to create an open-access resource of data and biological samples for future type 1 diabetes research.Methods and analysis Using the National Institute for Health Research Clinical Research Network, patients >5 years of age diagnosed clinically with type 1 diabetes (and their siblings) are recruited within 6 months of diagnosis. Participants agree to have their clinical, laboratory and demographic data stored on a secure database, for their clinical progress to be monitored using information held by NHS Digital, and to be contacted about additional research, in particular immunotherapy and other interventions. An optional blood sample is taken for islet autoantibody measurement and storage of blood and DNA for future analyses. Data will be analysed statistically to describe the presentation of incident type 1 diabetes in a contemporary UK population.Ethics and dissemination Ethical approval was obtained from the independent NHS Research Ethics Service. Results will be presented at national and international meetings and submitted for publication to peer-reviewed journals.

The dynamic nature of the nuclear envelope (NE) is often underestimated. The NE protects, regulates, and organizes the eukaryote genome and adapts to epigenetic changes and to its environment. The NE morphology is characterized by a wide range of diversity and abnormality such as invagination and blebbing, and it is a diagnostic factor for pathologies such as cancer. Recently, the micronuclei, a small nucleus that contains a full chromosome or a fragment thereof, has gained much attention. The NE of micronuclei is prone to collapse, leading to DNA release into the cytoplasm with consequences ranging from the activation of the cGAS/STING pathway, an innate immune response, to the creation of chromosomal instability. The discovery of those mechanisms has revolutionized the understanding of some inflammation-related diseases and the origin of complex chromosomal rearrangements, as observed during the initiation of tumorigenesis. Herein, we will highlight the complexity of the NE biology and discuss the clinical symptoms observed in NE-related diseases. The interplay between innate immunity, genomic instability, and nuclear envelope leakage could be a major focus in future years to explain a wide range of diseases and could lead to new classes of therapeutics., Authors were/are funded by grants from the Spanish association against the Cancer AECC (Aecc investigator to V.C. INVES20033COMA) and from the Fundación Vencer el Cáncer (to B.R.G. and V.C). B.R.G. was/is funded by grants from Consejería de Salud, Fundación Pública AndaluzaProgreso y Salud, Junta de Andalucía (PI-0727-2010, December 2010 to B.R.G. and PI-0001- 2020, December 2020 to B.R.G.); Consejería de Economía, Innovación y Ciencia, Junta de Andalucía (P10-CTS-6359 to B.R.G.); Ministerio de Economía y Competitividad, Instituto de Salud Carlos III, cofunded by Fondos FEDER (PI10/00871, January 2011and PI13/00593, January 2014 to B.R.G.); Ministerio de Economía y Competitividad, Plan Nacional(BFU2017-83588-P, January 2018 to B.R.G.); JuvenileDiabetes Research Foundation JDRF (17-2013-372, August 2013 and 2-SRA-2019-837-S-B, August 2019 to B.R.G.).

Diabetes is a chronic metabolic disease caused by an absolute or relative deficiency in functional pancreatic β‐cells that leads to defective control of blood glucose. Current treatments for diabetes, despite their great beneficial effects on clinical symptoms, are not curative treatments, leading to a chronic dependence on insulin throughout life that does not prevent the secondary complications associated with diabetes. The overwhelming increase in DM incidence has led to a search for novel antidiabetic therapies aiming at the regeneration of the lost functional β‐cells to allow the re‐establishment of the endogenous glucose homeostasis. Here we review several aspects that must be considered for the development of novel and successful regenerative therapies for diabetes: first, the need to maintain the heterogeneity of islet β‐cells with several subpopulations of β‐cells characterized by different transcriptomic profiles correlating with differences in functionality and in resistance/behavior under stress conditions; second, the existence of an intrinsic islet plasticity that allows stimulus‐mediated transcriptome alterations that trigger the transdifferentiation of islet non‐β‐cells into β‐cells; and finally, the possibility of using agents that promote a fully functional/mature β‐cell phenotype to reduce and reverse the process of dedifferentiation of β‐cells during diabetes., Authors were/are funded by grants from Consejería de Salud, Fundación Pública Andaluza Progreso y Salud, Junta de Andalucía (PI-0727-2010, December 2010 to B.R.G., PI-0085-2013, December 2013 to P.I.L., and PI-0001-2020, December 2020 to B.R.G. and P.I.L.); Consejería de Economía, Innovación y Ciencia, Junta de Andalucía (P10-CTS-6359 to B.R.G.); Ministerio de Economía y Competitividad, Instituto de Salud Carlos III, cofunded by Fondos FEDER (PI10/00871, January 2011 and PI13/00593, January 2014 to B.R.G.); Ministerio de Economía y Competitividad, Plan Nacional (BFU2017-83588-P, January 2018 to B.R.G. and PRE2018-084907, January 2018 to E.M.-V.); Juvenile Diabetes Research Foundation JDRF (17-2013-372, August 2013 and 2-SRA-2019-837-S-B, August 2019 to B.R.G:); and Fundacion Vencer el Cancer (to B.R.G.). Special thanks to ALUSVI (Asociación Lucha y Sonríe por la Vida, Pilas), a local Andalusian association, and the Fundacion DiabetesCero for their unconditional financial support.

The vast majority of type 1 diabetes (T1D) genetic association signals lie in noncoding regions of the human genome. Many have been predicted to affect the expression and secondary structure of long noncoding RNAs (lncRNAs), but the contribution of these lncRNAs to the pathogenesis of T1D remains to be clarified. Here, we performed a complete functional characterization of a lncRNA that harbors a single nucleotide polymorphism (SNP) associated with T1D, namely, Lnc13. Human pancreatic islets harboring the T1D-associated SNP risk genotype in Lnc13 (rs917997*CC) showed higher STAT1 expression than islets harboring the heterozygous genotype (rs917997*CT). Up-regulation of Lnc13 in pancreatic β-cells increased activation of the proinflammatory STAT1 pathway, which correlated with increased production of chemokines in an allele-specific manner. In a mirror image, Lnc13 gene disruption in β-cells partially counteracts polyinosinic-polycytidylic acid (PIC)-induced STAT1 and proinflammatory chemokine expression. Furthermore, we observed that PIC, a viral mimetic, induces Lnc13 translocation from the nucleus to the cytoplasm promoting the interaction of STAT1 mRNA with (poly[rC] binding protein 2) (PCBP2). Interestingly, Lnc13-PCBP2 interaction regulates the stability of the STAT1 mRNA, sustaining inflammation in β-cells in an allele-specific manner. Our results show that the T1D-associated Lnc13 may contribute to the pathogenesis of T1D by increasing pancreatic β-cell inflammation. These findings provide information on the molecular mechanisms by which disease-associated SNPs in lncRNAs influence disease pathogenesis and open the door to the development of diagnostic and therapeutic approaches based on lncRNA targeting., This work was supported by a Research Project Grant from the Basque Department of Health (2015111068) and a Research Grant from Fundación de la Sociedad Española de Diabetes (FSED) to I.S. I.G.-M. and A.O.-G. were supported by Predoctoral Fellowship Grants from the UPV/EHU (Universidad del Pais Vasco/Euskal Herriko Unibertsitatea) and the Basque Department of Education, respectively. A.C.-R. was funded by a Grant from the Spanish Ministry of Science, Innovation and Universities (PGC2018-097573-A-I00). D.L.E. received support from the Fonds National de la Recherche 420 Scientifique (FNRS), Welbio Grant CR-2015A-06. B.R.G. was funded by the Spanish Ministry of Science, Innovation and Universities; the European Fund for Regional Development (FEDER) (Grant BFU2017-83588-P); and the Juvenile Diabetes Research Foundation (Grants 17-2013-372 and 2-SRA-2019-837-S-B).

Global rates of diabetes mellitus are increasing, and treatment of the disease consumes a growing proportion of healthcare spending across the world. Pancreatic β-cells, responsible for insulin production, decline in mass in Type 1 and, to a more limited degree, in Type 2 diabetes. However, the extent and rate of loss in both diseases differs between patients resulting in the need for the development of novel diagnostic tools, which could quantitatively assess changes in mass of β-cells over time and potentially lead to earlier diagnosis and improved treatments. Exendin-4, a potent analogue of glucagon-like-peptide 1 (GLP-1), binds to the receptor GLP-1R, whose expression is enriched in β-cells. GLP-1R has thus been used in the past as a means of targeting probes for a wide variety of imaging modalities to the endocrine pancreas. However, exendin-4 conjugates designed specifically for MRI contrast agents are an under-explored area. In the present work, the synthesis and characterization of an exendin-4-dota(ga)-Gd(III) complex, GdEx, is reported, along with its in vivo behaviour in healthy and in β-cell-depleted C57BL/6J mice. Compared to the ubiquitous probe, [Gd(dota)](-), GdEx shows selective uptake by the pancreas with a marked decrease in accumulation observed after the loss of β-cells elicited by deleting the microRNA processing enzyme, DICER. These results open up pathways towards the development of other targeted MRI contrast agents based on similar chemistry methodology.

International audience; Recombinant interferon alpha is used to treat several immunological, oncological, and infectious diseases,1 and various serious side-effects have been associated with its therapeutic use. We want to bring attention to the development of neuromyelitis optica spectrum disorder (NMOSD) [...]

Type 1 diabetes mellitus (T1DM) is an autoimmune disease that targets the destruction of islet beta-cells resulting in insulin deficiency, hyperglycemia and death if untreated. Despite advances in medical devices and longer-acting insulin, there is still no robust therapy to substitute and protect beta-cells that are lost in T1DM. Attempts to refrain from the autoimmune attack have failed to achieve glycemic control in patients highlighting the necessity for a paradigm shift in T1DM treatment. Paradoxically, beta-cells are present in T1DM patients indicating a disturbed equilibrium between the immune attack and beta-cell regeneration reminiscent of unresolved wound healing that under normal circumstances progression towards an anti-inflammatory milieu promotes regeneration. Thus, the ultimate T1DM therapy should concomitantly restore immune self-tolerance and replenish the beta-cell mass similar to wound healing. Recently the agonistic activation of the nuclear receptor LRH-1/NR5A2 was shown to induce immune self-tolerance, increase beta-cell survival and promote regeneration through a mechanism of alpha-to-beta cell phenotypic switch. This trans-regeneration process appears to be facilitated by a pancreatic anti-inflammatory environment induced by LRH-1/NR5A2 activation. Herein, we review the literature on the role of LRH1/NR5A2 in immunity and islet physiology and propose that a cross-talk between these cellular compartments is mandatory to achieve therapeutic benefits., The research of B.R.G is/was supported by the Juvenile Diabetes Research Foundation (17-2013-372 and 2-SRA-2019-837-S-B), the DiabetesCero Foundation, Vencer el Cancer, ALUSVI, the Consejeria de Salud, Fundacion Publica Andaluza Progreso y Salud, Junta de Andalucia (PI-0727-2010), Consejeria de Economia, Innovacion y Ciencia (P10.CTS.6359) and the Ministerio de Economia y Competidividad co-funded by Fondos FEDER (PI10/00871, PI13/00593 and BFU2017-83588-P).

The Collaborative Working Group “Noma Project Team”., Cell therapy is a progressively growing field that is rapidly moving from preclinical model development to clinical application. Outcomes obtained from clinical trials reveal the therapeutic potential of stem cell-based therapy to deal with unmet medical treatment needs for several disorders with no therapeutic options. Among adult stem cells, mesenchymal stem cells (MSCs) are the leading cell type used in advanced therapies for the treatment of autoimmune, inflammatory and vascular diseases. To date, the safety and feasibility of autologous MSC-based therapy has been established; however, their indiscriminate use has resulted in mixed outcomes in preclinical and clinical studies. While MSCs derived from diverse tissues share common properties depending on the type of clinical application, they markedly differ within clinical trials in terms of efficacy, resulting in many unanswered questions regarding the application of MSCs. Additionally, our experience in clinical trials related to critical limb ischemia pathology (CLI) shows that the therapeutic efficacy of these cells in different animal models has only been partially reproduced in humans through clinical trials. Therefore, it is crucial to develop new research to identify pitfalls, to optimize procedures and to clarify the repair mechanisms used by these cells, as well as to be able to offer a next generation of stem cell that can be routinely used in a cost-effective and safe manner in stem cell-based therapies targeting CLI., The authors are supported by the Fundación Progreso y Salud, Consejería de Salud, Junta de Andalucía; FEDER co-funded grants from Instituto de Salud Carlos III (Red TerCel-Grant RD16/0011/0034 and RD16/0011/0013; PIC18/00010; PI16/00259, PI17/02104, and PI-0272-2017), Fundación Andaluza de I+D and Al-Andalus Biopharma Project (FAID-2018-1); and Juvenile Diabetes Research Foundation, JDRF 2-SRA-2019-837-S-B I to (BS). VC-G is a recipient of a Sara Borrell contract (CD16/00118) from FEDER co-funded grants from Instituto de Salud Carlos III.

Introduction: Insulin resistance has been proposed to accelerate progression to type 1 diabetes (T1D) in antibody positive relatives of affected individuals. We hypothesised that overweight individuals with confirmed T1D would be less likely to present with diabetic ketoacidosis (DKA), signifying an earlier onset of T1D, due to concomitant insulin resistance. Methods: The ADDRESS-2 study recruits incident clinician-assigned T1D cases within 6-months of diagnosis and systematically assesses pancreatic autoimmunity by GAD-65, IA-2 and ZnT8 antibodies. People with at least two positive antibodies were selected to confirm diagnosis of T1D and categorised for adiposity according to BMI (adults) or Z-scores (children). Odds ratios (OR) for presentation with DKA were compared, adjusted for potential confounders and sub-analysed by whether adult or child at recruitment. Results: 31% (969/ 3132) were positive for two or more pancreatic antibodies. Of these 44% (424/969) presented with DKA. The proportions with DKA varied significantly by adiposity: 59% underweight (16/27), 47% normal (280/601), 39% overweight (103/263), 30% obese (19/63) and 40% severely obese (6/15) (p=0.02). When adjusted for age, being overweight or obese was associated with lower risk of DKA in adults (OR 0.58, p=0.006; 0.44, p=0.03, respectively) not children (OR 0.9, p=0.81; 0.51, p=0.12, respectively). Higher adiposity category was associated with higher daily insulin-requirements independent of age, with obesity associated with a 4 unit/day increase (p=0.03) and severe obesity, 11 units/day increase (p=0.008). Conclusion: Adults with T1D are less likely to present with DKA if overweight or obese. Despite smaller proportions of DKA, insulin requirements are higher. These data suggest that, in adults, T1D presentation is unmasked by the insulin resistance of obesity prior to absolute insulin deficiency and ketoacidosis. Disclosure S. Misra: None. A. Kaur: None. I.F. Godsland: None. H.C. Walkey: None. D.G. Johnston: None. N. Oliver: Advisory Panel; Self; Dexcom, Inc., Roche Diabetes Care Health and Digital Solutions, Medtronic. Research Support; Self; Dexcom, Inc., Roche Diabetes Care Health and Digital Solutions. Other Relationship; Self; Novo Nordisk A/S, Takeda Development Centre Europe Ltd..

Background & Aims The exocrine pancreas consists of acinar cells that produce digestive enzymes transported to the intestine through a branched ductal epithelium. Chronic pancreatitis is characterized by progressive inflammation, fibrosis, and loss of acinar tissue. These changes of the exocrine tissue are risk factors for pancreatic cancer. The cause of chronic pancreatitis cannot be identified in one quarter of patients. Here, we investigated how duct dysfunction could contribute to pancreatitis development. Methods The transcription factor Hnf1b, first expressed in pancreatic progenitors, is strictly restricted to ductal cells from late embryogenesis. We previously showed that Hnf1b is crucial for pancreas morphogenesis but its postnatal role still remains unelucidated. To investigate the role of pancreatic ducts in exocrine homeostasis, we inactivated the Hnf1b gene in vivo in mouse ductal cells. Results We uncovered that postnatal Hnf1b inactivation in pancreatic ducts leads to chronic pancreatitis in adults. Hnf1bΔduct mutants show dilatation of ducts, loss of acinar cells, acinar-to-ductal metaplasia, and lipomatosis. We deciphered the early events involved, with down-regulation of cystic disease–associated genes, loss of primary cilia, up-regulation of signaling pathways, especially the Yap pathway, which is involved in acinar-to-ductal metaplasia. Remarkably, Hnf1bΔduct mutants developed pancreatic intraepithelial neoplasia and promote pancreatic intraepithelial neoplasia progression in concert with KRAS. We further showed that adult Hnf1b inactivation in pancreatic ducts is associated with impaired regeneration after injury, with persistent metaplasia and initiation of neoplasia. Conclusions Loss of Hnf1b in ductal cells leads to chronic pancreatitis and neoplasia. This study shows that Hnf1b deficiency may contribute to diseases of the exocrine pancreas and gains further insight into the etiology of pancreatitis and tumorigenesis., Graphical abstract

Background.The use of miRNAs as biomarkers for Type 1 Diabetes (T1D) risk is attractive as T1D is usually diagnosed in front of acute symptoms. As miR-375 is highly expressed in the endocrine pancreas, we postulated that its circulating level might reflect beta cell alterations and might be altered in the blood of T1D patients recently diagnosed.Methods.Sera were obtained from 22 T1D children at onset of the disease, before subcutaneous insulin treatment, and from 10 nondiabetic pediatric controls. MiR-375 seric level was quantified by stem-loop RT-PCR-based assay. MiRNAs regulations in isolated human islets in response to high glucose concentrations were determined by TaqMan Low-Density Array.Results.The abundance of miR-375, among the 410 miRNAs detected in human islets, mirrored its well-established role in rodent islet biology. Upregulated miRNAs targeted genes involved in islet homeostasis and regulation of beta cell mass. Downregulated miRNAs, including miR-375, were involved in pancreas secretion and protein turnover. Seric level of miR-375 was lower in T1D children versus age-matched controls, without any correlations with HbA1c, glycaemia, and number of autoantibodies.Conclusion.Altered circulating level of miR-375 at onset of T1D might be a general biomarker of metabolic alterations and inflammation associated with the disease.

Summary Clonal hemopoiesis driven by leukemia-associated gene mutations can occur without evidence of a blood disorder. To investigate this phenomenon, we interrogated 15 mutation hot spots in blood DNA from 4,219 individuals using ultra-deep sequencing. Using only the hot spots studied, we identified clonal hemopoiesis in 0.8% of individuals under 60, rising to 19.5% of those ≥90 years, thus predicting that clonal hemopoiesis is much more prevalent than previously realized. DNMT3A-R882 mutations were most common and, although their prevalence increased with age, were found in individuals as young as 25 years. By contrast, mutations affecting spliceosome genes SF3B1 and SRSF2, closely associated with the myelodysplastic syndromes, were identified only in those aged >70 years, with several individuals harboring more than one such mutation. This indicates that spliceosome gene mutations drive clonal expansion under selection pressures particular to the aging hemopoietic system and explains the high incidence of clonal disorders associated with these mutations in advanced old age., Graphical Abstract, Highlights • Clonal hemopoiesis is an almost inevitable consequence of aging in humans • Spliceosome gene mutations drove clonal hemopoiesis only in persons aged ≥70 years • NPM1 mutations behave as gatekeepers for leukemogenesis, McKerrell et al. employ ultra-deep sequencing to show that age-related clonal hemopoiesis is much more common than previously realized. They find that clonal hemopoiesis, driven by mutations in spliceosome genes SF3B1 and SRSF2, was noted exclusively in individuals aged 70 years or older and that NPM1 mutations are not seen in association with this phenomenon, endorsing their close association with leukemogenesis.

Cobo-Vuilleumier, Nadia et al., Type 1 diabetes mellitus (T1DM) is due to the selective destruction of islet beta cells by immune cells. Current therapies focused on repressing the immune attack or stimulating beta cell regeneration still have limited clinical efficacy. Therefore, it is timely to identify innovative targets to dampen the immune process, while promoting beta cell survival and function. Liver receptor homologue-1 (LRH-1) is a nuclear receptor that represses inflammation in digestive organs, and protects pancreatic islets against apoptosis. Here, we show that BL001, a small LRH-1 agonist, impedes hyperglycemia progression and the immune-dependent inflammation of pancreas in murine models of T1DM, and beta cell apoptosis in islets of type 2 diabetic patients, while increasing beta cell mass and insulin secretion. Thus, we suggest that LRH-1 agonism favors a dialogue between immune and islet cells, which could be druggable to protect against diabetes mellitus., This work was funded by grants from the Juvenile Diabetes Research Foundation (17-2013-372 to B.R.G.), the Consejeria de Salud, Fundacion Publica Andaluza Progreso y Salud, Junta de Andalucia (PI-0727-2010 to B.R.G. and P10CTS6505 to B.S.), Consejeria de Economia, Innovacion y Ciencia (P10.CTS.6359 to B.R.G.), the Ministerio de Economia y Competidividad co-funded by Fondos FEDER (PI10/00871, PI13/00593, and BFU2017-83588-P to B.R.G.; PI14/01015, RD12/0019/0028, and RD16/0011/0034 to B.S.; PI16/00259 to A.H.) and Deutsche Forschungsgemeinschaft (GRK-1789 ´CEMMA´ and DFG SCHI-505/6-1 to R.S.). Special thanks to the families of the DiabetesCero Foundation that graciously supported this work (to B.R.G.). A.M.M. is a recipient of a Miguel Servet grant (CP14/00105) from the Instituto de Salud Carlos III co-funded by Fondos FEDER whereas E.F.M. is a recipient of a Juan de la Cierva Fellowship. I.G.H.G. is supported by a fellowship from Amarna Therapeutics. In some instances, human islets were procured through the European Consortium for Islet Transplantation funded by Juvenile Diabetes Research Foundation (3-RSC-2016-162-I-X).

International audience; The gut microbiota is essential for the normal function of the gut immune system and microbiota alterations are associated with autoimmune disorders. However, how the gut microbiota prevents autoimmunity in distant organs remains poorly defined. Here we reveal that gut microbiota conditioned innate lymphoid cells (ILCs) induce the expression of mouse β-defensin 14 (mBD14) by pancreatic endocrine cells, preventing autoimmune diabetes in the non-obese diabetic (NOD) mice. MBD14 stimulates, via Toll-like receptor 2, interleukin-4 (IL-4)-secreting B cells that induce regulatory macrophages, which in turn induce protective regulatory T cells. The gut microbiota-derived molecules, aryl hydrocarbon receptor (AHR) ligands and butyrate, promote IL-22 secretion by pancreatic ILCs, which induce expression of mBD14 by endocrine cells. Dysbiotic microbiota and low-affinity AHR allele explain the defective pancreatic expression of mBD14 observed in NOD mice. Our study reveals a yet unidentified crosstalk between ILCs and endocrine cells in the pancreas that is essential for the prevention of autoimmune diabetes development.

International audience; Chronic low grade inflammation is closely linked to obesity-associated insulin resistance. To examine how administration of the anti-inflammatory compound indomethacin, a general cyclooxygenase inhibitor, affected obesity development and insulin sensitivity, we fed obesity-prone male C57BL/6J mice a high fat/high sucrose (HF/HS) diet or a regular diet supplemented or not with indomethacin (±INDO) for 7 weeks. Development of obesity, insulin resistance, and glucose intolerance was monitored, and the effect of indomethacin on glucose-stimulated insulin secretion (GSIS) was measured in vivo and in vitro using MIN6 β-cells. We found that supplementation with indomethacin prevented HF/HS-induced obesity and diet-induced changes in systemic insulin sensitivity. Thus, HF/HS+INDO-fed mice remained insulin-sensitive. However, mice fed HF/HS+INDO exhibited pronounced glucose intolerance. Hepatic glucose output was significantly increased. Indomethacin had no effect on adipose tissue mass, glucose tolerance, or GSIS when included in a regular diet. Indomethacin administration to obese mice did not reduce adipose tissue mass, and the compensatory increase in GSIS observed in obese mice was not affected by treatment with indomethacin. We demonstrate that indomethacin did not inhibit GSIS per se, but activation of GPR40 in the presence of indomethacin inhibited glucose-dependent insulin secretion in MIN6 cells. We conclude that constitutive high hepatic glucose output combined with impaired GSIS in response to activation of GPR40-dependent signaling in the HF/HS+INDO-fed mice contributed to the impaired glucose clearance during a glucose challenge and that the resulting lower levels of plasma insulin prevented the obesogenic action of the HF/HS diet.

Hepatocyte growth factor (HGF) is a mitogen required for β-cell replication during pregnancy. To determine whether HGF/c-Met signaling is required for β-cell regeneration, we characterized mice with pancreatic deletion of the HGF receptor, c-Met (PancMet KO mice), in two models of reduced β-cell mass and regeneration: multiple low-dose streptozotocin (MLDS) and partial pancreatectomy (Ppx). We also analyzed whether HGF administration could accelerate β-cell regeneration in wild-type (WT) mice after Ppx. Mouse islets obtained 7 days post-Ppx displayed significantly increased c-Met, suggesting a potential role for HGF/c-Met in β-cell proliferation in situations of reduced β-cell mass. Indeed, adult PancMet KO mice displayed markedly reduced β-cell replication compared with WT mice 7 days post-Ppx. Similarly, β-cell proliferation was decreased in PancMet KO mice in the MLDS mouse model. The decrease in β-cell proliferation post-Ppx correlated with a striking decrease in D-cyclin levels. Importantly, PancMet KO mice showed significantly diminished β-cell mass, decreased glucose tolerance, and impaired insulin secretion compared with WT mice 28 days post-Ppx. Conversely, HGF administration in WT Ppx mice further accelerated β-cell regeneration. These results indicate that HGF/c-Met signaling is critical for β-cell proliferation in situations of diminished β-cell mass and suggest that activation of this pathway can enhance β-cell regeneration. © 2014 by the American Diabetes Association., This work was supported in part by grants from the National Institutes of Health (DK-067351 and DK-077096) and the Juvenile Diabetes Research Foundation (1-2007-3) to A.G.-O. S.E. was the recipient of a postdoctoral fellowship from the National Institutes of Health T32 Research Training grant (T32DK-07052-32). C.D. was the recipient of a research fellowship from the Lawson Wilkins Pediatric Endocrine Society.

This article is distributed under the terms of the Creative Commons Attribution Noncommercial License.-- et al., The paired box transcription factor Pax8 is critical for development of the eye, thyroid gland as well as the urinary and reproductive organs. In adult, Pax8 overexpression is associated with kidney, ovarian and thyroid tumors and has emerged as a specific marker for these cancers. Recently, Pax8 expression was also reported in human pancreatic islets and in neuroendocrine tumors, identifying Pax8 as a novel member of the Pax family expressed in the pancreas. Herein, we sought to provide a comprehensive analysis of Pax8 expression during pancreogenesis and in adult islets. Immunohistochemical analysis using the most employed Pax8 polyclonal antibody revealed strong nuclear staining in the developing mouse pancreas and in mature human and mouse islets. Astonishingly, Pax8 mRNA in mouse islets was undetectable while human islets exhibited low levels. These discrepancies raised the possibility of antibody cross-reactivity. This premise was confirmed by demonstrating that the polyclonal Pax8 antibody also recognized the islet-enriched Pax6 protein both by Western blotting and immunohistochemistry. Thus, in islets polyclonal Pax8 staining corresponds mainly to Pax6. In order to circumvent this caveat, a novel Pax8 monoclonal antibody was used to re-evaluate whether Pax8 was indeed expressed in islets. Surprisingly, Pax8 was not detected in neither the developing pancreas or in mature islets. Reappraisal of pancreatic neuroendocrine tumors using this Pax8 monoclonal antibody exhibited no immunostaining as compared to the Pax8 polyclonal antibody. In conclusion, Pax8 is not expressed in the pancreas and cast doubts on the value of Pax8 as a pancreatic neuroendocrine tumor marker., This work was supported by grants from the Fundacion Progreso y Salud (to B.R.G.), the Spanish Ministry of Science and Innovation, Instituto de Salud Carlos III (PI10/00871 to B.R.G) and the Juvenile Diabetes Research Foundation (9-2004-384 to E.C.I.T Plan Nacional de I+D+I 2008-2011, ISCIII (PI08/0018) co-funded by Fondos FEDER (to A.R.) and Consejería de Salud, Junta de Andalucía (PI0008 to A.R.).

et al., [Objective]: To establish the role of the transcription factor Pax4 in pancreatic islet expansion and survival in response to physiological stress and its impact on glucose metabolism, we generated transgenic mice conditionally and selectively overexpressing Pax4 or a diabetes-linked mutant variant (Pax4R129W) in β-cells. [Research and Methods]: Glucose homeostasis and β-cell death and proliferation were assessed in Pax4- or Pax4R129W-overexpressing transgenic animals challenged with or without streptozotocin. Isolated transgenic islets were also exposed to cytokines, and apoptosis was evaluated by DNA fragmentation or cytochrome C release. The expression profiles of proliferation and apoptotic genes and β-cell markers were studied by immunohistochemistry and quantitative RT-PCR. [Results]: Pax4 but not Pax4R129W protected animals against streptozotocin-induced hyperglycemia and isolated islets from cytokine-mediated β-cell apoptosis. Cytochrome C release was abrogated in Pax4 islets treated with cytokines. Interleukin-1β transcript levels were suppressed in Pax4 islets, whereas they were increased along with NOS2 in Pax4R129W islets. Bcl-2, Cdk4, and c-myc expression levels were increased in Pax4 islets while MafA, insulin, and GLUT2 transcript levels were suppressed in both animal models. Long-term Pax4 expression promoted proliferation of a Pdx1-positive cell subpopulation while impeding insulin secretion. Suppression of Pax4 rescued this defect with a concomitant increase in pancreatic insulin content. [Conclusions]: Pax4 protects adult islets from stress-induced apoptosis by suppressing selective nuclear factor-κB target genes while increasing Bcl-2 levels. Furthermore, it promotes dedifferentiation and proliferation of β-cells through MafA repression, with a concomitant increase in Cdk4 and c-myc expression., This work was funded by the Swiss National Science Foundation (grants 310030-119763 to B.R.G.; 310000-116750/1 to C.B.W. and B.R.G.; 310000-109402, 310000-122423, and CR32I3-129987 to P.M.; and 3100A0103867 NCCR “Frontiers-in-Genetics” to P.L.H.), the Juvenile Diabetes Research Foundation (grants 7-2005-1158 to C.B.W. and B.R.G.; 40-2011-11 to P.M.; and 26-2007-928 and 26-2008-640 to P.L.H.), the EU (FP-7, BETAIMAGE 222980, IMI IMIDIA, and C2008-T7 to P.M. and FP-6, β Cell Therapy Consortium to P.L.H.), the National Institutes of Health/National Institutes of Diabetes and Digestive and Kidney Diseases (the Beta Cell Biology Consortium to P.L.H.), and the Foundation Progreso y Salud, Junta de Andalucia (to B.R.G.). K.H.H.H. was supported by a GeMet/Novo Nordisk fellowship. No other potential conflicts of interest relevant to this article were reported.

Yeste, Ada et al., Type 1 diabetes (T1D) is a T cell-dependent autoimmune disease that is characterized by the destruction of insulin-producing β cells in the pancreas. The administration to patients of ex vivo-differentiated FoxP3 regulatory T (T) cells or tolerogenic dendritic cells (DCs) that promote T cell differentiation is considered a potential therapy for T1D; however, cell-based therapies cannot be easily translated into clinical practice. We engineered nanoparticles (NPs) to deliver both a tolerogenic molecule, the aryl hydrocarbon receptor (AhR) ligand 2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE), and the β cell antigen proinsulin (NP) to induce a tolerogenic phenotype in DCs and promote T cell generation in vivo. NP administration to 8-week-old nonobese diabetic mice suppressed autoimmune diabetes. NP induced a tolerogenic phenotype in DCs, which was characterized by a decreased ability to activate inflammatory effector T cells and was concomitant with the increased differentiation of FoxP3 T cells. The induction of a tolerogenic phenotype in DCs by NPs was mediated by the AhR-dependent induction of Socs2, which resulted in inhibition of nuclear factor κB activation and proinflammatory cytokine production (properties of tolerogenic DCs). Together, these data suggest that NPs constitute a potential tool to reestablish tolerance in T1D and potentially other autoimmune disorders., This work was supported by grants ES025530, AI075285, and AI093903 from the NIH, 17-2011-371 from the Juvenile Diabetes Research Foundation, the Boston Area Diabetes Endocrinology Research Center (to F.J.Q.), and PI14-1600 from the Spanish Ministry of Economy and Competitiveness (ISCIII) with FEDER cofunding (to D.P.). M.C.T. received support from an educational grant from Mallinckrodt Pharmaceuticals (A113307) and from a postdoctoral fellowship (0571-15-6) from Coordination for the Improvement of Higher Education Personnel, Brazil. I.D.M. received support from an educational grant from Questcor (A219074) and from a postdoctoral fellowship (FG 2036-A1/1) from the National Multiple Sclerosis Society. S.C.K. received support from The Helmsley Charitable Trust.

International audience; Heterozygous mutations in the human HNF1B gene are associated with maturity-onset diabetes of the young type 5 (MODY5) and pancreas hypoplasia. In mouse, Hnf1b heterozygous mutants do not exhibit any phenotype, whereas the homozygous deletion in the entire epiblast leads to pancreas agenesis associated with abnormal gut regionalization. Here, we examine the specific role of Hnf1b during pancreas development, using constitutive and inducible conditional inactivation approaches at key developmental stages. Hnf1b early deletion leads to a reduced pool of pancreatic multipotent progenitor cells (MPCs) due to decreased proliferation and increased apoptosis. Lack of Hnf1b either during the first or the secondary transitions is associated with cystic ducts. Ductal cells exhibit aberrant polarity and decreased expression of several cystic disease genes, some of which we identified as novel Hnf1b targets. Notably, we show that Glis3, a transcription factor involved in duct morphogenesis and endocrine cell development, is downstream Hnf1b. In addition, a loss and abnormal differentiation of acinar cells are observed. Strikingly, inactivation of Hnf1b at different time points results in the absence of Ngn3(+) endocrine precursors throughout embryogenesis. We further show that Hnf1b occupies novel Ngn3 putative regulatory sequences in vivo. Thus, Hnf1b plays a crucial role in the regulatory networks that control pancreatic MPC expansion, acinar cell identity, duct morphogenesis and generation of endocrine precursors. Our results uncover an unappreciated requirement of Hnf1b in endocrine cell specification and suggest a mechanistic explanation of diabetes onset in individuals with MODY5.

Cell therapy and the use of mAbs that interfere with T cell effector functions constitute promising approaches for the control of allograft rejection. In the current study, we investigated a novel approach combining administration of autologous tolerogenic dendritic cells with short-term treatment with CD3-specific Abs. Permanent acceptance of pancreatic islet allografts was achieved in mice treated with the combination therapy the day before transplantation but not in recipients treated with either therapy alone. The combination treatment induced a marked decrease in T cells infiltrating the allografts and a sustained reduction of antidonor responses. Importantly, CD4+Foxp3+ regulatory T cells appeared to play a crucial role in the long-term graft acceptance. Their frequency increased significantly in the spleen, draining lymph nodes, and transplanted islets and remained elevated over the long term; they exhibited increased donor-specific suppressive functions; and their removal at the time of transplantation abrogated the therapeutic effect of the combined therapy. These results support the therapeutic potential of protocols combining autologous dendritic cells and low-dose CD3 Abs, both currently in clinical development, and that act in synergy to control allogeneic immune responses and favor graft survival in a full-mismatch situation.

PMCID: PMC3935450.-- et al., Type 2 diabetes affects over 300 million people, causing severe complications and premature death, yet the underlying molecular mechanisms are largely unknown. Pancreatic islet dysfunction is central in type 2 diabetes pathogenesis, and understanding islet genome regulation could therefore provide valuable mechanistic insights. We have now mapped and examined the function of human islet cis-regulatory networks. We identify genomic sequences that are targeted by islet transcription factors to drive islet-specific gene activity and show that most such sequences reside in clusters of enhancers that form physical three-dimensional chromatin domains. We find that sequence variants associated with type 2 diabetes and fasting glycemia are enriched in these clustered islet enhancers and identify trait-associated variants that disrupt DNA binding and islet enhancer activity. Our studies illustrate how islet transcription factors interact functionally with the epigenome and provide systematic evidence that the dysregulation of islet enhancers is relevant to the mechanisms underlying type 2 diabetes., We thank the DIAGRAM and MAGIC consortia, the Singapore Prospective Study Program, the Singapore Consortium of Cohort Studies, the Singapore Indian Eye Study, the Singapore Malay Eye Study and Y.Y. Teo, E.S. Tai, T.Y. Wong, W.Y. Lim and X. Wang (National University of Singapore; funded by the National Medical Research Council of Singapore, Singapore Translational Researcher Award schemes, the Biomedical Research Council of Singapore and the National Research Foundation (NRF) Fellowship scheme). This work was carried out in part at the Centre Esther Koplowitz. This work was funded by grants from a European Foundation for the Study of Diabetes Lilly fellowship (L. Pasquali), the Ministerio de Economía y Competitividad (SAF2011-27086 to J.F., BFU2010-14839 and CSD2007-00008 to J.L.G.S.), the Innovative Medicines Initiative (DIRECT to M.I.M. and J.F.), the Andalusian Government (CVI-3488 to J.L.G.S.), the Biology of Liver and Pancreatic Development and Disease Marie Curie Initial Training Network (F.M. and J.F.), the Wellcome Trust (090532, 98381 and 090367 to M.I.M., 095101 to A.L.G., 101033 to J.F.), Juvenile Diabetes Research Foundation (31-2012-783 to T.B., F.P. and L. Piemonti) and Framework Programme 7 (HEALTH-F4-2007-201413 to M.I.M.).

Despite widespread usage of β-adrenergic receptor (AR) agonists and antagonists in current clinical practice, our understanding of their interactions with the immune system is surprisingly sparse. Among the AR expressed by dendritic cells (DC), β2-AR can modify in vitro cytokine release upon stimulation. Because DC play a pivotal role in CD8+ T cell immune responses, we examined the effects of β2-AR stimulation on MHC class I exogenous peptide presentation and cross-presentation capacities. We demonstrate that β2-AR agonist-exposed mature DC display a reduced ability to cross-present protein Ags while retaining their exogenous peptide presentation capability. This effect is mediated through the nonclassical inhibitory G (Gαi/0) protein. Moreover, inhibition of cross-presentation is neither due to reduced costimulatory molecule expression nor Ag uptake, but rather to impaired phagosomal Ag degradation. We observed a crosstalk between the TLR4 and β2-AR transduction pathways at the NF-κB level. In vivo, β2-AR agonist treatment of mice inhibits Ag protein cross-presentation to CD8+ T cells but preserves their exogenous MHC class I peptide presentation capability. These findings may explain some side effects on the immune system associated with stress or β-agonist treatment and pave the way for the development of new immunomodulatory strategies.

Background: Genome-wide association studies have identified determinants of chronic obstructive pulmonary disease, asthma, and lung function level; however, none have addressed decline in lung function.Objective: We conducted the first genome-wide association study on the age-related decrease in FEV1 and its ratio to forced vital capacity (FVC) stratified a priori by asthma status.Methods: Discovery cohorts included adults of European ancestry (1,441 asthmatic and 2,677 nonasthmatic participants: the Epidemiological Study on the Genetics and Environment of Asthma, the Swiss Cohort Study on Air Pollution and Lung and Heart Disease in Adults, and the European Community Respiratory Health Survey). The associations of FEV1 and FEV1/FVC ratio decrease with 2.5 million single nucleotide polymorphisms (SNPs) were estimated. Thirty loci were followed up by in silico replication (1,160 asthmatic and 10,858 nonasthmatic participants: Atherosclerosis Risk in Communities, the Framingham Heart Study, the British 1958 Birth Cohort, and the Dutch Asthma Study).Results: Main signals identified differed between asthmatic and nonasthmatic participants. None of the SNPs reached genome-wide significance. The association between the height-related gene DLEU7 and FEV1 decrease suggested for nonasthmatic participants in the discovery phase was replicated (discovery, P = 4.8 x 10(-6); replication, P = .03), and additional sensitivity analyses point to a relation to growth. The top ranking signal, TUSC3, which is associated with FEV1/FVC ratio decrease in asthmatic participants (P = 5.3 x 10(-8)), did not replicate. SNPs previously associated with cross-sectional lung function were not prominently associated with decline.Conclusions: Genetic heterogeneity of lung function might be extensive. Our results suggest that genetic determinants of longitudinal and cross-sectional lung function differ and vary by asthma status. (J Allergy Clin Immunol 2012;129:1218-28.)

International audience; Depletion of blood glutathione (GSH), a key antioxidant, is associated with type 1 diabetes mellitus (T1D) and contributes to the pathophysiology of diabetes complications. The aim of the current study was to determine whether acute normalization of blood glucose would restore GSH kinetics in adolescents with poorly controlled T1D. Ten 16.9 +/- 1.5-year-old (SE) adolescents who had had T1D for 8.5 +/- 1.9 years and were free of complications but were in poor control (hemoglobin A(1c), 9.2% +/- 0.5%) received two 5-hour intravenous infusions of L-[3,3-H-2(2)]cysteine in the postabsorptive state on 2 separate days after blood glucose had been maintained overnight at 246 +/- 24 mg/dL (hyperglycemia) or 118 +/- 23 mg/dL (euglycemia) using intravenous insulin infusion. Blood GSH fractional synthesis rates were determined by mass spectrometry from H-2(2)-cysteine incorporation into GSH. Neither blood GSH (551 +/- 169 vs 541 +/- 232 mu mol/L, P = .629) nor GSH fractional synthesis rate (84% +/- 30% vs 82% +/- 33% d(-1), P = .965) was altered by the short-term change in glycemic control. This finding suggests that, in adolescents with poorly controlled T1D, either (a) blood glucose per se does not regulate GSH metabolism or (b) GSH may only respond to sustained, more chronic changes in blood glucose level. (C) 2012 Elsevier Inc. All rights reserved.

International audience; Synthetic peptides are widely used in immunological research as epitopes to stimulate their cognate T cells. These preparations are never completely pure, but trace contaminants are commonly revealed by mass spectrometry quality controls. In an effort to characterize novel major histocompatibility complex (MHC) Class I-restricted β-cell epitopes in non-obese diabetic (NOD) mice, we identified islet-infiltrating CD8+ T cells recognizing a contaminating peptide. The amount of this contaminant was so small to be undetectable by direct mass spectrometry. Only after concentration by liquid chromatography, we observed a mass peak corresponding to an immunodominant islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)$_{206-214}$ epitope described in the literature. Generation of CD8+ T-cell clones recognizing IGRP$_{206-214}$) using a novel method confirmed the identity of the contaminant, further underlining the immunodominance of IGRP$_{206-214}$. If left undetected, minute impurities in synthetic peptide preparations may thus give spurious results.

et al., Liver receptor homolog (LRH-1) is an orphan nuclear receptor (NR5A2) that regulates cholesterol homeostasis and cell plasticity in endodermal-derived tissues. Estrogen increases LRH-1 expression conveying cell protection and proliferation. Independently, estrogen also protects isolated human islets against cytokine- induced apoptosis. Herein, we demonstrate that LRH-1 is expressed in islets, including β-cells, and that transcript levels are modulated by 17β-estradiol through the estrogen receptor (ER)α but not ERβ signaling pathway. Repression of LRH-1 by siRNA abrogated the protective effect conveyed by estrogen on rat islets against cytokines. Adenoviral-mediated overexpression of LRH-1 in human islets did not alter proliferation but conferred protection against cytokines and streptozotocin-induced apoptosis. Expression levels of the cell cycle genes cyclin D1 and cyclin E1 as well as the antiapoptotic gene bcl-xl were unaltered in LRH-1 expressing islets. In contrast, the steroidogenic enzymes CYP11A1 and CYP11B1 involved in glucocorticoid biosynthesis were both stimulated in transduced islets. In parallel, graded overexpression of LRH-1 dose- dependently impaired glucose-induced insulin secretion. Our results demonstrate the crucial role of the estrogen target gene nr5a2 in protecting human islets against-stressed-induced apoptosis. We postulate that this effect is mediated through increased glucocorticoid production that blunts the pro-inflammatory response of islets. © The Author 2011. Published by Oxford University Press. All rights reserved., This work was supported by grants from the Swiss National Science Foundation (310030-119763 to B.R.G., 310000-116750/1 to C.B.W., 32003B-120376 to D.B.), the Juvenile Diabetes Research Foundation (9-2004-384 to E.C.I.T.), the Fundacion Progreso y Salud (to B.R.G.) and the Junta de Andalucia, Consejeria de Salud (PI-0727-2010 to B.R.G.).

International audience; ABSTRACT : INTRODUCTION : Early randomized clinical trials of autologous bone marrow cardiac stem cell therapy have reported contradictory results highlighting the need for a better evaluation of protocol designs. This study was designed to quantify and compare whole body and heart cell distribution after intracoronary or peripheral intravenous injection of autologous bone marrow mononuclear cells in a porcine acute myocardial infarction model with late reperfusion. METHODS : Myocardial infarction was induced using balloon inflation in the left coronary artery in domestic pigs. At seven days post-myocardial infarction, 1 x 10(8) autologous bone marrow mononuclear cells were labeled with fluorescent marker and/or 99mTc radiotracer, and delivered using intracoronary or peripheral intravenous injection (leg vein). RESULTS : Scintigraphic analyses and Upsilon-emission radioactivity counting of harvested organs showed a significant cell fraction retained within the heart after intracoronary injection (6 +/- 1.7% of injected radioactivity at 24 hours), whereas following peripheral intravenous cell injection, no cardiac homing was observed at 24 hours and cells were mainly detected within the lungs. Importantly, no difference was observed in the percentage of retained cells within the myocardium in the presence or absence of myocardial infarction. Histological evaluation did not show arterial occlusion in both animal groups and confirmed the presence of bone marrow mononuclear cells within the injected myocardium area. CONCLUSIONS : Intravenous bone marrow mononuclear cell injection was ineffective to target myocardium. Myocardial cell distribution following intracoronary injection did not depend on myocardial infarction presence, a factor that could be useful for cardiac cell therapy in patients with chronic heart failure of non-ischemic origin or with ischemic myocardium without myocardial infarction.

OBJECTIVE The response of ventromedial hypothalamic (VMH) glucose-inhibited neurons to decreased glucose is impaired under conditions where the counterregulatory response (CRR) to hypoglycemia is impaired (e.g., recurrent hypoglycemia). This suggests a role for glucose-inhibited neurons in the CRR. We recently showed that decreased glucose increases nitric oxide (NO) production in cultured VMH glucose-inhibited neurons. These in vitro data led us to hypothesize that NO release from VMH glucose-inhibited neurons is critical for the CRR. RESEARCH DESIGN AND METHODS The CRR was evaluated in rats and mice in response to acute insulin-induced hypoglycemia and hypoglycemic clamps after modulation of brain NO signaling. The glucose sensitivity of ventromedial nucleus glucose-inhibited neurons was also assessed. RESULTS Hypoglycemia increased hypothalamic constitutive NO synthase (NOS) activity and neuronal NOS (nNOS) but not endothelial NOS (eNOS) phosphorylation in rats. Intracerebroventricular and VMH injection of the nonselective NOS inhibitor NG-monomethyl-l-arginine (l-NMMA) slowed the recovery to euglycemia after hypoglycemia. VMH l-NMMA injection also increased the glucose infusion rate (GIR) and decreased epinephrine secretion during hyperinsulinemic/hypoglycemic clamp in rats. The GIR required to maintain the hypoglycemic plateau was higher in nNOS knockout than wild-type or eNOS knockout mice. Finally, VMH glucose-inhibited neurons were virtually absent in nNOS knockout mice. CONCLUSIONS We conclude that VMH NO production is necessary for glucose sensing in glucose-inhibited neurons and full generation of the CRR to hypoglycemia. These data suggest that potentiating NO signaling may improve the defective CRR resulting from recurrent hypoglycemia in patients using intensive insulin therapy.

Previous studies have documented that the insulin-producing beta-cells of laboratory rodents are coupled by gap junction channels made solely of the connexin36 (Cx36) protein, and have shown that loss of this protein desynchronizes beta-cells, leading to secretory defects reminiscent of those observed in type 2 diabetes. Since human islets differ in several respects from those of laboratory rodents, we have now screened human pancreas, and islets isolated thereof, for expression of a variety of connexin genes, tested whether the cognate proteins form functional channels for islet cell exchanges, and assessed whether this expression changes with beta-cell function in islets of control and type 2 diabetics. Here, we show that (i) different connexin isoforms are differentially distributed in the exocrine and endocrine parts of the human pancreas; (ii) human islets express at the transcript level different connexin isoforms; (iii) the membrane of beta-cells harbors detectable levels of gap junctions made of Cx36; (iv) this protein is concentrated in lipid raft domains of the beta-cell membrane where it forms gap junctions; (v) Cx36 channels allow for the preferential exchange of cationic molecules between human beta-cells; (vi) the levels of Cx36 mRNA correlated with the expression of the insulin gene in the islets of both control and type 2 diabetics. The data show that Cx36 is a native protein of human pancreatic islets, which mediates the coupling of the insulin-producing beta-cells, and contributes to control beta-cell function by modulating gene expression., The Swiss National Science Foundation (310000-122430 to P.Me), the Juvenile Diabetes Research Foundation (1-2005-1084 to V.C., 1-2007-158 to P.Me), the National Institute of Health (DK55183 to V.C.), the European Union (FP6-Integrated Project EuroDia LSHM-CT-2006-518153 to P.Ma; FP-7 BETAIMAGE 222980 to P.Me), Novo Nordisk (to P.Me) and The Larry L. Hillblom Foundation (to V.C.). Image analysis was performed at The National Center for Microscopy and Imaging Research (NIH grant RR4050 to M. Ellisman). Fresh human islets were provided by the Cell Isolation and Transplantation Center

Allergic asthma is a chronic disease characterized by airway obstruction in response to allergen exposure. It results from an inappropriate T helper type 2 response to environmental airborne antigens and affects 300 million individuals. Its prevalence has increased markedly in recent decades, most probably as a result of changes in environmental factors. Exposure to environmental antigens during infancy is crucial to the development of asthma. Epidemiological studies on the relationship between breastfeeding and allergic diseases have reached conflicting results. Here, we have investigated whether the exposure of lactating mice to an airborne allergen affects asthma development in progeny. We found that airborne antigens were efficiently transferred from the mother to the neonate through milk and that tolerance induction did not require the transfer of immunoglobulins. Breastfeeding-induced tolerance relied on the presence of transforming growth factor (TGF)-beta during lactation, was mediated by regulatory CD4+ T lymphocytes and depended on TGF-beta signaling in T cells. In conclusion, breast milk-mediated transfer of an antigen to the neonate resulted in oral tolerance induction leading to antigen-specific protection from allergic airway disease. This study may pave the way for the design of new strategies to prevent the development of allergic diseases.

International audience; Type 1 diabetes is an immune-mediated disease, in which T cells of the adaptive immune system mediate beta cell destruction. Recently the innate immune system has been linked to etiopathogenesis of several autoimmune diseases including type 1 diabetes, as innate effector cells (e.g. dendritic cells, monocytes/macrophages and NK cells) can prime and promote or regulate (auto)immune responses. We have previously developed an experimental autoimmune diabetes (EAD) model with insulin peptide B:9-23 immunization in transgenic H-2 d mice expressing the costimulatory molecule B7.1 in their islets (under the Rat Insulin Promotor, RIP). We compared the induction of diabetes with polyinosinic–polycytidylic acid (Poly I:C), a mimic of double stranded viral RNA versus insulin B:9-23 peptide in mice following backcrossing of the B7.1 transgene on to BALB/c mice from original B7.1 C57Bl/6 mice. We find that diabetes induction by Poly I:C is C57Bl/6 associated, whereas B:9-23 peptide induced diabetes and induction of insulin autoantibodies (IAA) are dependent on BALB/c genes. This B:9-23 peptide induced diabetes is consistent with MHC class II H-2 d being necessary for the response to this peptide. Of note Poly I:C induction of diabetes was lost while B:9-23 induction was retained with backcrossing to BALB/c mice. Interaction of genes and environment (antigenic epitope and viral mimic) can be important in the pathogenesis of immune mediated diabetes and activation of the innate immune system (e.g. Poly I:C) may be one key determinant.

Subject Areas: carotid arteries; emulsions; fatty acids; gene expression; heparin; hypothalamus; neurons; oxidation.; International audience; Variations in plasma fatty acid (FA) concentrations are detected by FA sensing neurons in specific brain areas such as the hypothalamus. These neurons play a physiological role in the control of food intake and the regulation of hepatic glucose production. Le Foll et al. previously showed in vitro that at least 50% of the FA sensing in ventromedial hypothalamic (VMH) neurons is attributable to the interaction of long chain FA with FA translocase/CD36 (CD36). The present work assessed whether in vivo effects of hypothalamic FA sensing might be partly mediated by CD36 or intracellular events such as acylCoA synthesis or β-oxidation. To that end, a catheter was implanted in the carotid artery toward the brain in male Wistar rats. After 1 wk recovery, animals were food-deprived for 5 h, then 10 min infusions of triglyceride emulsion, Intralipid +/- heparin (IL, ILH, respectively) or saline/heparin (SH) were carried out and food intake was assessed over the next 5 h. Experimental groups included: 1) Rats previously injected in ventromedian nucleus (VMN) with shRNA against CD36 or scrambled RNA; 2) Etomoxir (CPT1 inhibitor) or saline co-infused with ILH/SH; and 3) Triacsin C (acylCoA synthase inhibitor) or saline co-infused with ILH/SH. ILH significantly lowered food intake during refeeding compared to SH (p

http://www.ncbi.nlm.nih.gov/pubmed/23894333; International audience; AIMS: Hypoglycemia is a severe side effect of intensive insulin therapy. Recurrent hypoglycemia (RH) impairs the counter-regulatory response (CRR) which restores euglycemia. During hypoglycemia, ventromedial hypothalamus (VMH) production of nitric oxide (NO) and activation of its receptor soluble guanylyl cyclase (sGC) are critical for the CRR. Hypoglycemia also increases brain reactive oxygen species (ROS) production. NO production in the presence of ROS causes protein S-nitrosylation. S-nitrosylation of sGC impairs its function and induces desensitization to NO. We hypothesized that during hypoglycemia, the interaction between NO and ROS increases VMH sGC S-nitrosylation levels and impairs the CRR to subsequent episodes of hypoglycemia. VMH ROS production and S-nitrosylation were quantified following three consecutive daily episodes of insulin-hypoglycemia (RH model). The CRR was evaluated in rats in response to acute insulin-induced hypoglycemia or via hypoglycemic-hyperinsulinemic clamps. Pretreatment with the anti-oxidant N-acetyl-cysteine (NAC) was used to prevent increased VMH S-nitrosylation.RESULTS: Acute insulin-hypoglycemia increased VMH ROS levels by 49±6.3%. RH increased VMH sGC S-nitrosylation. Increasing VMH S-nitrosylation with intracerebroventricular injection of the nitrosylating agent S-nitroso-L-cysteine (CSNO) was associated with decreased glucagon secretion during hypoglycemic clamp. Finally, in RH rats pre-treated with NAC (0.5% in drinking water for 9 days) hypoglycemia-induced VMH ROS production was prevented and glucagon and epinephrine production was not blunted in response to subsequent insulin-hypoglycemia.CONCLUSION: These data suggest that NAC may be clinically useful in preventing impaired CRR in patients undergoing intensive-insulin therapy.