Syntaxin 4 Eenrichment in β-cells Prevents Conversion to Autoimmune Diabetes in Non-Obese Diabetic (NOD) Mice

Syntaxin 4 (STX4), a plasma membrane-localized SNARE protein, regulates human islet β-cell insulin secretion and preservation of β-cell mass. We found that human type 1 diabetic (T1D) and non-obese diabetic (NOD) mouse islets show reduced β-cell STX4 expression, consistent with decreased STX4 expression as a potential driver of T1D phenotypes. To test this hypothesis, we generated inducible β-cell-specific STX4-expressing NOD mice (NOD-iβSTX4).<b> </b>Of NOD-iβSTX4 mice, 73% had sustained normoglycemia versus <20% of control NOD (NOD-Ctrl) mice, by 25 weeks of age. At 12 weeks of age, prior to diabetes conversion, NOD-iβSTX4 mice demonstrated superior whole-body glucose tolerance and β-cell glucose responsiveness than NOD-Ctrl mice. Higher β-cell mass and reduced β-cell apoptosis were also detected in NOD-iβSTX4 pancreata compared with those of NOD-Ctrl mice. Single-cell RNA‐sequencing revealed that islets from NOD-iβSTX4 had markedly reduced IFNƔ signaling and TNFα signaling via NF-ĸB in islet β-cells, including reduced expression of the chemokine CCL5; CD4<sup>+</sup> Treg cells were also enriched in NOD-iβSTX4 islets. These results provide a deeper mechanistic understanding of STX4 function in β-cell protection and warrant further investigation of STX4 enrichment as a strategy to reverse or prevent T1D in humans or protect β-cell grafts.

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Syntaxin 4 Eenrichment in β-cells Prevents Conversion to Autoimmune Diabetes in Non-Obese Diabetic (NOD) Mice

10.2337/figshare.16613851.v1 ◽

2021 ◽

Author(s):

Eunjin Oh ◽

Erika M. McCown ◽

Miwon Ahn ◽

Pablo A. Garcia ◽

Sergio Branciamore ◽

...

Keyword(s):

Autoimmune Diabetes ◽

Cell Mass ◽

Nod Mice ◽

Treg Cells ◽

Whole Body ◽

Cell Protection ◽

Β Cells ◽

Β Cell ◽

Syntaxin 4 ◽

Glucose Responsiveness

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InsB9-23 Gene Transfer To Hepatocytes-Based Combined Therapy Abrogates Recurrence of Type-1 Diabetes After Islet Transplantation

10.2337/figshare.13135592 ◽

2020 ◽

Author(s):

Ada Admin ◽

Fabio Russo ◽

Antonio Citro ◽

Giorgia Squeri ◽

Francesca Sanvito ◽

...

Keyword(s):

T Regulatory Cells ◽

Lentiviral Vector ◽

Autoimmune Diabetes ◽

Therapeutic Option ◽

Combined Therapy ◽

Cell Mass ◽

Nod Mice ◽

Β Cell ◽

Suboptimal Dose ◽

Islets Transplantation

The induction of antigen (Ag)-specific tolerance represents a therapeutic option for autoimmune diabetes. We demonstrated that administration of lentiviral vector enabling expression of insulinB9-23 (LV.InsB) in hepatocytes, arrests β cell destruction in pre-diabetic NOD mice, by generating InsB9-23-specific FoxP3+T regulatory cells (Tregs). LV.InsB in combination with a suboptimal dose of anti-CD3 mAb (combined therapy, 1X5µg CT5) reverts diabetes and prevents recurrence of autoimmunity following islets transplantation in ~50% of NOD mice. We investigated whether CT optimization could lead to abrogation of recurrence of autoimmunity. Therefore, allo-islets were transplanted after optimized CT tolerogenic conditioning (1X25µg CT25). Diabetic NOD mice conditioned with CT25 when glycaemia was <500mg/dL, remained normoglycaemic for 100 days after allo-islets transplantation, displayed reduced insulitis, but independently from the graft. Accordingly, cured mice showed T cell unresponsiveness to InsB9-23 stimulation and increased Tregs frequency in islets infiltration and pancreatic LN. Additional studies revealed a complex mechanism of Ag-specific immune regulation driven by CT25, in which both Tregs and PDL1 co-stimulation cooperate to control diabetogenic cells, while transplanted islets play a crucial role, although transient, recruiting diabetogenic cells. Therefore, CT25 before allo-islets transplantation represents an Ag-specific immunotherapy to resolve autoimmune diabetes in the presence of residual endogenous β cell mass.

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β-Cell adaptation in 60% pancreatectomy rats that preserves normoinsulinemia and normoglycemia

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.2000.279.1.e68 ◽

2000 ◽

Vol 279 (1) ◽

pp. E68-E73 ◽

Cited By ~ 30

Author(s):

Ye Qi Liu ◽

Peter W. Nevin ◽

Jack L. Leahy

Keyword(s):

Insulin Secretion ◽

Regulatory Element ◽

Cell Mass ◽

Maximal Velocity ◽

Β Cells ◽

Β Cell ◽

Biochemical Basis ◽

Adaptive Mechanisms ◽

Sense And Respond ◽

Glucose Responsiveness

Islet β-cells are the regulatory element of the glucose homeostasis system. When functioning normally, they precisely counterbalance changes in insulin sensitivity or β-cell mass to preserve normoglycemia. This understanding seems counter to the dogma that β-cells are regulated by glycemia. We studied 60% pancreatectomy rats (Px) 4 wk postsurgery to elucidate the β-cell adaptive mechanisms. Nonfasting glycemia and insulinemia were identical in Px and sham-operated controls. There was partial regeneration of the excised β-cells in the Px rats, but it was limited in scope, with the pancreas β-cell mass reaching 55% of the shams (40% increase from the time of surgery). More consequential was a heightened glucose responsiveness of Px islets so that glucose utilization and insulin secretion per milligram of islet protein were both 80% augmented at normal levels of glycemia. Investigation of the biochemical basis showed a doubled glucokinase maximal velocity in Px islets, with no change in the glucokinase protein concentration after adjustment for the different β-cell mass in Px and sham islets. Hexokinase activity measured in islet extracts was also minimally increased, but the glucose 6-phosphate concentration and basal glucose usage of Px islets were not different from those in islets from sham-operated rats. The dominant β-cell adaptive response in the 60% Px rats was an increased catalytic activity of glucokinase. The remaining β-cells thus sense, and respond to, perceived hyperglycemia despite glycemia actually being normal. β-Cell mass and insulin secretion are both augmented so that whole pancreas insulin output, and consequently glycemia, are maintained at normal levels.

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Transient Depletion of Foxp3+ Regulatory T Cells Selectively Promotes Aggressive β Cell Autoimmunity in Genetically Susceptible DEREG Mice

Frontiers in Immunology ◽

10.3389/fimmu.2021.720133 ◽

2021 ◽

Vol 12 ◽

Author(s):

Deepika Watts ◽

Marthe Janßen ◽

Mangesh Jaykar ◽

Francesco Palmucci ◽

Marc Weigelt ◽

...

Keyword(s):

T Cells ◽

Treg Cell ◽

Autoimmune Diabetes ◽

Cell Activity ◽

Spontaneous Diabetes ◽

Nod Mice ◽

Treg Cells ◽

Β Cell ◽

Cell Ablation ◽

Tcr Repertoire

Type 1 diabetes (T1D) represents a hallmark of the fatal multiorgan autoimmune syndrome affecting humans with abrogated Foxp3+ regulatory T (Treg) cell function due to Foxp3 gene mutations, but whether the loss of Foxp3+ Treg cell activity is indeed sufficient to promote β cell autoimmunity requires further scrutiny. As opposed to human Treg cell deficiency, β cell autoimmunity has not been observed in non-autoimmune-prone mice with constitutive Foxp3 deficiency or after diphtheria toxin receptor (DTR)-mediated ablation of Foxp3+ Treg cells. In the spontaneous nonobese diabetic (NOD) mouse model of T1D, constitutive Foxp3 deficiency did not result in invasive insulitis and hyperglycemia, and previous studies on Foxp3+ Treg cell ablation focused on Foxp3DTR NOD mice, in which expression of a transgenic BDC2.5 T cell receptor (TCR) restricted the CD4+ TCR repertoire to a single diabetogenic specificity. Here we revisited the effect of acute Foxp3+ Treg cell ablation on β cell autoimmunity in NOD mice in the context of a polyclonal TCR repertoire. For this, we took advantage of the well-established DTR/GFP transgene of DEREG mice, which allows for specific ablation of Foxp3+ Treg cells without promoting catastrophic autoimmune diseases. We show that the transient loss of Foxp3+ Treg cells in prediabetic NOD.DEREG mice is sufficient to precipitate severe insulitis and persistent hyperglycemia within 5 days after DT administration. Importantly, DT-treated NOD.DEREG mice preserved many clinical features of spontaneous diabetes progression in the NOD model, including a prominent role of diabetogenic CD8+ T cells in terminal β cell destruction. Despite the severity of destructive β cell autoimmunity, anti-CD3 mAb therapy of DT-treated mice interfered with the progression to overt diabetes, indicating that the novel NOD.DEREG model can be exploited for preclinical studies on T1D under experimental conditions of synchronized, advanced β cell autoimmunity. Overall, our studies highlight the continuous requirement of Foxp3+ Treg cell activity for the control of genetically pre-installed autoimmune diabetes.

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Selective Deletion of Pten in Pancreatic β Cells Leads to Increased Islet Mass and Resistance to STZ-Induced Diabetes

Molecular and Cellular Biology ◽

10.1128/mcb.26.7.2772-2781.2006 ◽

2006 ◽

Vol 26 (7) ◽

pp. 2772-2781 ◽

Cited By ~ 92

Author(s):

Bangyan L. Stiles ◽

Christine Kuralwalla-Martinez ◽

Wei Guo ◽

Caroline Gregorian ◽

Ying Wang ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Mass ◽

Cell Protection ◽

Β Cells ◽

Β Cell ◽

Islet Mass ◽

Pten Loss ◽

Lipid Phosphatase ◽

Streptozotocin Induced Diabetes ◽

Phosphatase And Tensin Homologue

ABSTRACT Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is a lipid phosphatase. PTEN inhibits the action of phosphatidylinositol-3-kinase and reduces the levels of phosphatidylinositol triphosphate, a crucial second messenger for cell proliferation and survival, as well as insulin signaling. In this study, we deleted Pten specifically in the insulin producing β cells during murine pancreatic development. Pten deletion leads to increased cell proliferation and decreased cell death, without significant alteration of β-cell differentiation. Consequently, the mutant pancreas generates more and larger islets, with a significant increase in total β-cell mass. PTEN loss also protects animals from developing streptozotocin-induced diabetes. Our data demonstrate that PTEN loss in β cells is not tumorigenic but beneficial. This suggests that modulating the PTEN-controlled signaling pathway is a potential approach for β-cell protection and regeneration therapies.

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Syntaxin 4 enrichment in β-cells prevents conversion to autoimmune diabetes in non-obese diabetic (NOD) mice

Diabetes ◽

10.2337/db21-0170 ◽

2021 ◽

pp. db210170

Author(s):

Eunjin Oh ◽

Erika M. McCown ◽

Miwon Ahn ◽

Pablo A. Garcia ◽

Sergio Branciamore ◽

...

Keyword(s):

Autoimmune Diabetes ◽

Nod Mice ◽

Β Cells ◽

Syntaxin 4

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Detecting Insulitis in Type 1 Diabetes with Ultrasound Phase-change Contrast Agents

10.1101/2020.10.28.359687 ◽

2020 ◽

Author(s):

David G. Ramirez ◽

Awaneesh K. Upadhyay ◽

Vinh T. Pham ◽

Mark Ciccaglione ◽

Mark A Borden ◽

...

Keyword(s):

Type 1 Diabetes ◽

Contrast Agents ◽

Cell Mass ◽

Nod Mice ◽

Therapeutic Interventions ◽

Β Cells ◽

Β Cell ◽

Diabetes Onset ◽

Ultrasound Contrast

AbstractType 1 diabetes (T1D) results from immune infiltration and destruction of insulin-producing β-cells within the pancreatic islets of Langerhans (insulitis), resulting in loss of glucose homeostasis. Early diagnosis during pre-symptomatic T1D would allow for therapeutic intervention prior to substantial loss of β-cell mass at T1D onset. There are limited methods to track the progression of insulitis and β-cell mass decline in pre-symptomatic T1D. During insulitis, the islet microvasculature increases permeability, such that sub-micron sized particles can extravasate and accumulate within the islet microenvironment. Ultrasound is a widely deployable and cost-effective clinical imaging modality. However, conventional microbubble contrast agents are restricted to the vasculature. Sub-micron sized nanodroplet (ND) phasechange agents can be vaporized into micron-sized bubbles; serving as a circulating microbubble precursor. We tested if NDs extravasate into the immune-infiltrated islet microenvironment. We performed ultrasound contrast-imaging following ND infusion in NOD mice and NOD;Rag1ko controls, and tracked diabetes development. We measured the biodistribution of fluorescently labeled NDs, with histological analysis of insulitis. Ultrasound contrast signal was elevated in the pancreas of 10w NOD mice following ND infusion and vaporization, but was absent in both the non-infiltrated kidney of NOD mice and pancreas of Rag1ko controls. High contrast elevation also correlated with rapid diabetes onset. In pancreata of NOD mice, infiltrated islets and nearby exocrine tissue were selectively labeled with fluorescent NDs. Thus, contrast ultrasound imaging with ND phase-change agents can detect insulitis prior to diabetes onset. This will be important for monitoring disease progression to guide and assess preventative therapeutic interventions for T1D.SignificanceThere is a need for imaging methods to detect type1 diabetes (T1D) progression prior to clinical diagnosis. T1D is a chronic disease that results from autoreactive T cells infiltrating the islet of Langerhans and destroying insulin-producing β-cells. Overt disease takes years to present and is only diagnosed after significant β-cells loss. As such, the possibility of therapeutic intervention to preserve β-cell mass is hampered by an inability to follow pre-symptomatic T1D progression. There are immunotherapies that can delay T1D development. However identifying ‘at risk’ individuals, and tracking whether therapeutic interventions are impacting disease progression, prior to T1D onset, is lacking. A method to detect insulitis and β-cell mass decline would present an opportunity to guide therapeutic treatments to prevent T1D.

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Effective Destruction of Fas-deficient Insulin-producing β Cells in Type 1 Diabetes

Journal of Experimental Medicine ◽

10.1084/jem.20030698 ◽

2003 ◽

Vol 198 (7) ◽

pp. 1103-1106 ◽

Cited By ~ 52

Author(s):

Irina Apostolou ◽

Zhenyue Hao ◽

Klaus Rajewsky ◽

Harald von Boehmer

Keyword(s):

T Cells ◽

Cell Death ◽

Type 1 Diabetes ◽

Autoimmune Diabetes ◽

Nod Mice ◽

Β Cells ◽

Β Cell ◽

Influenza Hemagglutinin ◽

Insulin Promoter

In type 1 diabetes, autoimmune T cells cause destruction of pancreatic β cells by largely unknown mechanism. Previous analyses have shown that β cell destruction is delayed but can occur in perforin-deficient nonobese diabetic (NOD) mice and that Fas-deficient NOD mice do not develop diabetes. However, because of possible pleiotropic functions of Fas, it was not clear whether the Fas receptor was an essential mediator of β cell death in type 1 diabetes. To directly test this hypothesis, we have generated a β cell–specific knockout of the Fas gene in a transgenic model of type 1 autoimmune diabetes in which CD4+ T cells with a transgenic TCR specific for influenza hemagglutinin (HA) are causing diabetes in mice that express HA under control of the rat insulin promoter. Here we show that the Fas-deficient mice develop autoimmune diabetes with slightly accelerated kinetics indicating that Fas-dependent apoptosis of β cells is a dispensable mode of cell death in this disease.

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Beta-cell specific insulin resistance promotes glucose-stimulated insulin hypersecretion

10.1101/2020.10.15.338160 ◽

2020 ◽

Author(s):

Søs Skovsø ◽

Evgeniy Panzhinskiy ◽

Jelena Kolic ◽

Derek A. Dionne ◽

Xiao-Qing Dai ◽

...

Keyword(s):

Insulin Resistance ◽

Glucose Tolerance ◽

Cell Mass ◽

Whole Body ◽

Calcium Oscillation ◽

Male Mice ◽

Β Cells ◽

Β Cell ◽

Specific Insulin

AbstractInsulin receptor (Insr) protein can be found at higher levels in pancreatic β-cells than in most other cell types, but the consequences of β-cell insulin resistance remain enigmatic. Ins1cre allele was used to delete Insr specifically in β-cells of both female and male mice which were compared to Ins1cre-containing littermate controls at multiple ages and on multiple diets. RNA-seq of recombined β-cells revealed significant differences in multiple pathways previously implicated in insulin secretion and cellular fate, including rewired Ras and NFκB signaling. Male, but not female, βInsrKO mice had reduced oxygen consumption rate, while action potential and calcium oscillation frequencies were increased in Insr knockout β-cells from female, but not male mice. Female βInsrKO and βInsrHET mice exhibited elevated insulin release in perifusion experiments, during hyperglycemic clamps, and following i.p. glucose challenge. Deletion of Insr did not reduce β-cell mass up to 9 months of age, nor did it impair hyperglycemia-induced proliferation. Based on our data, we adapted a mathematical model to include β-cell insulin resistance, which predicted that β-cell Insr knockout would improve glucose tolerance depending on the degree of whole-body insulin resistance. Indeed, glucose tolerance was significantly improved in female βInsrKO and βInsrHET mice when compared to controls at 9, 21 and 39 weeks. We did not observe improved glucose tolerance in adult male mice or in high fat diet-fed mice, corroborating the prediction that global insulin resistance obscures the effects of β-cell specific insulin resistance. We further validated our in vivo findings using the Ins1-CreERT transgenic line and found improved glucose tolerance 4 weeks after tamoxifen-mediated Insr deletion. Collectively, our data show that loss of β-cell Insr alone is sufficient to drive glucose-induced hyperinsulinemia, thereby improving glucose homeostasis in otherwise insulin sensitive dietary and age contexts.

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