Cells In Pancreatic Lymph Nodes Research Articles

Pam3CSK4 enhanced beta cell loss and diabetogenesis: The roles of IFN-gamma and IL-17

Toll like receptors are primary sensors of both innate and adaptive immune systems. They activate APCs and influence T-cell function in inflammatory autoimmune response. Studies have shown that TLR manipulation may lead to either tolerance or trigger autoimmunity. Using diabetogenic and subdiabetogenic multiple low doses of streptozotocin, we demonstrate here that Pam3 CYS-CK4 a TLR-2 agonist, enhances and promotes diabetes in C57BL/6 male mice following increased apoptosis of β islet cells. FACS analysis of isolated pancreatic lymph node cells revealed significant increased number of macrophages, dendritic cells, CD4(+) TNF-α(+), CD4(+) IFN-γ(+) and most significantly, CD4(+) IL-17(+) and reduced number of CD25(+)Fox p3(+) T cells after Pam3CSK4 treatment. Genetic deletion of IFN-γ prevents whereas deletion of IL-17 reduced severity of Pam3CSK4-induced enhancement of diabetes. TLR-2 agonist-enhanced diabetogenesis is also influenced by enhanced influx of antigen presenting cells and suppression of regulatory T cell activity.

Resistance of diabetes in aged NOD mice is mediated by CD4+CD25+Foxp3+ regulatory T cells (P4144)

Abstract We investigated the role of CD4+CD25+FoxP3+ regulatory T cells on diabetes resistance in aged NOD mice that do not develop diabetes in pathogen-free housing. At 60 weeks age, <80% NOD mice developed diabetes. Adoptive transfer of lymphocytes from diabetic mice to aged nodiabetic NOD mice did not significantly increase the onset of diabetes. When 1x107 lymphocytes from age NOD mice were co-transferred with 5x106 lymphocytes from diabetes NOD mice into NOD.scid mice, all mice developed diabetes at 6 weeks. When 2x107 lymphocytes from aged NOD mice were co-transferred, none developed diabetes at 6 weeks and only 50% NOD.scid mice developed diabetes at 12 weeks. When 1x106 CD4+CD25- T cells was cotransferred with 5x106 lymphocytes from diabetic NOD mice, diabetes was developed in all NOD.scid mice. When 1x106 CD4+CD25+ T cells were used, only 17% mice developed diabetes at 12 weeks. The percentages of CD4+CD25+Foxp3+ T cells in the pancreatic lymph nodes, spleen and peripheral blood were 20.1±5.3%, 14.6±4.1% and 9.8±3.5% in aged NOD mice; 5.8±1.5%, 8.6±0.7% and 6.4±1.7% in control diabetic NOD mice. When a low dose of cyclosphamiade was given, diabetes was developed in 75% young NOD mice and in 18% aged NOD mice. However, when the high dose was given, 58% aged NOD mice had diabetes and CD4+CD25+Foxp3+ T cells in pancreatic lymph nodes were 7.0±1.7%. Our data demonstrated that immunoregulation through CD4+CD25+Foxp3+ T cells mediated the resistance of diabetes in aged NOD mice.

Adoptive cell transfer from multiple low dose streptozotocin diabetic donors can induce diabetes in healthy young C57/BL6 mice

Event Abstract Back to Event Adoptive cell transfer from multiple low dose streptozotocin diabetic donors can induce diabetes in healthy young C57/BL6 mice Nemanja Zdravkovic1*, Sladjana Pavlovic1 and Miodrag L. Lukic1 1 Faculty of Medical Sciences, University of Kragujevac, Serbia, Center for Molecular Medicine and Stem Cell Research, Serbia Multiple low dose of streptozoticin (MLD-STZ) diabetes in C57/BL6 mice is the result of T cell–dependent process directed against the pancreatic beta cells. However, diabetes induction after adoptive transfer of splenocytes and pancreatic lymph node cells (pLNc) has not been established. Therefore, we investigated whether splenocytes (1x108 viable cells) and pLNc (5x106 viable cells) from diabetic donors can induce diabetes in young (6-wk-old) C57/BL6 mice. Our work shows that i.p. transfer of splenocytes and pLNc from MLD-STZ mice altered i.p. glucose tolerance, stimulated the onset of glycosuria and intra-insulitis in young recipients, which was accompanied by the increased levels of TNF-α, IFN-γ, IL-17, and decreased level of IL-10 in the sera. The precise role of different types of mononuclear cells in diabetes induction has been determined by FACS analysis. The number of CD4+TNF-α+, CD4+IFN-γ+, CD4+IL-17+, CD3+CD8+, CD19+, F4/80+IL-12+ cells was significantly increased (p<0.001), while the number of CD3-CD8-NK1.1+ cells was decreased (p<0.05) in pancreatic lymph nodes and pancreas of recipients in comparison to healthy mice. In contrast with pancreatic lymph nodes, the number of CD19+ B cells in pancreas was similar in the recipients and healthy mice, suggesting that effector Th1, Th17, CD3+CD8+ T cells and M1 macrophages generated infiltrates in the islets of Langerhans in young C57/BL6 recipients. Our study provided the first evidence that splenocytes and pLNc of MLD-STZ diabetic male C57/BL6 mice can transfer diabetes to young syngeneic mice and identifies a new strategy for the examination of autoimmune diabetes. Acknowledgements Supported by the Serbian Ministry of Science (Grant # ON 175069) and from the Faculty of Medical Sciences, University of Kragujevac (grant JP 08/10), Serbia. Keywords: Diabetes Mellitus, Type 1, transfer diabetes, autoimmunity models, splenocytes, pancreatic lymph node cells Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Translational immunology and immune intervention Citation: Zdravkovic N, Pavlovic S and Lukic ML (2013). Adoptive cell transfer from multiple low dose streptozotocin diabetic donors can induce diabetes in healthy young C57/BL6 mice. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00688 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 12 Jun 2013; Published Online: 22 Aug 2013. * Correspondence: Prof. Nemanja Zdravkovic, Faculty of Medical Sciences, University of Kragujevac, Serbia, Center for Molecular Medicine and Stem Cell Research, Kragujevac, 34000, Serbia, zdravkovic_nemanja@yahoo.com Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Nemanja Zdravkovic Sladjana Pavlovic Miodrag L Lukic Google Nemanja Zdravkovic Sladjana Pavlovic Miodrag L Lukic Google Scholar Nemanja Zdravkovic Sladjana Pavlovic Miodrag L Lukic PubMed Nemanja Zdravkovic Sladjana Pavlovic Miodrag L Lukic Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Treg cells in pancreatic lymph nodes: the possible role in diabetogenesis and β cell regeneration in a T1D model

Previously, we established a model in which physiologically adequate function of the autologous β cells was recovered in non-obese diabetic (NOD) mice after the onset of hyperglycemia by rendering them hemopoietic chimera. These mice were termed antea-diabetic. In the current study, we addressed the role of T regulatory (Treg) cells in the mechanisms mediating the restoration of euglycemia in the antea-diabetic NOD model. The data generated in this study demonstrated that the numbers of Treg cells were decreased in unmanipulated NOD mice, with the most profound deficiency detected in the pancreatic lymph nodes (PLNs). The impaired retention of the Treg cells in the PLNs correlated with the locally compromised profile of the chemokines involved in their trafficking, with the most prominent decrease observed in SDF-1. The amelioration of autoimmunity and restoration of euglycemia observed in the antea-diabetic mice was associated with restoration of the Treg cell population in the PLNs. These data indicate that the function of the SDF-1/CXCR4 axis and the retention of Treg cells in the PLNs have a potential role in diabetogenesis and in the amelioration of autoimmunity and β cell regeneration in the antea-diabetic model. We have demonstrated in the antea-diabetic mouse model that lifelong recovery of the β cells has a strong correlation with normalization of the Treg cell population in the PLNs. This finding offers new opportunities for testing the immunomodulatory regimens that promote accumulation of Treg cells in the PLNs as a therapeutic approach for type 1 diabetes (T1D).

Dynamic changes of the Th17/Tc17 and regulatory T cell populations interfere in the experimental autoimmune diabetes pathogenesis

A balance between proinflammatory (Th17 and Tc17) and anti-inflammatory (regulatory T cells) subsets of T cells is essential to maintain immunological tolerance and prevent the onset of several autoimmune diseases, including type 1 diabetes. However, the kinetics of these subsets and disease severity during the streptozotocin (STZ)-induced diabetes course has not been determined. Thus, susceptible C57BL/6 mice were administrated with multiple low doses of STZ and we evaluated the frequency/absolute number of these T cell subsets in the pancreatic lymph nodes (PLNs) and spleen and Th1, Th17, Treg cytokine production in the pancreatic tissue. At different time points of the disease progression (6, 11, 18 and 25 days after the last STZ administration), the histopathological alterations were also evaluated by H&E and immunohistochemistry staining. During the initial phase of diabetes development (day 6), we noted increased numbers of CD4+ and CD8+ T cells in spleen and PLNs. At the same time, the frequencies of Th17 and Tc17 cells in PLNs were also enhanced. In addition, the early augment of interferon gamma (IFN-γ), tumoral necrosis factor (TNF-α), IL-6 and IL-17 levels in pancreatic tissue correlated with pancreatic islet inflammation and mild β-cell damage. Notably, the absolute number of Treg cells increased in PLNs during over time when compared to control group. Interestingly, increased IL-10 levels were associated with control of the inflammatory process during the late phase of the type 1 diabetes (day 25). In agreement, mice lacking the expression of IL-17 receptor (Il17r) showed impairment in STZ-induced diabetes progression, reduced peri-insulitis and beta cells preservation when compared with wild-type mice. Our findings suggest that dynamic changes of pathogenic Th17/Tc17 and regulatory T cell subsets numbers is associated with early strong inflammation in the pancreatic islets followed by late regulatory profile during the experimental STZ-induced diabetes course.

Endogenous Expansion of Regulatory T Cells Leads to Long-Term Islet Graft Survival in Diabetic NOD Mice

Donor pancreatic lymph node cells (PLNC) protect islet transplants in Non-obese diabetic (NOD) mice. We hypothesized that induced FoxP3(+) regulatory T cells (Tregs) were required for long-term islet engraftment. NOD or NOD.NON mice were treated with ALS (antilymphocyte serum) and transplanted with NOR islets +/-PLNC (5 × 10(7) ). In vivo proliferation and expansion of FoxP3(+) Tregs was monitored in spleen and PLN from ALS- and ALS/PLNC-treated recipient mice. Anti-CD25 depletion was used to determine the necessity of Tregs for tolerance. FoxP3(+) numbers significantly increased in ALS/PLNC-treated recipients compared to ALS-treated mice. In ALS/PLNC-treated mice, recipient-derived Tregs localized to the transplanted islets, and this was associated with intact, insulin-producing β cells. Proliferation and expansion of FoxP3(+) Tregs was markedly increased in PLNC-treated mice with accepted islet grafts, but not in diabetic mice not receiving PLNC. Deletion of Tregs with anti-CD25 antibodies prevented islet graft tolerance and resulted in rejection. Adoptive transfer of Tregs to secondary NOD.scid recipients inhibited autoimmunity by cotransferred NOD effector T cells. Treg expansion induced by ALS/PLNC-treatment promoted long term islet graft survival. Strategies leading to Treg proliferation and localization to the transplant site represent a therapeutic approach to controlling recurrent autoimmunity.

Oral administration of Capsaicin, a ligand of VR1, can modulate dendritic cell population and induces CD4+FoxP3- Tr-1 cells in the GALT (178.14)

Abstract VR1 is a non-selective cation channel, highly expressed by sensory neurons and cells of the immune system e.g. dendritic cells (DCs) and macrophages. Capsaicin (CP), the pungent component of chili peppers is a chemical ligand of VR1. We have recently reported that oral administration of CP inhibits proliferation and activation of auto-reactive T cells and protects mice from development of Type-1 diabetes (T1D) in a dose dependent manner. Here we demonstrate that orally administered CP can lead to an increase in IL-27 producing DCs, and IL10 producing CD4+/FoxP3- T cells (Tr1) but decrease in the frequency of CD4+/IL17+ T cells (TH17) in the pancreatic lymph node (PLN). We also demonstrate that splenocytes and PLN cells when adoptively transferred from capsaicin fed mouse to a naïve NOD mouse leads to protection of the recipient mouse from T1D. Our results thus demonstrate that engagement of VR1 on immune cells can have profound effect on gut mediated immune tolerance and autoimmunity.

Oral ingestion of Capsaicin, the pungent component of chili pepper, enhances a discreet population of macrophages and confers protection from autoimmune diabetes.

Vanilloid receptor 1 (VR1) is expressed on immune cells as well as on sensory neurons. Here we report that VR1 can regulate immunological events in the gut in response to its ligand Capsaicin (CP), a nutritional factor, the pungent component of chili peppers. Oral administration of CP attenuates the proliferation and activation of autoreactive T cells in pancreatic lymph nodes (PLNs) but not other lymph nodes, and protects mice from development of type 1 diabetes (T1D). This is a general phenomenon and not restricted to one particular strain of mice. Engagement of VR1 enhances a discreet population of CD11b(+)/F4/80(+) macrophages in PLN, which express anti-inflammatory factors interleukin (IL)-10 and PD-L1. This population is essential for CP-mediated attenuation of T-cell proliferation in an IL-10-dependent manner. Lack of VR1 expression fails to inhibit proliferation of autoreactive T cells, which is partially reversed in (VR1(+/+) → VR1(-/-)) bone marrow chimeric mice, implying the role of VR1 in crosstalk between neuronal and immunological responses in vivo. These findings imply that endogenous ligands of VR1 can have profound effect on gut-mediated immune tolerance and autoimmunity by influencing the nutrient-immune interactions.

Opposing roles for Fas ligand and IL-10 in regulating insulitis initiation (101.32)

Abstract Type 1 diabetes (TID), leading autoimmune disease of childhood, is caused by destruction of beta-cells by autoreactive T cells. Despite daily insulin injections, patients often develop cardiovascular complications and there is a need for identifying therapeutic targets to prevent the disease without impinging on host defense. Fas ligand (FasL) can be such a potential target. Fas/FasL interactions primarily regulate T cell homeostasis but not activation. Nevertheless, mutation of FasL (gld) prevents autoimmune diabetes in NOD mice, the widely used model for T1D. Although homozygous gld mutations cause age-dependent lymphoproliferation, limiting the gld mutation to one allele (NOD-gld/+) or treating NOD-wt mice with FasL-neutralizing mAb inhibits insulitis and prevents disease development without causing lymphoproliferation or immunosuppression. Here we show that the gld mutation did not inhibit proliferation and expansion of diabetogenic T cells in pancreatic lymph nodes but inhibited their accumulation in the pancreata. A significant proportion of B cells found in pancreata of NOD-gld/+ mice surface-expressed CD5 and produced IL-10. Neutralization of IL-10 allowed CD4+ T cell accumulation in the pancreas and development of insulitis in NOD-gld/+ mice. The results provide novel mechanistic insights into the disease pathogenesis with potential implications for developing novel T1D therapy.

Dry olive leaf extract (DOLE) down-regulates the progression of experimental immune-mediated diabetes by modulation of cytokine profile in the draining lymph nodes

We have recently demonstrated the beneficial effects of dry olive leaf extract (DOLE) in two preclinical models of type 1 diabetes. Here we analyze the potential mechanisms underlying diabetes amelioration at the level of lymph node drainage. Treatment of C57BL/6 mice with DOLE during induction of diabetes with multiple low doses of streptozotocin (MLD-SZ) modulated cytokine expression and production in pancreatic lymph node cells, thereby changing the balance between potentially pathogenic and down-regulating cytokines. These results support the immunoregulatory potential of DOLE which takes place at the level of lymph node drainage and preserves the target tissue from autoimmune attack.

A key role for Fas ligand on B cells in unleashing T cell diabetogenicity by suppression of IL-10-mediated islet tolerance (143.34)

Abstract A key role for the Fas pathway in promoting organ-specific-autoimmunity is indicated by resistance of mice bearing the lpr mutation of Fas or gld mutation of Fas ligand (FasL) to several organ-specific autoimmune diseases, including autoimmune diabetes and multiple sclerosis. However, the underlying mechanism remains elusive. Here we show that FasL expressed on B cells plays a key role in facilitating activation of diabetogenic T cells in pancreatic lymph nodes (PLNs) and pancreatic islet accumulation resulting in initiation of insulitis in the autoimmune diabetes-prone NOD mice. Expression of FasL on diabetogenic T cells themselves is not sufficient to mediate its diabetogenic role. By contrast, expression of FasL on B cells leads to deletion of localized IL-10-producing B cells, the presence of which in PLNs and pancreatic islets is associated with suppression of diabetogenic T cell priming and islet accumulation, respectively. Consequently, FasL inactivation by gld mutation or antibody blockade preserves the local IL-10-producing B cells and protects islets from diabetogenic T cells by a mechanism that is reversed by IL-10R blockade in vivo. The results provide novel insights into how FasL orchestrates the pathogenic relationship among B cell subsets in its implications for diabetogenic T cells.

Activation of Aryl Hydrocarbon Receptor by Tcdd Prevents Diabetes in Nod Mice and Increases Foxp3 + T Cells in Pancreatic Lymph Nodes

The ligand-activated transcription factor, aryl hydrocarbon receptor (AHR), is a novel inducer of adaptive Tregs. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), the most potent AHR ligand, induces adaptive CD4+CD25+ Tregs during an acute graft-versus-host (GvH) response and prevents the generation of allospecific cytotoxic T lymphocytes. TCDD also suppresses the induction of experimental autoimmune encephalitis in association with an expanded population of Foxp3+ Tregs. In this study, we show that chronic treatment of NOD mice with TCDD potently suppresses the development of autoimmune Type 1 diabetes in parallel with greatly reduced pancreatic islet insulitis and an expanded population of CD4+CD25+Foxp3+ cells in the pancreatic lymph nodes. When treatment with TCDD was terminated after 15 weeks (23 weeks of age), mice developed diabetes over the next 8 weeks in association with lower numbers of Tregs and decreased activation of AHR. Analysis of the expression levels of several genes associated with inflammation, T-cell activation and/or Treg function in pancreatic lymph node cells failed to reveal any differences associated with TCDD treatment. Taken together, the data suggest that AHR activation by TCDD-like ligands may represent a novel avenue for treatment of immune-mediated diseases.

The VR1 ligand capsaicin confers protection from autoimmune diabetes by suppressing autoreactive T cells in an APC dependent manner (50.38)

Abstract Sensory nerve fibers innervating the pancreas play a critical role in the etiology of autoimmune diabetes. These sensory neurons express a nonselective cation channel vanilloid receptor 1 (VR1) which has been identified as a receptor for acid, temperatures &amp;gt; 43°C and Capsaicin (CP), the principal pungent ingredient in chili peppers. We have reported that dendritic cells (DCs), a key cell type in immune responses, have the receptor for CP, and engagement of this receptor has powerful immune consequences. However very little is known about the function of VR1 expressing pancreas resident antigen presenting cells (APCs) including DCs in Type 1 diabetes. Here we have explored the immunomodulatory activity of CP in the non-obese diabetic (NOD) mouse and found that oral administration of CP delays the onset of diabetes and reduces the incidence of diabetes in NOD mice in a dose dependent manner. Moreover, splenocytes from CP treated mice are able to transfer protection in recipient NOD mice. CP administration attenuates the proliferation of pancreatic antigen-specific autoreactive T cells in pancreatic lymph nodes (PLN) both in vivo and in vitro in two distinct mouse models of diabetes, NOD and RIP-OVA. We have further found that this CP dependent modulation of autoimmunity requires both VR1 expression and a population of myeloid CD11b+ cells present in the PLN. These results reveal that CP is an immunologically potent ligand that when administered orally suppresses autoimmune diabetes by inhibiting the proliferation of autoreactive T cells in an APC dependent manner.

Upregulating CD4+CD25+FOXP3+ Regulatory T Cells in Pancreatic Lymph Nodes in Diabetic NOD Mice by Adjuvant Immunotherapy

Immunotherapy with Complete Freund's adjuvant (CFA) is effective in ameliorating autoimmunity in diabetic nonobese diabetic (NOD) mice. We investigated whether CFA treatment up-regulates CD4+CD25+Foxp3+ regulatory T cells and increases transforming growth factor (TGF)-beta1 production in diabetic NOD mice. New-onset diabetic NOD mice were treated with CFA and exendin-4, a potent analog of glucagon-like peptide-1. Reversal of diabetes was determined by monitoring blood glucose level. Ameliorating autoimmunity through immunoregulation was assessed by adoptive transfer. Regulatory T cells in the peripheral blood, spleen, thymus, and pancreatic nodes were measured. TGF-beta1 in plasma and the insulin content in the pancreas were also measured. Immunostainings for insulin and BrdU were performed. New-onset diabetes could be reversed in 38% of NOD mice treated with CFA alone and in 86% of NOD mice treated with both CFA and exendin-4. Diabetes adoptive transfer by splenocytes from CFA-treated NOD mice was delayed. The percentage of CD4+CD25+Foxp3+ regulatory T cells in the pancreatic lymph nodes of CFA-treated NOD mice was significantly increased at 1, 5, and 15 to 17 weeks after treatment. TGF-beta1 in the plasma of CFA-treated NOD mice was also significantly increased. Combining CFA with exendin-4 treatment significantly increased the insulin content and the numbers of insulin and BrdU double-labeled beta cells in the islets. Our results demonstrated that CFA treatment ameliorates autoimmunity in diabetic NOD mice by up-regulating CD4=CD25+Foxp3+ regulatory T cells and increasing TGF-beta1 production. Exendin-4 enhanced the effect of CFA on reversing diabetes in NOD mice by stimulating beta-cell replication.

Diet and Dysfunctional Metabolism in BB Rats

This review deals with the age- and diet-related metabolic and functional anomalies found in BB rats, currently considered as an animal model of type 1 diabetes mellitus. The metabolism of nutrients, the secretion of insulin, the production of NO, the effects of IL-1β and the activity of the mitochondrial FAD-linked glycerophosphate dehydrogenase in pancreatic islets are first considered. The metabolism and mitotic activity in Peyers patch cells and both mesenteric and pancreatic lymph node cells are then evoked. The occurrence of intestinal dysfunction is mentioned, with emphasis on recent findings made in BB rats before weaning. The participation of zonulins and mucins in the perturbation of gut permeability are also discussed. Last, attention is drawn to the perturbation of the anomeric specificity of glucose-induced insulin release, considered as a manifestation of pancreatic islet beta-cell glucotoxicity. Keywords: BioBreeding rats, type 1 diabetes, diet, pancreatic islets, lymph node cells, gut, mucins, anomeric specificity of glucose-induced insulin release

Galectin 3 is required for MLD-STZ induced diabetes in mice (130.38)

Abstract Galectin-3, member of ancient lectin family characterized by specific binding of β galactosides, has an antiapoptotic function in T cells, macrophages and islet cells. It was therefore of interest to evaluate the susceptibility to multiple low dose induced diabetes (MLD-STZ) in galectin (gal-3) deficient C57BL/6 mice. Gal-3 −/− and gal-3+/+ mice were treated with 5 daily injections of 40 mg/kg STZ and diabetes development evaluated by glycemia and immunohistochemistry of the pancreas. Gal-3+/+ mice developed delayed sustained and progressive hyperglucemia and mononuclear cell infiltrates in the islets. Gal-3 −/− demonstrated only mild glycemia with minimal islet pathology as evaluated by the number of infiltrating cells and insulin content. There was higher number of apoptotic cells in the islets of galectin-3 knock out than in the control gal-3+/+ mice. RT PCR analysis of pancreatic lymph node cells 17 days after diabetes induction revealed the presence of IL-23 and IL-17 in gal-3+/+ but not in Gal-3 −/− mice. TNF-α and INF-γ INOS expression was also attenuated in Gal-3 −/− mice. We concluded that antiapoptotic effect of gal-3 in diabetogenic cells favors the induction of disease. The data are also compatible with our previous finding that IL-23-Th17 axis plays a role in diabetogenesis (Eur J Immunol, 36: 216–23, 2006). (Supported by Sheikh Hamdan Awards for Medical Research).

Dendritic Cells Expressing Transgenic Galectin-1 Delay Onset of Autoimmune Diabetes in Mice

Type 1 diabetes (T1D) is a disease caused by the destruction of the beta cells of the pancreas by activated T cells. Dendritic cells (DC) are the APC that initiate the T cell response that triggers T1D. However, DC also participate in T cell tolerance, and genetic engineering of DC to modulate T cell immunity is an area of active research. Galectin-1 (gal-1) is an endogenous lectin with regulatory effects on activated T cells including induction of apoptosis and down-regulation of the Th1 response, characteristics that make gal-1 an ideal transgene to transduce DC to treat T1D. We engineered bone marrow-derived DC to synthesize transgenic gal-1 (gal-1-DC) and tested their potential to prevent T1D through their regulatory effects on activated T cells. NOD-derived gal-1-DC triggered rapid apoptosis of diabetogenic BDC2.5 TCR-transgenic CD4+ T cells by TCR-dependent and -independent mechanisms. Intravenously administered gal-1-DC trafficked to pancreatic lymph nodes and spleen and delayed onset of diabetes and insulitis in the NODrag1(-/-) lymphocyte adoptive transfer model. The therapeutic effect of gal-1-DC was accompanied by increased percentage of apoptotic T cells and reduced number of IFN-gamma-secreting CD4+ T cells in pancreatic lymph nodes. Treatment with gal-1-DC inhibited proliferation and secretion of IFN-gamma of T cells in response to beta cell Ag. Unlike other DC-based approaches to modulate T cell immunity, the use of the regulatory properties of gal-1-DC on activated T cells might help to delete beta cell-reactive T cells at early stages of the disease when the diabetogenic T cells are already activated.

CD38 Is Required for the Peripheral Survival of Immunotolerogenic CD4+ Invariant NK T Cells in Nonobese Diabetic Mice

T cell-mediated autoimmune type-1 diabetes (T1D) in NOD mice partly results from this strain's numerical and functional defects in invariant NK T (iNKT) cells. T1D is inhibited in NOD mice treated with the iNKT cell superagonist alpha-galactosylceramide through a process involving enhanced accumulation of immunotolerogenic dendritic cells in pancreatic lymph nodes. Conversely, T1D is accelerated in NOD mice lacking CD38 molecules that play a role in dendritic cell migration to inflamed tissues. Unlike in standard NOD mice, alpha-galactosylceramide pretreatment did not protect the CD38-deficient stock from T1D induced by an adoptively transferred pancreatic beta cell-autoreactive CD8 T cell clone (AI4). We found that in the absence of CD38, ADP-ribosyltransferase 2 preferentially activates apoptotic deletion of peripheral iNKT cells, especially the CD4+ subset. Therefore, this study documents a previously unrecognized role for CD38 in maintaining survival of an iNKT cell subset that preferentially contributes to the maintenance of immunological tolerance.

Abrogation of recurrent autoimmunity in the NOD mouse: A critical role for host interleukin 4

We previously established a clinically relevant strategy to abrogate recurrent autoimmunity and enable long-term islet graft survival, involving antilymphocyte serum (ALS)-depletion of recipient T cells and intraportal administration of donor pancreatic lymph node cells (PLNCs) along with islet grafts. In this study, we investigated whether Th2 cytokines were required for the tolerizing ability of ALS/PLNC treatment in islet transplantation. ALS-treated diabetic NOD recipient mice, and NOD recipient mice deficient in interleukin 4 (IL-4-/-) or 10 (IL-4/10-/-) were transplanted with NOR or NOD.scid islets intraportally along with donor PLNC. Blood glucose levels were monitored to access graft function, sections of graft-bearing livers were histologically examined, and ELISPOT assays were used to assess cytokine profile and frequency of islet-reactive CD4 T cells. We found that ALS/PLNC was not effective in prolonging islet graft survival in diabetic NOD hosts deficient in either IL-4 (NOD.IL-4-/-) or in IL-4 and IL-10 (NOD.IL4-/-/10-/-) (mean survival time, 36 days), contrasting the long-term survival of islet grafts in wild-type NOD mice (mean survival time, > 80 days). In contrast, PLNC deficient in IL-4 promoted long-term graft survival in wild-type NOD hosts similar to that in wild-type PLNC. In wild-type NOD recipients of either wild-type PLNC or IL-4-/- PLNC, the host autoantigen-specific CD4 T cells produced predominately IL-4 coincident with long-term graft survival, whereas, in NOD.IL-4-/- recipients with rejected grafts, the autoreactive T cells produced interferon gamma and low amounts of IL-4. These data demonstrate that abrogation of recurrent autoimmunity requires host IL-4 and that manipulation of the autoreactive cytokine profile in long-term diabetes may be an effective strategy for islet transplant therapies.

T-Cell Homing to the Pancreas in Autoimmune Mouse Models of Diabetes: In Vivo MR Imaging

To evaluate the efficiency of T-cell labeling with anionic magnetic nanoparticles (AMNPs) and in vivo magnetic resonance (MR) imaging monitoring of T-cell homing to the pancreas. In vivo MR images of pancreas were obtained with a 7-T MR system in 12 NOD (nonobese diabetic) mice at 11 and 20 days after injection of AMNP-loaded or unloaded T cells. Homing of loaded T cells in pancreatic lymph nodes was detected by the presence of a focal dark spot with T2* effect in a caudal area of the pancreas. Detection of loaded T cells in pancreatic islets was evaluated by comparison of histograms of MR signal intensity generated in whole pancreas in mice injected with loaded and unloaded T cells. Homing of loaded T cells was confirmed at transmission electronic microscopy (TEM). Fifty-six mice underwent all experiments. Focal dark spots with T2* effect were observed at 11 days in all three mice injected with loaded T cells and in none of the three mice injected with unloaded T cells. At 20 days, a more diffuse negative enhancement of the whole pancreas was noticed in one mouse injected with loaded T cells than in three mice injected with unloaded T cells. Presence of loaded T cells was confirmed with TEM. In vitro and in vivo tests confirmed that survival and function were not altered by loading. The ability of MR imaging to depict cell homing in living organisms at least 20 days after cell labeling was demonstrated, opening the way of follow-up in autoimmune diseases and cell therapy.