Tannic Acid Capsules Decrease Redox-dependent Pro-inflammatory Responses to Prolong Islet Graft Survival After Transplantation

Type 1 diabetes (T1D) is an autoimmune disease resulting in pancreatic b-cell lysis via reactive oxygen species (ROS), pro-inflammatory cytokines, and islet-infiltrating leukocytes. One promising treatment for T1D is islet transplantation, but its clinical application is constrained due to islet availability, adverse effects of immunosuppressants on islet function, and declining graft survival. Islet encapsulation may provide an immunoprotective barrier to aid in preserving function and curbing immune-mediated rejection post transplantation. We previously demonstrated that a cytoprotective nanothin multilayer coating consisting of tannic acid (TA), an immunomodulatory antioxidant, and poly (N-vinylpyrrolidone) (PVPON), curtailed immune responses involved in transplant rejection. We hypothesized that (PVPON/TA) coating may hinder the synthesis of redox-sensitive chemokines that recruit immune cells to sites of islet engraftment and blunt ROS-dependent cues necessary for M1 macrophage differentiation. (PVPON/TA) coatings elicited a 7-fold decrease in M1 macrophage-derived pro-inflammatory chemokines CCL5 and CXCL10, blunted M1 macrophage activation markers, and curbed diabetogenic CD4 T cell migration approximately 2-fold. Importantly, murine islet encapsulation with (PVPON/TA) material restored euglycemia in immunodeficient-diabetic mice post transplantation. Our studies demonstrate that (PVPON/TA) multilayer coatings represent a promising immunomodulatory biomaterial for islet transplantation. Future studies will evaluate the efficiency of PVPON/TA coated islets in curbing hyperglycemia by allotransplantation into diabetic rodent models.

Islet Encapsulation: A Long-Term Treatment for Type 1 Diabetes

Type 1 diabetes (T1D) is a chronic condition in which patients suffer from high blood glucose levels due to the body's inability to produce sufficient insulin. Continuous insulin administration and T1D management are difficult, often leading to hypoglycemic events, insulin resistance, and lower quality of life. Major advancements have been made in recent years, including clinical islet transplantation, but their application is limited by rapid immune rejection and islet destruction. Thus, a necessary paradigm shift has been observed in recent times toward biomaterial-based islet transplantation therapy. The use of biomaterial-based encapsulation addresses major limitations, including immune rejection and hypoxia, and provides a proper cell microenvironment offering greater islet viability. Presently, researchers are more focused on developing a clinically translatable therapy for T1D with the existing knowledge of advanced biomaterial technology. In this review article, we provide a historical perspective, highlighting the developments in the field of islet encapsulation and transplantation, and focus on cutting-edge advancements with modern bioengineering from a clinical perspective.

Recent papers have shown that the initial event in the pathogenesis of autoimmune type 1 diabetes (T1D) comprises sensing of molecular patterns released from apoptotic β-cells by innate immune receptors such as toll-like receptor (TLR). We have reported that apoptotic β-cells undergoing secondary necrosis called 'late apoptotic' β-cells stimulate dendritic cells (DCs) and induce diabetogenic T cell priming through TLR2. The role of other innate immune receptors such as TLR7 or TLR9 in the initiation of T1D has also been suggested. We hypothesized that TLR2 blockade could inhibit T1D at the initial step of T1D. Indeed, when a TLR2 agonist, Pam3CSK4 was administered chronically, the development of T1D in nonobese diabetic (NOD) mice was inhibited. Diabetogenic T cell priming by DCs was attenuated by chronic treatment with Pam3CSK4, indicating DC tolerance. For the treatment of established T1D, immune tolerance alone is not enough because β-cell mass is critically reduced. We employed TLR2 tolerance in conjunction with islet transplantation, which led to reversal of newly established T1D. Dipeptidyl peptidase 4 (DPP4) inhibitors are a new class of anti-diabetic agents that have beneficial effects on β-cells. We investigated whether a combination of DPP4 inhibition and TLR2 tolerization could reverse newly established T1D without islet transplantation. We could achieve normoglycemia by TLR2 tolerization in combination with DPP4 inhibition but not by TLR2 tolerization or DPP4 inhibition alone. β-cell mass was significantly increased by combined treatment with TLR2 tolerization and DPP4 inhibition. These results suggest the possibility that a novel strategy of TLR tolerization will be available for the inhibition or treatment of established T1D when combined with measures increasing critically reduced β-cell mass of T1D patients such as DPP4 inhibition or stem cell technology.

Background/Objectives: A stem cell therapy for type 1 diabetes (T1D) is experimentally available but only to those few humans in whom the use of systemic immunosuppression can be justified. For others with T1D, a means to deliver the islets needs to be perfected. We have previously bioengineered a removable device for this purpose and now wish to test the effect of adding extracellular matrix (ECM) derived from decellularised human pancreas to it. Methods: The complete device consists of encapsulated pluripotent stem cell differentiated islets seeded into tubular scaffolds of polycaprolactone made by melt electrospin writing and to which ECM was added. The seeded device was implanted either subcutaneously (SC) or intraperitoneally (IP) into streptozotocin diabetic immunodeficient mice. The outcome over the next few months was compared with that achieved in diabetic mice implanted IP with encapsulated islets alone. Results: The device seeded with encapsulated islets but not containing ECM functioned less well than encapsulated islets implanted alone, with lower human C-peptide production. However, when ECM was added to the seeded device and whether implanted SC or IP, islets functioned as efficiently as those implanted without use of a scaffold. Conclusions: These data provide optimism for the use of seeded scaffolds in diabetic humans in whom a single scaffold seeded with multiple encapsulated islets can more readily be removed if needed for safety reasons than can multiple encapsulated islets not seeded into a scaffold.

Tissue ischemia promotes vasculogenesis through chemokine-induced recruitment of bone marrow-derived endothelial progenitor cells (EPCs). Diabetes significantly impairs this process. Because hyperglycemia increases reactive oxygen species in a number of cell types, and because many of the defects responsible for impaired vasculogenesis involve HIF1-regulated genes, we hypothesized that HIF1 function is impaired in diabetes because of reactive oxygen species-induced modification of HIF1alpha by the glyoxalase 1 (GLO1) substrate methylglyoxal. Decreasing superoxide in diabetic mice by either transgenic expression of manganese superoxide dismutase or by administration of an superoxide dismutase mimetic corrected post-ischemic defects in neovascularization, oxygen delivery, and chemokine expression, and normalized tissue survival. In hypoxic fibroblasts cultured in high glucose, overexpression of GLO1 prevented reduced expression of both the EPC mobilizing chemokine stromal cell-derived factor-1 (SDF-1) and of vascular epidermal growth factor, which modulates growth and differentiation of recruited EPCs. In hypoxic EPCs cultured in high glucose, overexpression of GLO1 prevented reduced expression of both the SDF-1 receptor CXCR4, and endothelial nitric-oxide synthase, an enzyme essential for EPC mobilization. HIF1alpha modification by methylglyoxal reduced heterodimer formation and HIF1alpha binding to all relevant promoters. These results provide a basis for the rational design of new therapeutics to normalize impaired ischemia-induced vasculogenesis in patients with diabetes.

ObjectivePresent clinicians with an updated overview of empagliflozin for the treatment of type 2 diabetes mellitus (T2DM), with focus on its use in combination regimens. MethodsUsing the Medline database, keyword searches were undertaken to identify literature reporting the use of empagliflozin treatment in clinical trials with a minimum duration of 12 weeks relating to patients with T2DM.

Type 1 diabetes (T1D) is a chronic autoimmune disease in which autoreactive T-cells and inflammation cause severe loss of pancreatic beta cells. Insulitis, the pathologic hallmark of T1D, is an inflammatory lesion consisting of immune cell infiltrates around and within the islets. New research initiatives and methodologies are advancing our understanding of pancreas pathology. Studies have revealed the predominant cellular types that infiltrate the islets, novel molecular aspects associated with insulitis, and the coexistence of additional pathological abnormalities. While insulitis is a critical element of T1D pathology and pathogenesis, it is typically present only in a modest proportion of islets at any given time, even at diagnosis, with overall limited relation to disease duration. Thus, the relative importance of insulitis as a determining factor of diabetes symptoms at disease onset appears to have been overestimated; growing evidence also shows that beta cell loss at diagnosis is more modest than previously thought. Thus, the sole targeting of the immune system may not afford full therapeutic efficacy if dysfunction affects beta cells that are not under immune attack and this is a key contributor to symptoms. Combination therapies that promote both immunoregulation and address beta cell dysfunction should be more effective in treating this chronic disease process. It remains a major goal to clarify the relation of insulitis with the dynamics of beta cell loss and coexisting mechanisms of dysfunction, according to clinical stage; such improved understanding is key to design therapeutic strategies that target multiple pathogenic mechanisms.

The Myrica rubra pomace polyphenols (MRPP) were used to study their hypoglycemic effects and mechanisms using Type 2 diabetes (T2D) (db/db) mice as the model. The results showed that the oral administration of MRPP (low-dose myricitrin, 50mg/kg BW by gavage; high-dose myricitrin, 250mg/kg BW by gavage; and Myrica rubra pomace, 500mg/kg BW by gavage) for 4 weeks significantly reduced fasting blood glucose, glycated serum protein, serum insulin, and insulin resistance index in mice, as well as the dyslipidemia in mice was improved. MRPP was able to improve the structural morphology of hepatocytes and pancreatic β-cells in diabetic mice. Its hypoglycemic mechanism may involve the upregulation of GLUT-4 and IRS-1 genes expression in the PI3K and AMPK signaling pathways, downregulation of GSK-3β, AMPK, PI3K, and AKT genes expression, and enhancement of the activity of enzymes related to glycogen synthesis and glucose metabolism. Research findings on 16S rRNA suggest that MRPP has the ability to alter the composition of the gut microbiota, impede the growth of harmful bacteria, and foster the growth of beneficial bacteria. Therefore, MRPP was capable of reshaping the specific gut microbial community and supporting its application as a novel supplement in functional foods for the treatment of T2D.

Type 1 diabetes (T1D) is caused by autoimmune disease that leads to the destruction of pancreatic β-cells. Transplantation of cadaveric pancreatic organs or pancreatic islets can restore normal physiology. However, there is a chronic shortage of cadaveric organs, limiting the treatment of the majority of patients on the pancreas transplantation waiting list. Here, we hypothesized that human iPS cells can be directly differentiated into insulin producing cells (IPCs) capable of secreting insulin. Using a series of pancreatic growth factors, we successfully generated iPS cells derived IPCs. Furthermore, to investigate the capability of these cells to secrete insulin in vivo, the differentiated cells were transplanted under the kidney capsules of diabetic immunodeficient mice. Serum glucose levels gradually declined to either normal or near normal levels over 150 days, suggesting that the IPCs were secreting insulin. In addition, using MRI, a 3D organoid appeared as a white patch on the transplanted kidneys but not on the control kidneys. These organoids showed neo-vascularization and stained positive for insulin and glucagon. All together, these data show that a pancreatic organ can be created in vivo providing evidence that iPS cells might be a novel option for the treatment of T1D.

Type 1 Diabetes (T1D) therapies have focused on restoring beta cell function via exogenous insulin or islet transplant. While promising, these approaches fail to account for the role of alpha cells and intra-islet communication in disease etiology. Alpha cells secrete glucagon and maintain effective paracrine signaling with beta cells to maintain glucose homeostasis and metabolic control. Mounting evidences suggest that impaired alpha cell function is a significant contributor to disease progression. Understanding triggers of alpha cell dysfunction and the resulting effects will be crucial for restoring both beta and alpha cell function in T1D. Lack of human alpha cell models have limited progress towards this goal. We recently reported a protocol to generate human alpha cells from stem cell sources (SC-alpha). Using these SC-alpha cells, we have established a T1D disease model to identify the triggers that induce alpha cell dysfunction. Importantly, we have identified defects in proglucagon processing that mimic those observed in diabetes and have identified conditions that modulate proglucagon processing and secretion. Here we demonstrate that exposure of SC-alpha cells to diabetogenic stressors such as hyperglycemia, hyperlipidemia, and ER stress impacts the processing of proglucagon and ultimately results in alpha cell dysfunction. We show that upon exposure to these stressors, SC-alpha cells secrete elevated glucagon compared to the control conditions and show a loss of glucose responsiveness. We also observed changes in the secretion of alternate proglucagon processing products, GLP-1, GLP-2, and glicentin. Interestingly, glicentin levels were affected by ER stress induction, but were unaffected by other stressors, suggesting that ER stress plays a direct role in the disruption of proglucagon processing, leading to alpha cell dysfunction. Identifying triggers of alpha cell dysfunction and assessing impacts in proglucagon processing will be a step towards restoring islet paracrine signaling. Disclosure S. Shrestha: None. Q. P. Peterson: None.

Type 1 diabetes (T1D) is a chronic autoimmune disease that leads to the progressive loss of pancreatic beta cells. Interferons (IFNs) contribute to the initiation and amplification of beta cell autoimmunity. STAT1 is the main mediator of IFN signalling but little is known on its complex activation processes and role in the progression of beta cell failure. We presently show that two T1D candidate genes (i.e. PTPN2 and BACH2) modulate STAT1 activation via two different pathways, namely the JAK/STAT, involved in the short-term phosphorylation of its tyrosine residue (Y701), and the MAPKs pathway, involved in the long-term phosphorylation of its serine residue (S727). Each STAT1 phosphorylation type can independently induce expression of the chemokine CXCL10, but both residues are necessary for the expression of MHC class I molecules. IFN-α-induced STAT1 activation is dynamic and residue-dependent, being STAT1-Y701 fast (detectable after 4h) but transitory (back to basal by 24h) while STAT1-S727 increases slowly (peak at 48h) and is associated with the long-term effects of IFN-α exposure. These pathways can be chemically dissociated in human beta cells by the use of JAK1/2, TYK2 or JNK1 inhibitors. The present findings provide a novel understanding of the dynamics of STAT1 activation and will be useful to develop novel and hopefully targeted (i.e. favouring individuals with particular polymorphisms) therapies for T1D and other autoimmune diseases.

Stem Cell Derived Beta Cells: The State of the Science.

Islet autoantibodies are the primary biomarkers used to predict type 1 diabetes (T1D) disease risk. They signal immune tolerance breach by islet autoantigen-specific B lymphocytes. T-B lymphocyte interactions that lead to expansion of pathogenic T cells underlie T1D development. Promising strategies to broadly prevent this T-B crosstalk include T cell elimination (anti-CD3, teplizumab), B cell elimination (anti-CD20, rituximab), and disruption of T cell costimulation/activation (CTLA-4/Fc fusion, abatacept). However, global disruption or depletion of immune cell subsets is associated with significant risk, particularly in children. Therefore, antigen-specific therapy is an area of active investigation for T1D prevention. We provide an overview of strategies to eliminate antigen-specific B lymphocytes as a means to limit pathogenic T cell expansion to prevent beta cell attack in T1D. Such approaches could be used to prevent T1D in at-risk individuals. Patients with established T1D would also benefit from such targeted therapies if endogenous beta cell function can be recovered or islet transplant becomes clinically feasible for T1D treatment.

Mitochondrial (mito) bioenergetics of circulating cells change as a function of disease in Alzheimer’s, fibromyalgia, cancer, and Type 2 diabetes. We are exploring the use of mito-bioenergetics as a biomarker of Type 1 diabetes (T1D) progression and immune cell activation. T1D is a polygenic autoimmune disease in which an inflammatory leukocytic infiltrate into pancreatic islets results in destruction of insulin-producing β-cells. Autoantibody synthesis is currently used as a biomarker, but is a poor predictor of T1D since the majority of β-cells are destroyed prior to autoantibody seroconversion. We hypothesize that T1D is a chronic inflammatory disease in which conditions of oxidative stress will diminish the mitochondrial reserve capacity of immune cells and provide a surrogate marker for T1D. We compared mito-bioenergetics of murine immune cells derived from the spleen, pancreatic lymph node and bone marrow between diabetic and pre-diabetic Non-Obese Diabetic (NOD) mice. We show that mitochondrial reserve capacity is reduced in circulating CD4+ T cells from diabetic mice, but not in pre-diabetic mice during T1D progression. To explore the role of mitochondrial oxidative stress on diabetogenic CD4+ T cell effector responses, we used mito-TEMPO to scavenge mitochondrial reactive oxygen species (mROS). Mito-TEMPO reduced IFNγ synthesis in a primary recall assay by 0.24-fold and the combination of mito and cytosolic TEMPO increased Treg-associated IL-2 by 2.5-fold, suggesting that mROS have a causal role in modulating T cell effector responses critical for T1D pathology. We plan to perform human translational studies on mito-bioenergetics of PBMCs from recent onset T1D patients and further dissect the role of mROS on T1D.