NN1213 - A Potent, Long-Acting, and Selective Analog of Human Amylin.

Amylin, a member of the calcitonin family, acts via amylin receptors in the hindbrain and hypothalamus to suppress appetite. Native ligands of these receptors are peptides with short half-lives. Conjugating fatty acids to these peptides can increase their half-lives.

Identification of the novel HLA allele HLA-DRB1*03:201 by next-generation sequencing.

HLA-DRB1*03:201 differs from HLA-DRB1*03:01 in exon 3 at codon 178 resulting in a proline to serine substitution.

Detection of the novel HLA allele, HLA-DRB1*08:112, identified in a Danish family.

HLA-DRB1*08:112 differs from HLA-DRB1*08:01 in exon 2 at amino acid 62; asparagine to lysine substitution.

Detection of the novel HLA allele, HLA-DQA1*01:65, identified in a Danish donor.

HLA-DQA1*01:65 differs from HLA-DQA1*01:03 in exon 1 at amino acid -7 a valine to methionine substitution.

RAGE binds preamyloid IAPP intermediates and mediates pancreatic β cell proteotoxicity.

Islet amyloidosis is characterized by the aberrant accumulation of islet amyloid polypeptide (IAPP) in pancreatic islets, resulting in β cell toxicity, which exacerbates type 2 diabetes and islet transplant failure. It is not fully clear how IAPP induces cellular stress or how IAPP-induced toxicity can be prevented or treated. We recently defined the properties of toxic IAPP species.

The EndoC-βH1 cell line is a valid model of human beta cells and applicable for screenings to identify novel drug target candidates.

Objective: To characterize the EndoC-βH1 cell line as a model for human beta cells and evaluate its beta cell functionality, focusing on insulin secretion, proliferation, apoptosis and ER stress, with the objective to assess its potential as a screening platform for identification of novel anti-diabetic drug candidates.

Methods: EndoC-βH1 was transplanted into mice for validation of in vivo functionality. Insulin secretion was evaluated in cells cultured as monolayer and as pseudoislets, as well as in diabetic mice.

Time-resolved studies define the nature of toxic IAPP intermediates, providing insight for anti-amyloidosis therapeutics.

Islet amyloidosis by IAPP contributes to pancreatic β-cell death in diabetes, but the nature of toxic IAPP species remains elusive. Using concurrent time-resolved biophysical and biological measurements, we define the toxic species produced during IAPP amyloid formation and link their properties to induction of rat INS-1 β-cell and murine islet toxicity. These globally flexible, low order oligomers upregulate pro-inflammatory markers and induce reactive oxygen species.

Islet remodeling in female mice with spontaneous autoimmune and streptozotocin-induced diabetes.

Islet alpha- and delta-cells are spared autoimmune destruction directed at beta-cells in type 1 diabetes resulting in an apparent increase of non-beta endocrine cells in the islet core. We determined how islet remodeling in autoimmune diabetes compares to streptozotocin (STZ)-induced diabetes. Islet cell mass, proliferation, and immune cell infiltration in pancreas sections from diabetic NOD mice and mice with STZ-induced diabetes was assessed using quantitative image analysis.

Advances and challenges in islet transplantation: islet procurement rates and lessons learned from suboptimal islet transplantation.

The initial step in successful islet transplantation is procurement of healthy donor islets. Given the limited number of donor pancreata selected for islet isolation and that islets from multiple donors are typically required to obtain insulin independence, it is critical to improve pancreas procurement rates and yield of islets for transplantation. Islets are delicate microorgans that are susceptible to apoptosis, hypoxia, and ischemia during isolation, culture, and the peritransplant period.

Differentiation of mouse embryonic stem cells into endoderm without embryoid body formation.

Pluripotent embryonic stem cells hold a great promise as an unlimited source of tissue for treatment of chronic diseases such as Type 1 diabetes. Herein, we describe a protocol using all-trans-retinoic acid, basic fibroblast growth factor and dibutyryl cAMP (DBcAMP) in the absence of embryoid body formation, for differentiation of murine embryonic stem cells into definitive endoderm that may serve as pancreatic precursors. The produced cells were analyzed by quantitative PCR, immunohistochemistry and static insulin release assay for markers of trilaminar embryo, and pancreas.

XIAP inhibition of β-cell apoptosis reduces the number of islets required to restore euglycemia in a syngeneic islet transplantation model.

Clinical pancreatic islet transplantation has great promise as a treatment for type 1 diabetes but despite recent advances, it is still limited by the need for lifelong immunosuppression, restricted availability of donor islets, and uncertainty regarding long-term graft survival. Using a syngeneic, suboptimal islet transplantation model, we asked whether adenoviral overexpression of an anti-apoptotic protein, the X-linked inhibitor of apoptosis protein (XIAP) would protect transplanted islet cells from death and reduce the number of islets required for successful transplantation. Transplantation of 100 XIAP-expressing islets into the kidney capsule of syngeneic Balb/c mice restored euglycemia in 86% of recipients, where transplantation of 100 islets transduced with a control adenovirus expressing LacZ restored euglycemia in only 27% of recipients.

The flavanol (-)-epigallocatechin 3-gallate inhibits amyloid formation by islet amyloid polypeptide, disaggregates amyloid fibrils, and protects cultured cells against IAPP-induced toxicity.

Islet amyloid polypeptide (IAPP, amylin) is the major protein component of the islet amyloid deposits associated with type 2 diabetes. The polypeptide lacks a well-defined structure in its monomeric state but readily assembles to form amyloid. Amyloid fibrils formed from IAPP, intermediates generated in the assembly of IAPP amyloid, or both are toxic to β-cells, suggesting that islet amyloid formation may contribute to the pathology of type 2 diabetes.

The sulfated triphenyl methane derivative acid fuchsin is a potent inhibitor of amyloid formation by human islet amyloid polypeptide and protects against the toxic effects of amyloid formation.

Islet amyloid polypeptide (IAPP), also known as amylin, is responsible for amyloid formation in type 2 diabetes. The formation of islet amyloid is believed to contribute to the pathology of the disease by killing beta-cells, and it may also contribute to islet transplant failure. The design of inhibitors of amyloid formation is an active area of research, but comparatively little attention has been paid to inhibitors of IAPP in contrast to the large body of work on beta-amyloid, and most small-molecule inhibitors of IAPP amyloid are generally effective only when used at a significant molar excess.

Islet allograft rejection is independent of toll-like receptor signaling in mice.

Background: Islet transplantation is a promising therapy for type 1 diabetes; however, most islet grafts fail within 5 years. Innate immunity has been suggested to play a role in islet allograft rejection, potentially mediated by toll-like receptors (TLRs), a class of innate immune receptors. Lack of TLR4, in particular, has been reported to improve allograft survival.

The X-linked inhibitor of apoptosis protein enhances survival of murine islet allografts.

Allotransplantation of pancreatic islets represents a promising approach to treat type 1 diabetes. Destruction of beta-cells in islet allografts involves multiple immune mechanisms that lead to activation of caspases and apoptotic cell death. The X-linked inhibitor of apoptosis (XIAP) inhibits apoptosis induced by a variety of triggers, primarily by preventing the activation of caspases.

Synergistic inhibition of tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis in human pancreatic beta cells by Bcl-2 and X-linked inhibitor of apoptosis.

To better understand the cytokine death-signal transduction pathways in human beta cells, we investigated the inhibitory effects of Bcl-2 (protooncogene bcl-2) and X-linked inhibitor of apoptosis (XIAP) on TRAIL (TNF-related apoptosis-inducing ligand)-induced human beta-cell destruction. A panel of Bcl-2-overexpressing transfectants of the human beta-cell lines NES2Y and CM was developed by transfection with a pEFpGKpuro vector containing Bcl-2 or an empty vector as a control. TRAIL-induced cytotoxicity and apoptosis of Bcl-2-overexpressing beta cells were clearly decreased, in comparison with wild-type cells and the empty vector transfectants.

Increasing the yield of human mononuclear cells and low serum conditions for in vitro generation of macrophages with M-CSF.

The yield of mononuclear cells extracted from peripheral blood using standard protocols is frequently inadequate when working with material of limited availability. In addition, the regular usage of autologous and fetal calf serum (FCS) to generate human macrophages in vitro may complicate antigen uptake, processing and presentation on HLA molecules. We optimized the yield of mononuclear cells from 34+/-3% to 65+/-5% by collecting both the interface and more than half of the overlayering supernatant, followed by three washes at 4 degrees C.