Normal Beta Cells Research Articles

The Membrane Phospholipidomics Research of Oxidatively Damaged INS-1 Pancreatic Beta Cells Intervened by the Effective Constituents ofAnemarrhenae AsphodeloidesRhizoma.

The rhizoma of Anemarrhenae asphodeloides has a long history of hypoglycemic use in Chinese traditional medicine. In this paper, 400 μmol/L H2O2 induced normal INS-1 pancreatic beta cells to establish experimental model of oxidative damage. Quercetin was used as a positive drug, and mangiferin and its ethanolic extract were selected as therapeutic agents in an oxidative damage model to evaluate the ameliorative effect of the active ingredients of Anemarrhenae asphodeloides rhizoma on oxidative damage in INS-1 pancreatic β-cells. Buliding a qualitative analysis method of membrane phospholipids of INS-1 pancreatic beta cells and identified 82 phospholipids based on the UPLC/Q-TOF MS technology, which could provide a database for further statistics analysis. OPLS-DA was used to screen the phospholipid biomarkers from the raw data. Exploring the biological significances of these biomarkers, and discussing the toxic effect of the effective components of Anemarrhenae asphodeloidesrhizoma , on oxidatively damaged INS-1 pancreatic beta cell.

Direct Differentiation of Bone Marrow Mononucleated Cells Into Insulin-Producing Cells Using 4 Specific Soluble Factors.

Bone marrow-derived stem cells are self-renewing and multipotent adult stem cells that differentiate into several types of cells. Here, we investigated a unique combination of 4 differentiation-inducing factors (DIFs), including putrescine (Put), glucosamine (GlcN), nicotinamide, and BP-1-102, to develop a differentiation method for inducing mature insulin-producing cells (IPCs) and apply this method to bone marrow mononucleated cells (BMNCs) isolated from mice. BMNCs, primed with the 4 soluble DIFs, were differentiated into functional IPCs. BMNCs cultured under the defined conditions synergistically expressed multiple genes, including those for PDX1, NKX6.1, MAFA, NEUROG3, GLUT2, and insulin, related to pancreatic beta cell development and function. They produced insulin/C-peptide and PDX1, as assessed using immunofluorescence and flow cytometry. The induced cells secreted insulin in a glucose-responsive manner, similar to normal pancreatic beta cells. Grafting BMNC-derived IPCs under kidney capsules of mice with streptozotocin (STZ)-induced diabetes alleviated hyperglycemia by lowering blood glucose levels, enhancing glucose tolerance, and improving glucose-stimulated insulin secretion. Insulin- and PDX1-expressing cells were observed in the IPC-bearing graft sections of nephrectomized mice. Therefore, this study provides a simple protocol for BMNC differentiation, which can be a novel approach for cell-based therapy in diabetes mellitus.

In-silico Study for African Plants with Possible Beta-Cell Regeneration Effect through Inhibition of DYRK1A

Background: The continuous destruction of normal insulin-producing pancreatic beta cells is a contributing factor in all common forms of diabetes, due to insufficient production of insulin, especially in type 1 diabetes. There are attempts to betacells transplantation, but the cost and availability of donors pose a great challenge to the process. Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase A plays a crucial role in beta-cells destruction. Aims: Our research targets to identify plants for that can be utilized possible alternative approach of beta-cell replacement through a pharmacologically induced regeneration of new beta cells in-silico. Methods and Material: The 3D structure Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase A and 6511 phytochemicals were obtained from Protein Databank and African Natural Products Database respectively. They were appropriately prepared for molecular docking simulations. Molecular docking simulations were implemented, after validation of docking protocols, in AutoDock-Vina®, using virtual screening scripts. Phytocompounds with good binding affinities for Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase A were selected as frontrunners. The compounds were screened for toxicity and Lipinski’s rule confirmation using Datawarrior and then kinase inhibitory bioactivity prediction using Molinspiration. Results: Twelve phytocompounds were found to be predictably highly active in-silico against Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase A, druglike based on Lipinski’s rule, non-mutagenic, non-tumorigenic, no reproductive effect and non-irritant, with high bioactivity prediction. Conclusions: In-silico active phytocompounds against Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase A with their plant sources and physicochemical parameters were identified. Further studies will be carried out in-vitro and in-vivo in to validate the results of this study using plants containing the identified phytocompounds.

Overexpression of PDX1 and NKX6.1 as a Treatment for Type 1 Diabetes: A Research Protocol

Introduction: Type 1 diabetes is an autoimmune disorder characterized by the destruction of pancreatic islet beta cells responsible for insulin production, leading to insulin deficiency. The PDX1 and NKX6-1 genes are essential to beta cell development and maturation in the pancreas. Based on previous studies, Pdx1 is downregulated in diabetics, and higher Nkx6.1 expression has been shown to induce high cell turnover. This proposal aims to investigate the potential therapeutic effects of gene therapy on streptozotocin-induced non-obese diabetic/severe-combined immunodeficient mice in treating Type 1 Diabetes. It is hypothesized that the use of gene therapy to overexpress PDX1 and NKX6-1 into the pancreas of experimental diabetic mice will result in increased beta cell production and improved glucose metabolism. Methods: Both male and female streptozotocin-induced non-obese diabetic/severe-combined immunodeficient experimental mice will receive human islets from individuals 32-55 years old. The PDX1 and NKX6-1 genes will be overexpressed in a shuttle vector and incorporated into the gutless adenovirus vector backbone through co-transfection using the Cre293/lox P cell line. Verification of successful vectors containing the gutless adenovirus coupled with a helper plasmid will be done using gel electrophoresis. Incubation with beta cells will stimulate the transduction process after which beta cells will be delivered into mice pancreas’ through endoscopic retrograde cholangiopancreatography. An intraperitoneal glucose tolerance test, pancreatic biopsy with immunohistochemical staining, and islet calculations will be done. Results: An intraperitoneal glucose tolerance test is expected to show improved blood glucose levels in experimental mice post-treatment, while a pancreatic biopsy with immunohistochemical staining and subsequent islet diameter and volume calculations are expected to depict an increase in the number of beta cells. Discussion: The experimental mice are anticipated to show an improvement in blood glucose levels and increased production of beta cells following treatment due to the overexpression of PDX1 and NKX6-1. The normal control mice and diabetic untreated mice are expected to show normal and high glucose levels, as well as normal and deficient beta cells respectively. Conclusion: This proposal may provide a better understanding of the pathophysiology in Type 1 Diabetes and pave a new path for its treatment.

The decrease of histomorphometry and function of pancreas in male albino rats after induced by sleep deprivation

Background: Sleep disturbance accelerates type 2 diabetes mellitus by reducing insulin secretion and by the occurrence of difficulty in controlling sleep behaviour in humans so that it is necessary conduct a study by using experimental animals.Objective: This study aims to determine histophotometric differences of pancreas and differences of glucose tolerance in male albino rats (Rattus norvegicus) after induced by stress paradoxical sleep deprivation (PSD) and total sleep deprivation (TSD). Methods: This study applied a post-test only with control groups consisting of 30 male albino rats (Wistar strain) divided randomly into 3 groups: control group, PSD group and TSD group. The mean number and diameter of islets of Langerhans were calculated per slide and were averaged for each group at a magnification of 400 X. The percentage of beta cells in one islet of Langerhans was (ß-p) that was a number of normal beta cells (Bn) divided for the total beta cells (Bt) (normal and damaged) x 100%. The glucose level based on the sampling time was made a curve shape, and the area under the 0-to-120-minute curves (AUC0-120) was calculated by using a trapezoidal formula.Results: The lowest AUC0-120 level of glucose was the control group (14,082 ± 955mg/dL) and the highest was the TSD group (16,293 ± 2,195 mg/dL); the Post-Hoc LSD test showed a significant difference (p<0.05) between the control-PSD group and the TSD-control group. The mean result of pancreatic ß cells mass with the lowest number was in the TSD group (76.3 ± 4.8), and the highest result was in the control group (94.3 ± 2.7); Post-Hoc LSD showed a significant difference (p <0.05) in the control-PSD group and the TSD-control group. The lowest mean in cell diameter of islets of Langerhans was in the TSD group (0.132 ± 0.031 mm), and the highest mean was in the control group (0.213 ± 0.019 mm). Conclusion: Sleep deprivation could reduce pancreatic ß cell mass and cell diameter of islets of Langerhans and increase glucose tolerance levels.

Insulin-positive ductal cells do not migrate into preexisting islets during pregnancy

The adult pancreatic ductal system was suggested to harbor facultative beta-cell progenitors similar to the embryonic pancreas, and the appearance of insulin-positive duct cells has been used as evidence for natural duct-to-beta-cell reprogramming. Nevertheless, the phenotype and fate of these insulin-positive cells in ducts have not been determined. Here, we used a cell-tagging dye, CFDA-SE, to permanently label pancreatic duct cells through an intraductal infusion technique. Representing a time when significant increases in beta-cell mass occur, pregnancy was later induced in these CFDA-SE-treated mice to assess the phenotype and fate of the insulin-positive cells in ducts. We found that a small portion of CFDA-SE-labeled duct cells became insulin-positive, but they were not fully functional beta-cells based on the in vitro glucose response and the expression levels of key beta-cell genes. Moreover, these insulin-positive cells in ducts expressed significantly lower levels of genes associated with extracellular matrix degradation and cell migration, which may thus prevent their budding and migration into preexisting islets. A similar conclusion was reached through analysis of the Gene Expression Omnibus database for both mice and humans. Together, our data suggest that the contribution of duct cells to normal beta-cells in adult islets is minimal at best.

Enhanced structure and function of human pluripotent stem cell-derived beta-cells cultured on extracellular matrix.

The differentiation of human stem cells into insulin secreting beta‐like cells holds great promise to treat diabetes. Current protocols drive stem cells through stages of directed differentiation and maturation and produce cells that secrete insulin in response to glucose. Further refinements are now needed to faithfully phenocopy the responses of normal beta cells. A critical factor in normal beta cell behavior is the islet microenvironment which plays a central role in beta cell survival, proliferation, gene expression and secretion. One important influence on native cell responses is the capillary basement membrane. In adult islets, each beta cell makes a point of contact with basement membrane protein secreted by vascular endothelial cells resulting in structural and functional polarization. Interaction with basement membrane proteins triggers local activation of focal adhesions, cell orientation, and targeting of insulin secretion. This study aims to identifying the role of basement membrane proteins on the structure and function of human embryonic stem cell and induced pluripotent stem cell‐derived beta cells. Here, we show that differentiated human stem cells‐derived spheroids do contain basement membrane proteins as a diffuse web‐like structure. However, the beta‐like cells within the spheroid do not polarize in response to this basement membrane. We demonstrate that 2D culture of the differentiated beta cells on to basement membrane proteins enforces cell polarity and favorably alters glucose dependent insulin secretion.

Aberrant methylation underlies insulin gene expression in human insulinoma

Human insulinomas are rare, benign, slowly proliferating, insulin-producing beta cell tumors that provide a molecular “recipe” or “roadmap” for pathways that control human beta cell regeneration. An earlier study revealed abnormal methylation in the imprinted p15.5-p15.4 region of chromosome 11, known to be abnormally methylated in another disorder of expanded beta cell mass and function: the focal variant of congenital hyperinsulinism. Here, we compare deep DNA methylome sequencing on 19 human insulinomas, and five sets of normal beta cells. We find a remarkably consistent, abnormal methylation pattern in insulinomas. The findings suggest that abnormal insulin (INS) promoter methylation and altered transcription factor expression create alternative drivers of INS expression, replacing canonical PDX1-driven beta cell specification with a pathological, looping, distal enhancer-based form of transcriptional regulation. Finally, NFaT transcription factors, rather than the canonical PDX1 enhancer complex, are predicted to drive INS transactivation.

Heparanase and Type 1 Diabetes.

Type 1 diabetes (T1D) results from autoimmune destruction of insulin-producing beta cells in pancreatic islets. The degradation of the glycosaminoglycan heparan sulfate (HS) by the endo-β-D-glycosidase heparanase plays a critical role in multiple stages of the disease process. Heparanase aids (i) migration of inflammatory leukocytes from the vasculature to the islets, (ii) intra-islet invasion by insulitis leukocytes, and (iii) selective destruction of beta cells. These disease stages are marked by the solubilization of HS in the subendothelial basement membrane (BM), HS breakdown in the peri-islet BM, and the degradation of HS inside beta cells, respectively. Significantly, healthy islet beta cells are enriched in highly sulfated HS which is essential for their viability, protection from damage by reactive oxygen species (ROS), beta cell function and differentiation. Consequently, mouse and human beta cells but not glucagon-producing alpha cells (which contain less-sulfated HS) are exquisitely vulnerable to heparanase-mediated damage. In vitro, the death of HS-depleted mouse and human beta cells can be prevented by HS replacement using highly sulfated HS mimetics or analogues. T1D progression in NOD mice and recent-onset T1D in humans correlate with increased expression of heparanase by circulating leukocytes of myeloid origin and heparanase-expressing insulitis leukocytes. Treatment of NOD mice with the heparanase inhibitor and HS replacer, PI-88, significantly reduced T1D incidence by 50%, impaired the development of insulitis and preserved beta cell HS. These outcomes identified heparanase as a novel destructive tool in T1D, distinct from the conventional cytotoxic and apoptosis-inducing mechanisms of autoreactive T cells. In contrast to exogenous catalytically active heparanase, endogenous heparanase may function in HS homeostasis, gene expression and insulin secretion in normal beta cells and immune gene expression in leukocytes. In established diabetes, the interplay between hyperglycemia, local inflammatory cells (e.g. macrophages) and heparanase contributes to secondary micro- and macro-vascular disease. We have identified dual activity heparanase inhibitors/HS replacers as a novel class of therapeutic for preventing T1D progression and potentially for mitigating secondary vascular disease that develops with long-term T1D.

Combined Inhibition of DYRK1A, SMAD, and Trithorax Pathways Synergizes to Induce Robust Replication in Adult Human Beta Cells

Small-molecule inhibitors of dual-specificity tyrosine-regulated kinase 1A (DYRK1A) induce human beta cells to proliferate, generating a labeling index of 1.5%-3%. Here, we demonstrate that combined pharmacologic inhibition of DYRK1A and transforming growth factor beta superfamily (TGFβSF)/SMAD signaling generates remarkable further synergistic increases in human beta cell proliferation (average labeling index, 5%-8%, and as high as 15%-18%), and increases in both mouse and human beta cell numbers. This synergy reflects activation of cyclins and cdks by DYRK1A inhibition, accompanied by simultaneous reductions in key cell-cycle inhibitors (CDKN1C and CDKN1A). The latter results from interference with the basal Trithorax- and SMAD-mediated transactivation of CDKN1C and CDKN1A. Notably, combined DYRK1A and TGFβ inhibition allows preservation of beta cell differentiated function. These beneficial effects extend from normal human beta cells and stem cell-derived human beta cells to those from people with type 2 diabetes, and occur both invitro and invivo.

Loss of intra-islet heparan sulfate is a highly sensitive marker of type 1 diabetes progression in humans.

Type 1 diabetes (T1D) is an autoimmune disease in which insulin-producing beta cells in pancreatic islets are progressively destroyed. Clinical trials of immunotherapies in recently diagnosed T1D patients have only transiently and partially impacted the disease course, suggesting that other approaches are required. Our previous studies have demonstrated that heparan sulfate (HS), a glycosaminoglycan conventionally expressed in extracellular matrix, is present at high levels inside normal mouse beta cells. Intracellular HS was shown to be critical for beta cell survival and protection from oxidative damage. T1D development in Non-Obese Diabetic (NOD) mice correlated with loss of islet HS and was prevented by inhibiting HS degradation by the endoglycosidase, heparanase. In this study we investigated the distribution of HS and heparan sulfate proteoglycan (HSPG) core proteins in normal human islets, a role for HS in human beta cell viability and the clinical relevance of intra-islet HS and HSPG levels, compared to insulin, in human T1D. In normal human islets, HS (identified by 10E4 mAb) co-localized with insulin but not glucagon and correlated with the HSPG core proteins for collagen type XVIII (Col18) and syndecan-1 (Sdc1). Insulin-positive islets of T1D pancreases showed significant loss of HS, Col18 and Sdc1 and heparanase was strongly expressed by islet-infiltrating leukocytes. Human beta cells cultured with HS mimetics showed significantly improved survival and protection against hydrogen peroxide-induced death, suggesting that loss of HS could contribute to beta cell death in T1D. We conclude that HS depletion in beta cells, possibly due to heparanase produced by insulitis leukocytes, may function as an important mechanism in the pathogenesis of human T1D. Our findings raise the possibility that intervention therapy with dual activity HS replacers/heparanase inhibitors could help to protect the residual beta cell mass in patients recently diagnosed with T1D.

Insights into beta cell regeneration for diabetes via integration of molecular landscapes in human insulinomas

Although diabetes results in part from a deficiency of normal pancreatic beta cells, inducing human beta cells to regenerate is difficult. Reasoning that insulinomas hold the “genomic recipe” for beta cell expansion, we surveyed 38 human insulinomas to obtain insights into therapeutic pathways for beta cell regeneration. An integrative analysis of whole-exome and RNA-sequencing data was employed to extensively characterize the genomic and molecular landscape of insulinomas relative to normal beta cells. Here, we show at the pathway level that the majority of the insulinomas display mutations, copy number variants and/or dysregulation of epigenetic modifying genes, most prominently in the polycomb and trithorax families. Importantly, these processes are coupled to co-expression network modules associated with cell proliferation, revealing candidates for inducing beta cell regeneration. Validation of key computational predictions supports the concept that understanding the molecular complexity of insulinoma may be a valuable approach to diabetes drug discovery.

Insulin receptor alternative splicing is regulated by insulin signaling and modulates beta cell survival.

Type 2 Diabetes (T2DM) affects more than 300 million people worldwide. One of the hallmarks of T2DM is peripheral insulin resistance, in part due to unproductive insulin signaling through the insulin receptor. The insulin receptor (INSR) exists as two isoforms, INSR-A and INSR-B, which results from skipping or inclusion of exon 11 respectively. What determines the relative abundance of the different insulin receptor splice variants is unknown. Moreover, it is not yet clear what the physiological roles of each of the isoforms are in normal and diseased beta cells. In this study, we show that insulin induces INSR exon 11 inclusion in pancreatic beta cells in both human and mouse. This occurs through activation of the Ras-MAPK/ERK signaling pathway and up-regulation of the splicing factor SRSF1. Induction of exon 11 skipping by a splice-site competitive antisense oligonucleotide inhibited the MAPK-ERK signaling pathway downstream of the insulin receptor, sensitizing the pancreatic β-cell line MIN6 to stress-induced apoptosis and lipotoxicity. These results assign to insulin a regulatory role in INSR alternative splicing through the Ras-MAPK/ERK signaling pathway. We suggest that in beta cells, INSR-B has a protective role, while INSR-A expression sensitizes beta cells to programmed cell death.

Breaking dogma for future therapy using stem cell - Where we have reached?

Breaking dogma for future therapy using stem cell - Where we have reached?

A cytosensor based on NiO nanoparticle-enhanced surface plasmon resonance for detection of the breast cancer cell line MCF-7

The article describes the use of a designed histidine (His)-tagged arginine-glycine-aspartic acid (RGD) peptide as a linker to bind integrin to captured cancer cells, and how to amplify the surface plasmon resonance (SPR) signal after binding of NiO nanoparticles (NiO-NPs) via the His-tag on the peptide. Specifically, breast cancer cells were captured via the interaction between human mucin (MUC-1) and a gold surface modified with a MUC-selective aptamer. The resulting cytosensor exhibits specificity and high sensitivity which is due to the enhancement of the SPR signal by NiO-NPs. The breast cancer cell line MCF-7 can be easily distinguished from normal islet beta cells by using this biosensor. Implementation of the His-tagged RGD peptide modified with NiO-NPs resulted in 20-fold enhancement of the SPR signal at the limit of detection. Hence, the actual limit of detection is lowered to 136 cells per mL. In our perception, this cytosensor has a large clinical potential in that it may also be used to detect various other kinds of tumor cells.

Different responses of mouse islets and MIN6 pseudo-islets to metabolic stimulation: a note of caution.

MIN6 cells and MIN6 pseudo-islets are popular surrogates for the use of primary beta cells and islets. Even though it is generally agreed that the stimulus-secretion coupling may deviate from that of beta cells or islets, direct comparisons are rare. The present side-by-side comparison of insulin secretion, cytosolic Ca(2+) concentration ([Ca(2+)] i ) and oxygen consumption rate (OCR) points out where similarities and differences exist between MIN6 cells and normal mouse beta cells. In mouse islets and MIN6 pseudo-islets depolarization by 40 mM KCl was a more robust insulinotropic stimulus than 30 mM glucose. In MIN6 pseudo-islets, but not in mouse islets, the response to 30 mM glucose was much lower than to 40 mM KCl and could be suppressed by a preceding stimulation with 40 mM KCl. In MIN6 pseudo-islets, glucose was less effective to raise [Ca(2+)] i than in primary islets. In marked contrast to islets, the OCR response of MIN6 pseudo-islets to 30 mM glucose was smaller than to 40 mM KCl and was further diminished by a preceding stimulation with 40 mM KCl. The same pattern was observed when MIN6 pseudo-islets were cultured in 5 mM glucose. As with insulin secretion memory effects on the OCR remained after wash-out of a stimulus. The differences between MIN6 cells and primary beta cells were generally larger in the responses to glucose than to depolarization by KCl. Thus, the use of MIN6 cells in investigations on metabolic signalling requires particular caution.

Mouse Insulin Cells Expressing an Inducible RIPCre Transgene Are Functionally Impaired

We used cre-lox technology to test whether the inducible expression of Cre minimize the deleterious effect of the enzyme on beta cell function. We studied mice in which Cre is linked to a modified estrogen receptor (ER), and its expression is controlled by the rat insulin promoter (RIP). Following the injection of tamoxifen (TM), CreER- migrates to the nucleus and promotes the appearance of a reporter protein, enhanced yellow fluorescent protein (EYFP), in cells. Immunocytochemical analysis indicated that 46.6 ± 2.1% insulin cells of adult RIPCreER- EYFP expressed EYFP. RIPCreER-EYFP (+TM) mice were normoglycemic throughout the study, and their glucose tolerance test results were similar to control CD-1 mice. However, an extended exposure to reagents that stimulate insulin synthesis was detrimental to the survival of IN+EYFP+cells. The administration of an inhibitor of the enzyme dipeptidyl-peptidase (DPP4i), which prevents the cleavage of glucagon-like peptide (GLP-1), to adult RIPCreER-EYFP mice lead to a decrease in the percentage of IN+EYFP+ to 17.5 ± 1.73 and a significant increase in apoptotic cells in islets. Similarly, a 2-week administration of the GLP-1 analog exendin 4 (ex-4) induced an almost complete ablation of IN+ expressing a different reporter protein and a significant decrease in the beta cell mass and rate of beta cell proliferation. Since normal beta cells do not die when induced to increase insulin synthesis, our observations indicate that insulin cells expressing an inducible RIPCre transgene are functionally deficient. Studies employing these mice should carefully consider the pitfalls of the Cre-Lox technique.

Both PAX4 and MAFA Are Expressed in a Substantial Proportion of Normal Human Pancreatic Alpha Cells and Deregulated in Patients with Type 2 Diabetes

Pax4 and MafA (v-maf musculoaponeurotic fibrosarcoma oncogene homolog A) are two transcription factors crucial for normal functions of islet beta cells in the mouse. Intriguingly, recent studies indicate the existence of notable difference between human and rodent islet in terms of gene expression and functions. To better understand the biological role of human PAX4 and MAFA, we investigated their expression in normal and diseased human islets, using validated antibodies. PAX4 was detected in 43.0±5.0% and 39.1±4.0% of normal human alpha and beta cells respectively. We found that MAFA, detected in 88.3±6.3% insulin+cells as in the mouse, turned out to be also expressed in 61.2±6.4% of human glucagons+ cells with less intensity than in insulin+ cells, whereas MAFB expression was found not only in the majority of glucagon+ cells (67.2±7.6%), but also in 53.6±10.5% of human insulin+ cells. Interestingly, MAFA nuclear expression in both alpha and beta cells, and the percentage of alpha cells expressing PAX4 were found altered in a substantial proportion of patients with type 2 diabetes. Both MAFA and PAX4 display, therefore, a distinct expression pattern in human islet cells, suggesting more potential plasticity of human islets as compared with rodent islets.

GLP-1 could improve the similarity of IPCs and pancreatic beta cells in cellular ultrastructure and function

Transplantation of functional insulin-producing cells (IPCs) provides a novel mode for insulin replacement, but is often accompanied by many undesirable side effects. Our previous studies suggested that IPCs could not mimic the physiological regulation of insulin secretion performed by pancreatic beta cells. To obtain a better method through which to acquire more similar IPCs, we compared the difference between IPCs of the GLP-1 group and IPCs of the non-GLP-1 group in the morphological features in cellular level and physiological function. The levels of insulin secretion were measured by ELISA. The insulin and glucagon-like peptide-1 (GLP-1) mRNA gene expression was determined by real-time quantitative PCR. The morphological features were detected by atomic force microscopy (AFM) and laser confocal scanning microscopy (LCSM). Intracellular Ca(2+) levels and Glucagon-like peptide-1 receptor (GLP-1R) levels were determined by flow cytometer (FCM). We found that IPCs of the GLP-1 group had bigger membrane particle size and average roughness (Ra ) than IPCs of the non-GLP-1 group but still smaller than normal human pancreatic beta cells. The physiology function of IPCs of the GLP-1 group were much closer to normal human pancreatic beta cells than IPCs of the non-GLP-1 group. GLP-1 could improve the similarity of IPCs from human adipose tissue-derived mesenchymal stem cells and pancreatic beta cells in cellular ultrastructure and function.

Educational Exhibit Abstract No. 386 - Calcium arterial stimulation venous sampling for preoperative localization of occult insulinomas

Learning Objectives (1) To review the clinical features, diagnosis, and treatment of insulinomas, (2) to discuss the indications and advantages of intra-arterial calcium-stimulated venous sampling (ASVS) vs. portal venous sampling and imaging for preoperative diagnosis of occult insulinomas, (3) to discuss the physiologic rationale of ASVS, technique, and result interpretation. Background Insulinomas cause symptomatic hypoglycemia and are the most common islet cell tumors. Surgical resection is curative. Preoperative localization of an insulinoma is a prerequisite of successful surgical resection. Cross-sectional imaging modalities unreliably detect insulinomas less than 2 cm in size. ASVS is a highly sensitive test for localizing such occult insulinomas. Administration of calcium gluconate into the arterial supply of normal beta cells causes insulin release. The insulin surge from beta cells of an insulinoma is much greater allowing for tumor localization based on site of arterial injection. In the setting of multifocal pancreatic lesions, ASVS can also be used to identify the region that is hormonally most active prior to the resection. Clinical Findings/Procedure Details ASVS is performed in a fasting state. Arteries supplying the pancreas are selected and calcium gluconate is injected. Blood samples are obtained from hepatic veins simultaneously with each injection and evaluated for insulin levels. Location of the insulinoma is determined based on increase in the hepatic venous insulin levels corresponding to the pancreatic arterial territory stimulated. We will discuss the diagnostic algorithm, illustrate the ASVS technique in detail, and interpret the test results. We will also explain the physiologic rationale of the test, angiographic findings of insulinoma and potential use of cone beam CT scan in optimizing localization and identification. Conclusion and/or Teaching Points ASVS is the most accurate method to localize occult insulinomas preoperatively. Familiarity with this technique has the potential to improve clinical outcomes after surgical resection.