Metformin Ameliorates Dysfunctional Traits of Glibenclamide- and Glucose-Induced Insulin Secretion by Suppression of Imposed Overactivity of the Islet Nitric Oxide Synthase-NO System.

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Ingmar Lundquist

Full Text Available Metformin lowers diabetic blood glucose primarily by reducing hepatic gluconeogenesis and increasing peripheral glucose uptake. However, possible effects by metformin on beta-cell function are incompletely understood. We speculated that metformin might positively influence insulin secretion through impacting the beta-cell nitric oxide synthase (NOS-NO system, a negative modulator of glucose-stimulated insulin release. In short-time incubations with isolated murine islets either glibenclamide or high glucose augmented insulin release associated with increased NO production from both neural and inducible NOS. Metformin addition suppressed the augmented NO generation coinciding with amplified insulin release. Islet culturing with glibenclamide or high glucose revealed pronounced fluorescence of inducible NOS in the beta-cells being abolished by metformin co-culturing. These findings were reflected in medium nitrite-nitrate levels. A glucose challenge following islet culturing with glibenclamide or high glucose revealed markedly impaired insulin response. Metformin co-culturing restored this response. Culturing murine islets and human islets from controls and type 2 diabetics with high glucose or high glucose + glibenclamide induced a pronounced decrease of cell viability being remarkably restored by metformin co-culturing. We show here, that imposed overactivity of the beta-cell NOS-NO system by glibenclamide or high glucose leads to insulin secretory dysfunction and reduced cell viability and also, importantly, that these effects are relieved by metformin inhibiting beta-cell NO overproduction from both neural and inducible NOS thus ameliorating a concealed negative influence by NO induced by sulfonylurea treatment and/or high glucose levels. This double-edged effect of glibenclamide on the beta-cellsuggests sulfonylurea monotherapy in type 2 diabetes being avoided.

Advances and Challenges in Islet Transplantation: Islet Procurement Rates and Lessons Learned from Suboptimal Islet Transplantation

OpenAIRE

Annette Plesner; C. Bruce Verchere

2011-01-01

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...

G protein-coupled receptor 39 deficiency is associated with pancreatic islet dysfunction

DEFF Research Database (Denmark)

Holst, Birgitte; Egerod, Kristoffer L; Jin, Chunyu

2009-01-01

G protein-coupled receptor (GPR)-39 is a seven-transmembrane receptor expressed mainly in endocrine and metabolic tissues that acts as a Zn(++) sensor signaling mainly through the G(q) and G(12/13) pathways. The expression of GPR39 is regulated by hepatocyte nuclear factor (HNF)-1alpha and HNF-4...... tolerance both during oral and iv glucose tolerance tests, and Gpr39(-/-) mice had decreased plasma insulin response to oral glucose. Islet architecture was normal in the Gpr39 null mice, but expression of Pdx-1 and Hnf-1alpha was reduced. Isolated, perifused islets from Gpr39 null mice secreted less...

Construction of EMSC-islet co-localizing composites for xenogeneic porcine islet transplantation.

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Kim, Jung-Sik; Chung, Hyunwoo; Byun, Nari; Kang, Seong-Jun; Lee, Sunho; Shin, Jun-Seop; Park, Chung-Gyu

2018-03-04

Pancreatic islet transplantation is an ultimate solution for treating patients with type 1 diabetes (T1D). The pig is an ideal donor of islets for replacing scarce human islets. Besides immunological hurdles, non-immunological hurdles including fragmentation and delayed engraftment of porcine islets need solutions to succeed in porcine islet xenotransplantation. In this study, we suggest a simple but effective modality, a cell/islet co-localizing composite, to overcome these challenges. Endothelial-like mesenchymal stem cells (EMSCs), differentiated from bone-marrow derived mouse mesenchymal stem cells (MSCs), and MSCs evenly coated the surface of porcine islets (>85%) through optimized culture conditions. Both MSCs and EMSCs significantly reduced the fragmentation of porcine islets and increased the islet masses, designated as islet equivalents (IEQs). In fibrin in vitro and in vivo angiogenesis analysis, constructed EMSC-islet composites showed higher angiogenic potentials than naked islets, MSC-islet composites, or human endothelial cell-islet composites. This novel delivery method of porcine islets may have beneficial effects on the engraftment of transplanted islets by prevention of fragmentation and enhancement of revascularization. Copyright © 2018 Elsevier Inc. All rights reserved.

High-fat diet-induced insulin resistance does not increase plasma anandamide levels or potentiate anandamide insulinotropic effect in isolated canine islets.

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Orison O Woolcott

Full Text Available Obesity has been associated with elevated plasma anandamide levels. In addition, anandamide has been shown to stimulate insulin secretion in vitro, suggesting that anandamide might be linked to hyperinsulinemia.To determine whether high-fat diet-induced insulin resistance increases anandamide levels and potentiates the insulinotropic effect of anandamide in isolated pancreatic islets.Dogs were fed a high-fat diet (n = 9 for 22 weeks. Abdominal fat depot was quantified by MRI. Insulin sensitivity was assessed by the euglycemic-hyperinsulinemic clamp. Fasting plasma endocannabinoid levels were analyzed by liquid chromatography-mass spectrometry. All metabolic assessments were performed before and after fat diet regimen. At the end of the study, pancreatic islets were isolated prior to euthanasia to test the in vitro effect of anandamide on islet hormones. mRNA expression of cannabinoid receptors was determined in intact islets. The findings in vitro were compared with those from animals fed a control diet (n = 7.Prolonged fat feeding increased abdominal fat content by 81.3±21.6% (mean±S.E.M, P<0.01. In vivo insulin sensitivity decreased by 31.3±12.1% (P<0.05, concomitant with a decrease in plasma 2-arachidonoyl glycerol (from 39.1±5.2 to 15.7±2.0 nmol/L but not anandamide, oleoyl ethanolamide, linoleoyl ethanolamide, or palmitoyl ethanolamide. In control-diet animals (body weight: 28.8±1.0 kg, islets incubated with anandamide had a higher basal and glucose-stimulated insulin secretion as compared with no treatment. Islets from fat-fed animals (34.5±1.3 kg; P<0.05 versus control did not exhibit further potentiation of anandamide-induced insulin secretion as compared with control-diet animals. Glucagon but not somatostatin secretion in vitro was also increased in response to anandamide, but there was no difference between groups (P = 0.705. No differences in gene expression of CB1R or CB2R between groups were found.In canines, high-fat diet

Islet product characteristics and factors related to successful human islet transplantation from the Collaborative Islet Transplant Registry (CITR) 1999-2010.

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Balamurugan, A N; Naziruddin, B; Lockridge, A; Tiwari, M; Loganathan, G; Takita, M; Matsumoto, S; Papas, K; Trieger, M; Rainis, H; Kin, T; Kay, T W; Wease, S; Messinger, S; Ricordi, C; Alejandro, R; Markmann, J; Kerr-Conti, J; Rickels, M R; Liu, C; Zhang, X; Witkowski, P; Posselt, A; Maffi, P; Secchi, A; Berney, T; O'Connell, P J; Hering, B J; Barton, F B

2014-11-01

The Collaborative Islet Transplant Registry (CITR) collects data on clinical islet isolations and transplants. This retrospective report analyzed 1017 islet isolation procedures performed for 537 recipients of allogeneic clinical islet transplantation in 1999-2010. This study describes changes in donor and islet isolation variables by era and factors associated with quantity and quality of final islet products. Donor body weight and BMI increased significantly over the period (pIslet yield measures have improved with time including islet equivalent (IEQ)/particle ratio and IEQs infused. The average dose of islets infused significantly increased in the era of 2007-2010 when compared to 1999-2002 (445.4±156.8 vs. 421.3±155.4×0(3) IEQ; pIslet purity and total number of β cells significantly improved over the study period (pislets has remained consistently very high through this period, and differs substantially from nonclinical islets. In multivariate analysis of all recipient, donor and islet factors, and medical management factors, the only islet product characteristic that correlated with clinical outcomes was total IEQs infused. This analysis shows improvements in both quantity and some quality criteria of clinical islets produced over 1999-2010, and these parallel improvements in clinical outcomes over the same period. © 2014 The Authors. American Journal of Transplantation Published by Wiley Periodicals, Inc. on behalf of American Society of Transplant Surgeons.

Loss of end-differentiated β-cell phenotype following pancreatic islet transplantation.

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Anderson, S J; White, M G; Armour, S L; Maheshwari, R; Tiniakos, D; Muller, Y D; Berishvili, E; Berney, T; Shaw, J A M

2018-03-01

Replacement of pancreatic β-cells through deceased donor islet transplantation is a proven therapy for preventing recurrent life-threatening hypoglycemia in type 1 diabetes. Although near-normal glucose levels and insulin independence can be maintained for many years following successful islet transplantation, restoration of normal functional β-cell mass has remained elusive. It has recently been proposed that dedifferentiation/plasticity towards other endocrine phenotypes may play an important role in stress-induced β-cell dysfunction in type 2 diabetes. Here we report loss of end-differentiated β-cell phenotype in 2 intraportal islet allotransplant recipients. Despite excellent graft function and sustained insulin independence, all examined insulin-positive cells had lost expression of the end-differentiation marker, urocortin-3, or appeared to co-express the α-cell marker, glucagon. In contrast, no insulin + /urocortin-3 - cells were seen in nondiabetic deceased donor control pancreatic islets. Loss of end-differentiated phenotype may facilitate β-cell survival during the stresses associated with islet isolation and culture, in addition to sustained hypoxia following engraftment. As further refinements in islet isolation and culture are made in parallel with exploration of alternative β-cell sources, graft sites, and ultimately fully vascularized bioengineered insulin-secreting microtissues, differentiation status immunostaining provides a novel tool to assess whether fully mature β-cell phenotype has been maintained. © 2017 The American Society of Transplantation and the American Society of Transplant Surgeons.

Islet Product Characteristics and Factors Related to Successful Human Islet Transplantation From the Collaborative Islet Transplant Registry (CITR) 1999–2010

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Balamurugan, A N; Naziruddin, B; Lockridge, A; Tiwari, M; Loganathan, G; Takita, M; Matsumoto, S; Papas, K; Trieger, M; Rainis, H; Kin, T; Kay, T W; Wease, S; Messinger, S; Ricordi, C; Alejandro, R; Markmann, J; Kerr-Conti, J; Rickels, M R; Liu, C; Zhang, X; Witkowski, P; Posselt, A; Maffi, P; Secchi, A; Berney, T; O’Connell, P J; Hering, B J; Barton, F B

2014-01-01

The Collaborative Islet Transplant Registry (CITR) collects data on clinical islet isolations and transplants. This retrospective report analyzed 1017 islet isolation procedures performed for 537 recipients of allogeneic clinical islet transplantation in 1999–2010. This study describes changes in donor and islet isolation variables by era and factors associated with quantity and quality of final islet products. Donor body weight and BMI increased significantly over the period (p Islet yield measures have improved with time including islet equivalent (IEQ)/particle ratio and IEQs infused. The average dose of islets infused significantly increased in the era of 2007–2010 when compared to 1999–2002 (445.4 ± 156.8 vs. 421.3 ± 155.4 ×103 IEQ; p Islet purity and total number of β cells significantly improved over the study period (p islets has remained consistently very high through this period, and differs substantially from nonclinical islets. In multivariate analysis of all recipient, donor and islet factors, and medical management factors, the only islet product characteristic that correlated with clinical outcomes was total IEQs infused. This analysis shows improvements in both quantity and some quality criteria of clinical islets produced over 1999–2010, and these parallel improvements in clinical outcomes over the same period. PMID:25278159

Gene expression in rat models for inter-generational transmission of islet dysfunction and obesity

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Ruby C.Y. Lin

2014-12-01

Full Text Available Paternal high fat diet (HFD consumption triggers unique gene signatures, consistent with premature aging and chronic degenerative disorders, in both white adipose tissue (RpWAT and pancreatic islets of daughters. In addition to published data in Nature, 2010, 467, 963–966 (GSE: 19877, islet and FASEB J 2014, 28, 1830–1841 (GSE: 33551, RpWAT, we describe here additional details on systems-based approaches and analysis to develop our observations. Our data provides a resource for exploring the complex molecular mechanisms that underlie intergenerational transmission of obesity.

Hippocampal insulin resistance and cognitive dysfunction

NARCIS (Netherlands)

Biessels, Geert Jan; Reagan, Lawrence P.

2015-01-01

Clinical studies suggest a link between type 2 diabetes mellitus (T2DM) and insulin resistance (IR) and cognitive dysfunction, but there are significant gaps in our knowledge of the mechanisms underlying this relationship. Animal models of IR help to bridge these gaps and point to hippocampal IR as

Feasibility of islet magnetic resonance imaging using ferumoxytol in intraportal islet transplantation.

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Jin, Sang-Man; Oh, Seung-Hoon; Oh, Bae Jun; Shim, Wooyoung; Choi, Jin Myung; Yoo, Dongkyeom; Hwang, Yong Hwa; Lee, Jung Hee; Lee, Dong Yun; Kim, Jae Hyeon

2015-06-01

There is a clinical need for an alternative labeling agent for magnetic resonance imaging (MRI) in islet transplantation. We aimed to evaluate the feasibility of islet MRI using ferumoxytol, which is the only clinically-available ultrasmall superparamagnetic iron oxide. We compared islet function and viability of control islets and islets labeled with ferumoxytol and/or a heparin-protamine complex (HPF). Efficacy of ferumoxytol labeling was assessed in both ex vivo and in vivo models. Labeling for 48 h with HPF, but not up to 800 μg/mL ferumoxytol, deranged ex vivo islet viability and function. The T2∗ relaxation time was optimal when islets were labeled with 800 μg/mL of ferumoxytol for 48 h. Prussian blue stain, iron content assay, transmission electron microscopy (TEM) supported internalization of ferumoxytol particles. However, the labeling intensity in the ex vivo MRI of islets labeled with ferumoxytol was much weaker than that of islets labeled with ferucarbotran. In syngeneic intraportal islet transplantation, there was a correlation between the total area of visualized islets and the transplanted islet mass. In conclusion, islet MRI using ferumoxytol was feasible in terms of in vitro and in vivo efficacy and safety. However, the weak labeling efficacy is still a hurdle for the clinical application. Copyright © 2015 Elsevier Ltd. All rights reserved.

Pancreatic islet regeneration: Therapeutic potential, unknowns and controversy

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Ingrid L. Cockburn

2015-07-01

Full Text Available Glucose homeostasis in mammals is primarily maintained by the insulin-secreting β-cells contained within pancreas-resident islets of Langerhans. Gross disruption of this glucose regulation as a result of pancreatic dysfunction frequently results in diabetes, which is currently a major health concern in South Africa, as well as globally. For many years, researchers have realised that the pancreas, and specifically the islets of Langerhans, have a regenerative capacity, as islet mass has frequently been shown to increase following induced pancreatic injury. Given that gross β-cell loss contributes significantly to the pathogenesis of both type 1 and type 2 diabetes, endogenous pancreatic islet regeneration has been investigated extensively as a potential β-cell replacement therapy for diabetes. From the extensive research conducted on pancreatic regeneration, opposing findings and opinions have arisen as to how, and more recently even if, pancreatic regeneration occurs following induced injury. In this review, we outline and discuss the three primary mechanisms by which pancreatic regeneration is proposed to occur: neogenesis, β-cell replication and transdifferentiation. We further explain some of the advanced techniques used in pancreatic regeneration research, and conclude that despite the technologically advanced research tools available to researchers today, the mechanisms governing pancreatic regeneration may remain elusive until more powerful techniques are developed to allow for real-time, live-cell assessment of morphology and gene expression within the pancreas.

Rotational Transport of Islets: The Best Way for Islets to Get around?

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Rupert Oberhuber

2013-01-01

Full Text Available Islet transplantation is a valid treatment option for patients suffering from type 1 diabetes mellitus. To assure optimal islet cell quality, specialized islet isolation facilities have been developed. Utilization of such facilities necessitates transportation of islet cells to distant institutions for transplantation. Despite its importance, a clinically feasible solution for the transport of islets has still not been established. We here compare the functionality of isolated islets from C57BL/6 mice directly after the isolation procedure as well as after two simulated transport conditions, static versus rotation. Islet cell quality was assessed using real-time live confocal microscopy. In vivo islet function after syngeneic transplantation was determined by weight and blood sugar measurements as well as by intraperitoneal glucose tolerance tests. Vascularization of islets was documented by fluorescence microscopy and immunohistochemistry. All viability parameters documented comparable cell viability in the rotary group and the group transplanted immediately after isolation. Functional parameters assessed in vivo displayed no significant difference between these two groups. Moreover, vascularization of islets was similar in both groups. In conclusion, rotary culture conditions allows the maintenance of highest islet quality for at least 15 h, which is comparable to that of freshly isolated islets.

Isolated human islets require hyperoxia to maintain islet mass, metabolism, and function.

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Komatsu, Hirotake; Kang, Dongyang; Medrano, Leonard; Barriga, Alyssa; Mendez, Daniel; Rawson, Jeffrey; Omori, Keiko; Ferreri, Kevin; Tai, Yu-Chong; Kandeel, Fouad; Mullen, Yoko

2016-02-12

Pancreatic islet transplantation has been recognized as an effective treatment for Type 1 diabetes; however, there is still plenty of room to improve transplantation efficiency. Because islets are metabolically active they require high oxygen to survive; thus hypoxia after transplant is one of the major causes of graft failure. Knowing the optimal oxygen tension for isolated islets would allow a transplant team to provide the best oxygen environment during pre- and post-transplant periods. To address this issue and begin to establish empirically determined guidelines for islet maintenance, we exposed in vitro cultured islets to different partial oxygen pressures (pO2) and assessed changes in islet volume, viability, metabolism, and function. Human islets were cultured for 7 days in different pO2 media corresponding to hypoxia (90 mmHg), normoxia (160 mmHg), and hyerpoxia (270 or 350 mmHg). Compared to normoxia and hypoxia, hyperoxia alleviated the loss of islet volume, maintaining higher islet viability and metabolism as measured by oxygen consumption and glucose-stimulated insulin secretion responses. We predict that maintaining pre- and post-transplanted islets in a hyperoxic environment will alleviate islet volume loss and maintain islet quality thereby improving transplant outcomes. Copyright © 2016 Elsevier Inc. All rights reserved.

Impact of islet size on pancreatic islet transplantation and potential interventions to improve outcome.

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Zorzi, Daria; Phan, Tammy; Sequi, Marco; Lin, Yong; Freeman, Daniel H; Cicalese, Luca; Rastellini, Cristiana

2015-01-01

Better results have been recently reported in clinical pancreatic islet transplantation (ITX) due mostly to improved isolation techniques and immunosuppression; however, some limitations still exist. It is known that following transplantation, 30% to 60% of the islets are lost. In our study, we have investigated 1) the role of size as a factor affecting islet engraftment and 2) potential procedural manipulations to increase the number of smaller functional islets that can be transplanted. C57/BL10 mice were used as donors and recipients in a syngeneic islet transplant model. Isolated islets were divided by size (large, >300 μm; medium 150-300 μm; small, <150 μm). Each size was transplanted in chemically induced diabetic mice as full (600 IEQ), suboptimal (400 IEQ), and marginal mass (200 IEQ). Control animals received all size islets. Engraftment was defined as reversal of diabetes by day 7 posttransplantation. When the superiority of smaller islets was observed, strategies of overdigestion and fragmentation were adopted during islet isolation in the attempt to reduce islet size and improve engraftment. Smaller islets were significantly superior in engraftment compared to medium, large, and control (all sizes) groups. This was more evident when marginal mass data were compared. In all masses, success decreased as islet size increased. Once islets were engrafted, functionality was not affected by size. When larger islets were fragmented, a significant decrease in islet functionality was observed. On the contrary, if pancreata were slightly overdigested, although not as successful as small naive islets, an increase in engraftment was observed when compared to the control group. In conclusion, smaller islets are superior in engraftment following islet transplantation. Fragmentation has a deleterious effect on islet engraftment. Islet isolations can be performed by reducing islet size with slight overdigestion, and it can be safely adopted to improve clinical

Glucotoxicity promotes aberrant activation and mislocalization of Ras-related C3 botulinum toxin substrate 1 [Rac1] and metabolic dysfunction in pancreatic islet β-cells: reversal of such metabolic defects by metformin.

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Baidwan, Sartaj; Chekuri, Anil; Hynds, DiAnna L; Kowluru, Anjaneyulu

2017-11-01

Emerging evidence suggests that long-term exposure of insulin-secreting pancreatic β-cells to hyperglycemic (HG; glucotoxic) conditions promotes oxidative stress, which, in turn, leads to stress kinase activation, mitochondrial dysfunction, loss of nuclear structure and integrity and cell apoptosis. Original observations from our laboratory have proposed that Rac1 plays a key regulatory role in the generation of oxidative stress and downstream signaling events culminating in the onset of dysfunction of pancreatic β-cells under the duress of metabolic stress. However, precise molecular and cellular mechanisms underlying the metabolic roles of hyperactive Rac1 remain less understood. Using pharmacological and molecular biological approaches, we now report mistargetting of biologically-active Rac1 [GTP-bound conformation] to the nuclear compartment in clonal INS-1 cells, normal rat islets and human islets under HG conditions. Our findings also suggest that such a signaling step is independent of post-translational prenylation of Rac1. Evidence is also presented to highlight novel roles for sustained activation of Rac1 in HG-induced expression of Cluster of Differentiation 36 [CD36], a fatty acid transporter protein, which is implicated in cell apoptosis. Finally, our findings suggest that metformin, a biguanide anti-diabetic drug, at a clinically relevant concentration, prevents β-cell defects [Rac1 activation, nuclear association, CD36 expression, stress kinase and caspase-3 activation, and loss in metabolic viability] under the duress of glucotoxicity. Potential implications of these findings in the context of novel and direct regulation of islet β-cell function by metformin are discussed.

A Syntenic Cross Species Aneuploidy Genetic Screen Links RCAN1 Expression to β-Cell Mitochondrial Dysfunction in Type 2 Diabetes.

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Heshan Peiris

2016-05-01

Full Text Available Type 2 diabetes (T2D is a complex metabolic disease associated with obesity, insulin resistance and hypoinsulinemia due to pancreatic β-cell dysfunction. Reduced mitochondrial function is thought to be central to β-cell dysfunction. Mitochondrial dysfunction and reduced insulin secretion are also observed in β-cells of humans with the most common human genetic disorder, Down syndrome (DS, Trisomy 21. To identify regions of chromosome 21 that may be associated with perturbed glucose homeostasis we profiled the glycaemic status of different DS mouse models. The Ts65Dn and Dp16 DS mouse lines were hyperglycemic, while Tc1 and Ts1Rhr mice were not, providing us with a region of chromosome 21 containing genes that cause hyperglycemia. We then examined whether any of these genes were upregulated in a set of ~5,000 gene expression changes we had identified in a large gene expression analysis of human T2D β-cells. This approach produced a single gene, RCAN1, as a candidate gene linking hyperglycemia and functional changes in T2D β-cells. Further investigations demonstrated that RCAN1 methylation is reduced in human T2D islets at multiple sites, correlating with increased expression. RCAN1 protein expression was also increased in db/db mouse islets and in human and mouse islets exposed to high glucose. Mice overexpressing RCAN1 had reduced in vivo glucose-stimulated insulin secretion and their β-cells displayed mitochondrial dysfunction including hyperpolarised membrane potential, reduced oxidative phosphorylation and low ATP production. This lack of β-cell ATP had functional consequences by negatively affecting both glucose-stimulated membrane depolarisation and ATP-dependent insulin granule exocytosis. Thus, from amongst the myriad of gene expression changes occurring in T2D β-cells where we had little knowledge of which changes cause β-cell dysfunction, we applied a trisomy 21 screening approach which linked RCAN1 to β-cell mitochondrial

Continuous Quadrupole Magnetic Separation of Islets during Digestion Improves Purified Porcine Islet Viability.

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Weegman, Bradley P; Kumar Sajja, Venkata Sunil; Suszynski, Thomas M; Rizzari, Michael D; Scott Iii, William E; Kitzmann, Jennifer P; Mueller, Kate R; Hanley, Thomas R; Kennedy, David J; Todd, Paul W; Balamurugan, Appakalai N; Hering, Bernhard J; Papas, Klearchos K

2016-01-01

Islet transplantation (ITx) is an emerging and promising therapy for patients with uncontrolled type 1 diabetes. The islet isolation and purification processes require exposure to extended cold ischemia, warm-enzymatic digestion, mechanical agitation, and use of damaging chemicals for density gradient separation (DG), all of which reduce viable islet yield. In this paper, we describe initial proof-of-concept studies exploring quadrupole magnetic separation (QMS) of islets as an alternative to DG to reduce exposure to these harsh conditions. Three porcine pancreata were split into two parts, the splenic lobe (SPL) and the combined connecting/duodenal lobes (CDL), for paired digestions and purifications. Islets in the SPL were preferentially labeled using magnetic microparticles (MMPs) that lodge within the islet microvasculature when infused into the pancreas and were continuously separated from the exocrine tissue by QMS during the collection phase of the digestion process. Unlabeled islets from the CDL were purified by conventional DG. Islets purified by QMS exhibited significantly improved viability (measured by oxygen consumption rate per DNA, p < 0.03) and better morphology relative to control islets. Islet purification by QMS can reduce the detrimental effects of prolonged exposure to toxic enzymes and density gradient solutions and substantially improve islet viability after isolation.

Continuous Quadrupole Magnetic Separation of Islets during Digestion Improves Purified Porcine Islet Viability

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Bradley P. Weegman

2016-01-01

Full Text Available Islet transplantation (ITx is an emerging and promising therapy for patients with uncontrolled type 1 diabetes. The islet isolation and purification processes require exposure to extended cold ischemia, warm-enzymatic digestion, mechanical agitation, and use of damaging chemicals for density gradient separation (DG, all of which reduce viable islet yield. In this paper, we describe initial proof-of-concept studies exploring quadrupole magnetic separation (QMS of islets as an alternative to DG to reduce exposure to these harsh conditions. Three porcine pancreata were split into two parts, the splenic lobe (SPL and the combined connecting/duodenal lobes (CDL, for paired digestions and purifications. Islets in the SPL were preferentially labeled using magnetic microparticles (MMPs that lodge within the islet microvasculature when infused into the pancreas and were continuously separated from the exocrine tissue by QMS during the collection phase of the digestion process. Unlabeled islets from the CDL were purified by conventional DG. Islets purified by QMS exhibited significantly improved viability (measured by oxygen consumption rate per DNA, p<0.03 and better morphology relative to control islets. Islet purification by QMS can reduce the detrimental effects of prolonged exposure to toxic enzymes and density gradient solutions and substantially improve islet viability after isolation.

Roles of mitochondrial fragmentation and reactive oxygen species in mitochondrial dysfunction and myocardial insulin resistance

International Nuclear Information System (INIS)

Watanabe, Tomoyuki; Saotome, Masao; Nobuhara, Mamoru; Sakamoto, Atsushi; Urushida, Tsuyoshi; Katoh, Hideki; Satoh, Hiroshi; Funaki, Makoto; Hayashi, Hideharu

2014-01-01

Purpose: Evidence suggests an association between aberrant mitochondrial dynamics and cardiac diseases. Because myocardial metabolic deficiency caused by insulin resistance plays a crucial role in heart disease, we investigated the role of dynamin-related protein-1 (DRP1; a mitochondrial fission protein) in the pathogenesis of myocardial insulin resistance. Methods and Results: DRP1-expressing H9c2 myocytes, which had fragmented mitochondria with mitochondrial membrane potential (ΔΨ m ) depolarization, exhibited attenuated insulin signaling and 2-deoxy-D-glucose (2-DG) uptake, indicating insulin resistance. Treatment of the DRP1-expressing myocytes with Mn(III)tetrakis(1-methyl-4-pyridyl)porphyrin pentachloride (TMPyP) significantly improved insulin resistance and mitochondrial dysfunction. When myocytes were exposed to hydrogen peroxide (H 2 O 2 ), they increased DRP1 expression and mitochondrial fragmentation, resulting in ΔΨ m depolarization and insulin resistance. When DRP1 was suppressed by siRNA, H 2 O 2 -induced mitochondrial dysfunction and insulin resistance were restored. Our results suggest that a mutual enhancement between DRP1 and reactive oxygen species could induce mitochondrial dysfunction and myocardial insulin resistance. In palmitate-induced insulin-resistant myocytes, neither DRP1-suppression nor TMPyP restored the ΔΨ m depolarization and impaired 2-DG uptake, however they improved insulin signaling. Conclusions: A mutual enhancement between DRP1 and ROS could promote mitochondrial dysfunction and inhibition of insulin signal transduction. However, other mechanisms, including lipid metabolite-induced mitochondrial dysfunction, may be involved in palmitate-induced insulin resistance. - Highlights: • DRP1 promotes mitochondrial fragmentation and insulin-resistance. • A mutual enhancement between DRP1 and ROS ipromotes insulin-resistance. • Palmitate increases DRP1 expression and induces insulin-resistance. • Inhibition of DRP or ROS

Roles of mitochondrial fragmentation and reactive oxygen species in mitochondrial dysfunction and myocardial insulin resistance

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Watanabe, Tomoyuki [Internal Medicine III, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu 431-3192 (Japan); Saotome, Masao, E-mail: msaotome@hama-med.ac.jp [Internal Medicine III, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu 431-3192 (Japan); Nobuhara, Mamoru; Sakamoto, Atsushi; Urushida, Tsuyoshi; Katoh, Hideki; Satoh, Hiroshi [Internal Medicine III, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu 431-3192 (Japan); Funaki, Makoto [Clinical Research Center for Diabetes, Tokushima University Hospital, 2-50-1 Kuramoto-cho, Tokushima 770-8503 (Japan); Hayashi, Hideharu [Internal Medicine III, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu 431-3192 (Japan)

2014-05-01

Purpose: Evidence suggests an association between aberrant mitochondrial dynamics and cardiac diseases. Because myocardial metabolic deficiency caused by insulin resistance plays a crucial role in heart disease, we investigated the role of dynamin-related protein-1 (DRP1; a mitochondrial fission protein) in the pathogenesis of myocardial insulin resistance. Methods and Results: DRP1-expressing H9c2 myocytes, which had fragmented mitochondria with mitochondrial membrane potential (ΔΨ{sub m}) depolarization, exhibited attenuated insulin signaling and 2-deoxy-D-glucose (2-DG) uptake, indicating insulin resistance. Treatment of the DRP1-expressing myocytes with Mn(III)tetrakis(1-methyl-4-pyridyl)porphyrin pentachloride (TMPyP) significantly improved insulin resistance and mitochondrial dysfunction. When myocytes were exposed to hydrogen peroxide (H{sub 2}O{sub 2}), they increased DRP1 expression and mitochondrial fragmentation, resulting in ΔΨ{sub m} depolarization and insulin resistance. When DRP1 was suppressed by siRNA, H{sub 2}O{sub 2}-induced mitochondrial dysfunction and insulin resistance were restored. Our results suggest that a mutual enhancement between DRP1 and reactive oxygen species could induce mitochondrial dysfunction and myocardial insulin resistance. In palmitate-induced insulin-resistant myocytes, neither DRP1-suppression nor TMPyP restored the ΔΨ{sub m} depolarization and impaired 2-DG uptake, however they improved insulin signaling. Conclusions: A mutual enhancement between DRP1 and ROS could promote mitochondrial dysfunction and inhibition of insulin signal transduction. However, other mechanisms, including lipid metabolite-induced mitochondrial dysfunction, may be involved in palmitate-induced insulin resistance. - Highlights: • DRP1 promotes mitochondrial fragmentation and insulin-resistance. • A mutual enhancement between DRP1 and ROS ipromotes insulin-resistance. • Palmitate increases DRP1 expression and induces insulin-resistance

Inflammatory Response in Islet Transplantation

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Mazhar A. Kanak

2014-01-01

Full Text Available Islet cell transplantation is a promising beta cell replacement therapy for patients with brittle type 1 diabetes as well as refractory chronic pancreatitis. Despite the vast advancements made in this field, challenges still remain in achieving high frequency and long-term successful transplant outcomes. Here we review recent advances in understanding the role of inflammation in islet transplantation and development of strategies to prevent damage to islets from inflammation. The inflammatory response associated with islets has been recognized as the primary cause of early damage to islets and graft loss after transplantation. Details on cell signaling pathways in islets triggered by cytokines and harmful inflammatory events during pancreas procurement, pancreas preservation, islet isolation, and islet infusion are presented. Robust control of pre- and peritransplant islet inflammation could improve posttransplant islet survival and in turn enhance the benefits of islet cell transplantation for patients who are insulin dependent. We discuss several potent anti-inflammatory strategies that show promise for improving islet engraftment. Further understanding of molecular mechanisms involved in the inflammatory response will provide the basis for developing potent therapeutic strategies for enhancing the quality and success of islet transplantation.

Inflammatory Response in Islet Transplantation

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Kanak, Mazhar A.; Kunnathodi, Faisal; Lawrence, Michael C.; Levy, Marlon F.

2014-01-01

Islet cell transplantation is a promising beta cell replacement therapy for patients with brittle type 1 diabetes as well as refractory chronic pancreatitis. Despite the vast advancements made in this field, challenges still remain in achieving high frequency and long-term successful transplant outcomes. Here we review recent advances in understanding the role of inflammation in islet transplantation and development of strategies to prevent damage to islets from inflammation. The inflammatory response associated with islets has been recognized as the primary cause of early damage to islets and graft loss after transplantation. Details on cell signaling pathways in islets triggered by cytokines and harmful inflammatory events during pancreas procurement, pancreas preservation, islet isolation, and islet infusion are presented. Robust control of pre- and peritransplant islet inflammation could improve posttransplant islet survival and in turn enhance the benefits of islet cell transplantation for patients who are insulin dependent. We discuss several potent anti-inflammatory strategies that show promise for improving islet engraftment. Further understanding of molecular mechanisms involved in the inflammatory response will provide the basis for developing potent therapeutic strategies for enhancing the quality and success of islet transplantation. PMID:24883060

Beneficial effect of D-allose for isolated islet culture prior to islet transplantation.

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Kashiwagi, Hirotaka; Asano, Eisuke; Noguchi, Chisato; Sui, Li; Hossain, Akram; Akamoto, Shintaro; Okano, Keiichi; Tokuda, Masaaki; Suzuki, Yasuyuki

2016-01-01

Pretransplant restoration of islets damaged during isolation remains to be solved. In this study, we examined the effect of D-allose on islets isolated from rat pancreata prior to islet transplantation. Rat islets isolated from fresh pancreata were cultured overnight in Roswell Park Memorial Institute 1640 solution in the absence (group 1) or presence (group 2) of D-allose. Then we assessed stimulation index of insulin, and cure rate after islet transplantation to diabetic nude mice. We also measured malondialdehyde level and caspase 3 activity of islets after the overnight culture for assessment of the oxidative stress and the apoptosis. D-allose significantly improved insulin secretion of islets. The stimulation index in group 2 was significantly higher than in group 1. Cure rate after transplantation in group 2 was higher than in group 1 especially in the first week. The malondialdehyde level in group 2 was significantly lower than in group 1. But the caspase 3 activities in both groups did not differ. D-allose treatment of isolated islet culture prior to transplantation restored islet function and increased successful transplant rate. The results of this study suggested that D-allose improved function of damaged islets through its anti-oxidative activity. © 2015 Japanese Society of Hepato-Biliary-Pancreatic Surgery.

Small Islets Transplantation Superiority to Large Ones: Implications from Islet Microcirculation and Revascularization

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Wenjuan Li

2014-01-01

Full Text Available Pancreatic islet transplantation is a promising therapy to regain glycemic control in diabetic patients. The selection of ideal grafts is the basis to guarantee short-term effectivity and longevity of the transplanted islets. Contradictory to the traditional notion, recent findings implied the superiority of small islets for better transplantation outcomes rather than the large and intact ones. However, the mechanisms remain to be elucidated. Recent evidences emphasized the major impact of microcirculation on islet β-cell mass and function. And potentials in islet graft revascularization are crucial for their survival and preserved function in the recipient. In this study, we verified the distinct histological phenotype and functionality of small islets versus large ones both in vitro and in vivo. With efforts to exploring the differences in microcirculation and revascularization of islet grafts, we further evaluated local expressions of angiotensin and vascular endothelial growth factor A (VEGF-A at different levels. Our findings reveal that, apart from the higher density of insulin-producing β-cells, small islets express less angiotensin and more angiotrophic VEGF-A. We therefore hypothesized a logical explanation of the small islet superiority for transplantation outcome from the aspects of facilitated microcirculation and revascularization intrinsically in small islets.

Facilitated Engraftment of Isolated Islets Coated With Expanded Vascular Endothelial Cells for Islet Transplantation.

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Barba-Gutierrez, D Alonso; Daneri-Navarro, A; Villagomez-Mendez, J Jesus Alejandro; Kanamune, J; Robles-Murillo, A Karina; Sanchez-Enriquez, S; Villafan-Bernal, J Rafael; Rivas-Carrillo, J D

2016-03-01

Diabetes is complex disease, which involves primary metabolic changes followed by immunological and vascular pathophysiological adjustments. However, it is mostly characterized by an unbalanced decreased number of the β-cells unable to maintain the metabolic requirements and failure to further regenerate newly functional pancreatic islets. The objective of this study was to analyze the properties of the endothelial cells to facilitate the islet cells engraftment after islet transplantation. We devised a co-cultured engineer system to coat isolated islets with vascular endothelial cells. To assess the cell integration of cell-engineered islets, we stained them for endothelial marker CD31 and nuclei counterstained with DAPI dye. We comparatively performed islet transplantations into streptozotocin-induced diabetic mice and recovered the islet grafts for morphometric analyses on days 3, 7, 10, and 30. Blood glucose levels were measured continuously after islet transplantation to monitor the functional engraftment and capacity to achieve metabolic control. Cell-engineered islets showed a well-defined rounded shape after co-culture when compared with native isolated islets. Furthermore, the number of CD31-positive cells layered on the islet surface showed a direct proportion with engraftment capacities and less TUNEL-positive cells on days 3 and 7 after transplantation. We observed that vascular endothelial cells could be functional integrated into isolated islets. We also found that islets that are coated with vascular endothelial cells increased their capacity to engraft. These findings indicate that islets coated with endothelial cells have a greater capacity of engraftment and thus establish a definitely vascular network to support the metabolic requirements. Copyright © 2016 Elsevier Inc. All rights reserved.

Insulin Resistance and Endothelial Dysfunction Constitute a Common Therapeutic Target in Cardiometabolic Disorders

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A. Janus

2016-01-01

Full Text Available Insulin resistance and other risk factors for atherosclerosis, such as hypertension and hypercholesterolemia, promote endothelial dysfunction and lead to development of metabolic syndrome which constitutes an introduction to cardiovascular disease. The insulin resistance and endothelial dysfunction cross talk between each other by numerous metabolic pathways. Hence, targeting one of these pathologies with pleiotropic treatment exerts beneficial effect on another one. Combined and expletive treatment of hypertension, lipid disorders, and insulin resistance with nonpharmacological interventions and conventional pharmacotherapy may inhibit the transformation of metabolic disturbances to fully developed cardiovascular disease. This paper summarises the common therapeutic targets for insulin resistance, endothelial dysfunction, and vascular inflammatory reaction at molecular level and analyses the potential pleiotropic effects of drugs used currently in management of cardiovascular disease, metabolic syndrome, and diabetes.

Pancreatic islet transplantation

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Corrêa-Giannella Maria

2009-09-01

Full Text Available Abstract Background No formulation of exogenous insulin available to date has yet been able to mimic the physiological nictemeral rhythms of this hormone, and despite all engineering advancements, the theoretical proposal of developing a mechanical replacement for pancreatic β cell still has not been reached. Thus, the replacement of β cells through pancreas and pancreatic islet transplantation are the only concrete alternatives for re-establishing the endogenous insulin secretion in type 1 diabetic patients. Since only 1 to 1.5% of the pancreatic mass corresponds to endocrine tissue, pancreatic islets transplantation arises as a natural alternative. Data from the International Islet Transplant Registry (ITR from 1983 to December 2000 document a total of 493 transplants performed around the world, with progressively worse rates of post-transplant insulin independence. In 2000, the "Edmonton Protocol" introduced several modifications to the transplantation procedure, such as the use of a steroid-free immunosuppression regimen and transplantation of a mean islet mass of 11,000 islet equivalents per kilogram, which significantly improved 1-year outcomes. Although the results of a 5-year follow-up in 65 patients demonstrated improvement in glycemic instability in a significant portion of them, only 7.5% of the patients have reached insulin independence, indicating the need of further advances in the preservation of the function of transplanted islet. In addition to the scarcity of organs available for transplantation, islets transplantation still faces major challenges, specially those related to cell loss during the process of islet isolation and the losses related to the graft site, apoptosis, allorejection, autoimmunity, and immunosuppression. The main strategies to optimize islet transplantation aim at improving all these aspects. Conclusion Human islet transplantation should be regarded as an intervention that can decrease the frequency of

Combined lipidomic and proteomic analysis of isolated human islets exposed to palmitate reveals time-dependent changes in insulin secretion and lipid metabolism.

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Kirsten Roomp

Full Text Available Studies on the pathophysiology of type 2 diabetes mellitus (T2DM have linked the accumulation of lipid metabolites to the development of beta-cell dysfunction and impaired insulin secretion. In most in vitro models of T2DM, rodent islets or beta-cell lines are used and typically focus is on specific cellular pathways or organs. Our aim was to, firstly, develop a combined lipidomics and proteomics approach for lipotoxicity in isolated human islets and, secondly, investigate if the approach could delineate novel and/ or confirm reported mechanisms of lipotoxicity. To this end isolated human pancreatic islets, exposed to chronically elevated palmitate concentrations for 0, 2 and 7 days, were functionally characterized and their levels of multiple targeted lipid and untargeted protein species determined. Glucose-stimulated insulin secretion from the islets increased on day 2 and decreased on day 7. At day 7 islet insulin content decreased and the proinsulin to insulin content ratio doubled. Amounts of cholesterol, stearic acid, C16 dihydroceramide and C24:1 sphingomyelin, obtained from the lipidomic screen, increased time-dependently in the palmitate-exposed islets. The proteomic screen identified matching changes in proteins involved in lipid biosynthesis indicating up-regulated cholesterol and lipid biosynthesis in the islets. Furthermore, proteins associated with immature secretory granules were decreased when palmitate exposure time was increased despite their high affinity for cholesterol. Proteins associated with mature secretory granules remained unchanged. Pathway analysis based on the protein and lipid expression profiles implicated autocrine effects of insulin in lipotoxicity. Taken together the study demonstrates that combining different omics approaches has potential in mapping of multiple simultaneous cellular events. However, it also shows that challenges exist for effectively combining lipidomics and proteomics in primary cells. Our

High glucose suppresses human islet insulin biosynthesis by inducing miR-133a leading to decreased polypyrimidine tract binding protein-expression.

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Rikard G Fred

Full Text Available BACKGROUND: Prolonged periods of high glucose exposure results in human islet dysfunction in vitro. The underlying mechanisms behind this effect of high glucose are, however, unknown. The polypyrimidine tract binding protein (PTB is required for stabilization of insulin mRNA and the PTB mRNA 3'-UTR contains binding sites for the microRNA molecules miR-133a, miR-124a and miR-146. The aim of this study was therefore to investigate whether high glucose increased the levels of these three miRNAs in association with lower PTB levels and lower insulin biosynthesis rates. METHODOLOGY/PRINCIPAL FINDINGS: Human islets were cultured for 24 hours in the presence of low (5.6 mM or high glucose (20 mM. Islets were also exposed to sodium palmitate or the proinflammatory cytokines IL-1beta and IFN-gamma, since saturated free fatty acids and cytokines also cause islet dysfunction. RNA was then isolated for real-time RT-PCR analysis of miR-133a, miR-124a, miR-146, insulin mRNA and PTB mRNA contents. Insulin biosynthesis rates were determined by radioactive labeling and immunoprecipitation. Synthetic miR-133a precursor and inhibitor were delivered to dispersed islet cells by lipofection, and PTB was analyzed by immunoblotting following culture at low or high glucose. Culture in high glucose resulted in increased islet contents of miR-133a and reduced contents of miR-146. Cytokines increased the contents of miR-146. The insulin and PTB mRNA contents were unaffected by high glucose. However, both PTB protein levels and insulin biosynthesis rates were decreased in response to high glucose. The miR-133a inhibitor prevented the high glucose-induced decrease in PTB and insulin biosynthesis, and the miR-133a precursor decreased PTB levels and insulin biosynthesis similarly to high glucose. CONCLUSION: Prolonged high-glucose exposure down-regulates PTB levels and insulin biosynthesis rates in human islets by increasing miR-133a levels. We propose that this mechanism

Endothelial chimerism and vascular sequestration protect pancreatic islet grafts from antibody-mediated rejection

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Chen, Chien-Chia; Pouliquen, Eric; Broisat, Alexis; Andreata, Francesco; Racapé, Maud; Bruneval, Patrick; Kessler, Laurence; Ahmadi, Mitra; Bacot, Sandrine; Saison-Delaplace, Carole; Marcaud, Marina; Van Huyen, Jean-Paul Duong; Loupy, Alexandre; Villard, Jean; Demuylder-Mischler, Sandrine; Morelon, Emmanuel; Tsai, Meng-Kun; Kolopp-Sarda, Marie-Nathalie; Koenig, Alice; Mathias, Virginie; Ghezzi, Catherine; Dubois, Valerie; Defrance, Thierry

2017-01-01

Humoral rejection is the most common cause of solid organ transplant failure. Here, we evaluated a cohort of 49 patients who were successfully grafted with allogenic islets and determined that the appearance of donor-specific anti-HLA antibodies (DSAs) did not accelerate the rate of islet graft attrition, suggesting resistance to humoral rejection. Murine DSAs bound to allogeneic targets expressed by islet cells and induced their destruction in vitro; however, passive transfer of the same DSAs did not affect islet graft survival in murine models. Live imaging revealed that DSAs were sequestrated in the circulation of the recipients and failed to reach the endocrine cells of grafted islets. We used murine heart transplantation models to confirm that endothelial cells were the only accessible targets for DSAs, which induced the development of typical microvascular lesions in allogeneic transplants. In contrast, the vasculature of DSA-exposed allogeneic islet grafts was devoid of lesions because sprouting of recipient capillaries reestablished blood flow in grafted islets. Thus, we conclude that endothelial chimerism combined with vascular sequestration of DSAs protects islet grafts from humoral rejection. The reduced immunoglobulin concentrations in the interstitial tissue, confirmed in patients, may have important implications for biotherapies such as vaccines and monoclonal antibodies. PMID:29202467

Pancreatic islet cell tumor

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... cell tumors; Islet of Langerhans tumor; Neuroendocrine tumors; Peptic ulcer - islet cell tumor; Hypoglycemia - islet cell tumor ... stomach acid. Symptoms may include: Abdominal pain Diarrhea ... and small bowel Vomiting blood (occasionally) Glucagonomas make ...

Human Islet Amyloid Polypeptide

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Kosicka, Iga

2014-01-01

Diabetes mellitus type II is a metabolic disease affecting millions of people worldwide. The disease is associated with occurence of insoluble, fibrillar, protein aggregates in islets of Langerhans in the pancreas - islet amyloid. The main constituent of these protein fibers is the human islet...... of diabetes type II, while revealing the structure(s) of islet amyloid fibrils is necessary for potential design of therapeutic agents....

The Placental Secretome: Identifying Potential Cross-Talk Between Placenta and Islet β-Cells

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Robert Drynda

2018-02-01

Full Text Available Background/Aims: Insulin-secreting islet β-cells adapt to the insulin resistance associated with pregnancy by increasing functional β-cell mass, but the placental signals involved in this process are not well defined. In the current study, we analysed expression of G-protein coupled receptor (GPCR mRNAs in mouse islets and islet GPCR ligand mRNAs in placenta during pregnancy to generate an atlas of potential interactions between the placenta and β-cells to inform future functional studies of islet adaptive responses to pregnancy. Methods: Quantative RT-PCR arrays were used to measure mRNA expression levels of: (i 342 GPCRs in islets from non-pregnant mice, and in islets isolated from mice on gestational days 12 and 18; (ii 126 islet GPCR ligands in mouse placenta at gestational days 12 and 18. Results: At gestational day 12, a time of rapid expansion of the β-cell mass, 189 islet GPCR mRNAs were quantifiable, while 79 of the 126 known islet GPCR ligand mRNAs were detectable in placental extracts. Approximately half of the quantifiable placental GPCR ligand genes were of unknown function in β-cells. The expression of some islet GPCR and placental ligand mRNAs varied during pregnancy, with altered expression of both GPCR and ligand mRNAs by gestational day 18. Conclusion: The current study has revealed numerous potential routes for interaction between the placenta and islets, and offers an atlas to inform further functional studies of their roles in adaptive responses to pregnancy, and in the regulation of the β-cell mass.

A novel high-throughput assay for islet respiration reveals uncoupling of rodent and human islets.

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Jakob D Wikstrom

Full Text Available The pancreatic beta cell is unique in its response to nutrient by increased fuel oxidation. Recent studies have demonstrated that oxygen consumption rate (OCR may be a valuable predictor of islet quality and long term nutrient responsiveness. To date, high-throughput and user-friendly assays for islet respiration are lacking. The aim of this study was to develop such an assay and to examine bioenergetic efficiency of rodent and human islets.The XF24 respirometer platform was adapted to islets by the development of a 24-well plate specifically designed to confine islets. The islet plate generated data with low inter-well variability and enabled stable measurement of oxygen consumption for hours. The F1F0 ATP synthase blocker oligomycin was used to assess uncoupling while rotenone together with myxothiazol/antimycin was used to measure the level of non-mitochondrial respiration. The use of oligomycin in islets was validated by reversing its effect in the presence of the uncoupler FCCP. Respiratory leak averaged to 59% and 49% of basal OCR in islets from C57Bl6/J and FVB/N mice, respectively. In comparison, respiratory leak of INS-1 cells and C2C12 myotubes was measured to 38% and 23% respectively. Islets from a cohort of human donors showed a respiratory leak of 38%, significantly lower than mouse islets.The assay for islet respiration presented here provides a novel tool that can be used to study islet mitochondrial function in a relatively high-throughput manner. The data obtained in this study shows that rodent islets are less bioenergetically efficient than human islets as well as INS1 cells.

Islet Transplantation without Borders Enabling islet transplantation in Greece with international collaboration and innovative technology

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Papas, Klearchos K; Karatzas, Theodore; Berney, Thierry; Minor, Thomas; Pappas, Paris; Pattou, François; Shaw, James; Toso, Christian; Schuurman, Henk-Jan

2012-01-01

Recently, initiatives have been undertaken to establish an islet transplantation program in Athens, Greece. A major hurtle is the high cost associated with the establishment and maintenance of a clinical-grade islet manufacturing center. A collaboration was established with the University Hospitals of Geneva, Switzerland, to enable remote islet cell manufacturing with an established and validated fully operational team. However, remote islet manufacturing requires shipment of the pancreas from the procurement to the islet manufacturing site (in this case from anywhere in Greece to Geneva) and then shipment of the islets from the manufacturing site to the transplant site (from Geneva to Athens). To address challenges related to cold ischemia time of the pancreas and shipment time of islets, a collaboration was initiated with the University of Arizona, Tucson, USA. An international workshop was held in Athens, December 2011, to mark the start of this collaborative project. Experts in the field presented in three main sessions: [1] Islet transplantation: state-of-the-art, and the “network approach”; [2] Technical aspects of clinical islet transplantation and outcomes; and [3] Islet manufacturing – from the donated pancreas to the islet product. This manuscript presents a summary of the workshop. PMID:23330863

Autologous islet transplantation with remote islet isolation after pancreas resection for chronic pancreatitis.

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Tai, Denise S; Shen, Na; Szot, Gregory L; Posselt, Andrew; Feduska, Nicholas J; Habashy, Andrew; Clerkin, Barbara; Core, Erin; Busuttil, Ronald W; Hines, O Joe; Reber, Howard A; Lipshutz, Gerald S

2015-02-01

Autologous islet transplantation is an elegant and effective method for preserving euglycemia in patients undergoing near-total or total pancreatectomy for severe chronic pancreatitis. However, few centers worldwide perform this complex procedure, which requires interdisciplinary coordination and access to a sophisticated Food and Drug Administration-licensed islet-isolating facility. To investigate outcomes from a single institutional case series of near-total or total pancreatectomy and autologous islet transplantation using remote islet isolation. Retrospective cohort study between March 1, 2007, and December 31, 2013, at tertiary academic referral centers among 9 patients (age range, 13-47 years) with chronic pancreatitis and reduced quality of life after failed medical management. Pancreas resection, followed by transport to a remote facility for islet isolation using a modified Ricordi technique, with immediate transplantation via portal vein infusion. Islet yield, pain assessment, insulin requirement, costs, and transport time. Eight of nine patients had successful islet isolation after near-total or total pancreatectomy. Four of six patients with total pancreatectomy had islet yields exceeding 5000 islet equivalents per kilogram of body weight. At 2 months after surgery, all 9 patients had significantly reduced pain or were pain free. Of these patients, 2 did not require insulin, and 1 required low doses. The mean transport cost was $16,527, and the mean transport time was 3½ hours. Pancreatic resection with autologous islet transplantation for severe chronic pancreatitis is a safe and effective final alternative to ameliorate debilitating pain and to help prevent the development of surgical diabetes. Because many centers lack access to an islet-isolating facility, we describe our experience using a regional 2-center collaboration as a successful model to remotely isolate cells, with outcomes similar to those of larger case series.

Islet Assessment for Transplantation

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Papas, Klearchos K.; Suszynski, Thomas M.; Colton, Clark. K.

2010-01-01

Purpose of review There is a critical need for meaningful viability and potency assays that characterize islet preparations for release prior to clinical islet cell transplantation (ICT). Development, testing, and validation of such assays have been the subject of intense investigation for the past decade. These efforts are reviewed, highlighting the most recent results while focusing on the most promising assays. Recent Findings Assays based on membrane integrity do not reflect true viability when applied to either intact islets or dispersed islet cells. Assays requiring disaggregation of intact islets into individual cells for assessment introduce additional problems of cell damage and loss. Assays evaluating mitochondrial function, specifically mitochondrial membrane potential, bioenergetic status, and cellular oxygen consumption rate (OCR), especially when conducted with intact islets, appear most promising in evaluating their quality prior to ICT. Prospective, quantitative assays based on measurements of OCR with intact islets have been developed, validated and their results correlated with transplant outcomes in the diabetic nude mouse bioassay. Conclusion More sensitive and reliable islet viability and potency tests have been recently developed and tested. Those evaluating mitochondrial function are most promising, correlate with transplant outcomes in mice, and are currently being evaluated in the clinical setting. PMID:19812494

Islet Oxygen Consumption Rate (OCR) Dose Predicts Insulin Independence in Clinical Islet Autotransplantation.

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Papas, Klearchos K; Bellin, Melena D; Sutherland, David E R; Suszynski, Thomas M; Kitzmann, Jennifer P; Avgoustiniatos, Efstathios S; Gruessner, Angelika C; Mueller, Kathryn R; Beilman, Gregory J; Balamurugan, Appakalai N; Loganathan, Gopalakrishnan; Colton, Clark K; Koulmanda, Maria; Weir, Gordon C; Wilhelm, Josh J; Qian, Dajun; Niland, Joyce C; Hering, Bernhard J

2015-01-01

Reliable in vitro islet quality assessment assays that can be performed routinely, prospectively, and are able to predict clinical transplant outcomes are needed. In this paper we present data on the utility of an assay based on cellular oxygen consumption rate (OCR) in predicting clinical islet autotransplant (IAT) insulin independence (II). IAT is an attractive model for evaluating characterization assays regarding their utility in predicting II due to an absence of confounding factors such as immune rejection and immunosuppressant toxicity. Membrane integrity staining (FDA/PI), OCR normalized to DNA (OCR/DNA), islet equivalent (IE) and OCR (viable IE) normalized to recipient body weight (IE dose and OCR dose), and OCR/DNA normalized to islet size index (ISI) were used to characterize autoislet preparations (n = 35). Correlation between pre-IAT islet product characteristics and II was determined using receiver operating characteristic analysis. Preparations that resulted in II had significantly higher OCR dose and IE dose (p<0.001). These islet characterization methods were highly correlated with II at 6-12 months post-IAT (area-under-the-curve (AUC) = 0.94 for IE dose and 0.96 for OCR dose). FDA/PI (AUC = 0.49) and OCR/DNA (AUC = 0.58) did not correlate with II. OCR/DNA/ISI may have some utility in predicting outcome (AUC = 0.72). Commonly used assays to determine whether a clinical islet preparation is of high quality prior to transplantation are greatly lacking in sensitivity and specificity. While IE dose is highly predictive, it does not take into account islet cell quality. OCR dose, which takes into consideration both islet cell quality and quantity, may enable a more accurate and prospective evaluation of clinical islet preparations.

Islet Oxygen Consumption Rate (OCR Dose Predicts Insulin Independence in Clinical Islet Autotransplantation.

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Klearchos K Papas

Full Text Available Reliable in vitro islet quality assessment assays that can be performed routinely, prospectively, and are able to predict clinical transplant outcomes are needed. In this paper we present data on the utility of an assay based on cellular oxygen consumption rate (OCR in predicting clinical islet autotransplant (IAT insulin independence (II. IAT is an attractive model for evaluating characterization assays regarding their utility in predicting II due to an absence of confounding factors such as immune rejection and immunosuppressant toxicity.Membrane integrity staining (FDA/PI, OCR normalized to DNA (OCR/DNA, islet equivalent (IE and OCR (viable IE normalized to recipient body weight (IE dose and OCR dose, and OCR/DNA normalized to islet size index (ISI were used to characterize autoislet preparations (n = 35. Correlation between pre-IAT islet product characteristics and II was determined using receiver operating characteristic analysis.Preparations that resulted in II had significantly higher OCR dose and IE dose (p<0.001. These islet characterization methods were highly correlated with II at 6-12 months post-IAT (area-under-the-curve (AUC = 0.94 for IE dose and 0.96 for OCR dose. FDA/PI (AUC = 0.49 and OCR/DNA (AUC = 0.58 did not correlate with II. OCR/DNA/ISI may have some utility in predicting outcome (AUC = 0.72.Commonly used assays to determine whether a clinical islet preparation is of high quality prior to transplantation are greatly lacking in sensitivity and specificity. While IE dose is highly predictive, it does not take into account islet cell quality. OCR dose, which takes into consideration both islet cell quality and quantity, may enable a more accurate and prospective evaluation of clinical islet preparations.

Islet β cell failure in type 2 diabetes

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Prentki, Marc; Nolan, Christopher J.

2006-01-01

The major focus of this Review is on the mechanisms of islet β cell failure in the pathogenesis of obesity-associated type 2 diabetes (T2D). As this demise occurs within the context of β cell compensation for insulin resistance, consideration is also given to the mechanisms involved in the compensation process, including mechanisms for expansion of β cell mass and for enhanced β cell performance. The importance of genetic, intrauterine, and environmental factors in the determination of “susceptible” islets and overall risk for T2D is reviewed. The likely mechanisms of β cell failure are discussed within the two broad categories: those with initiation and those with progression roles. PMID:16823478

Islet oxygen consumption rate (OCR) dose predicts insulin independence for first clinical islet allotransplants

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Kitzmann, JP; O’Gorman, D; Kin, T; Gruessner, AC; Senior, P; Imes, S; Gruessner, RW; Shapiro, AMJ; Papas, KK

2014-01-01

Human islet allotransplant (ITx) for the treatment of type 1 diabetes is in phase III clinical registration trials in the US and standard of care in several other countries. Current islet product release criteria include viability based on cell membrane integrity stains, glucose stimulated insulin release (GSIR), and islet equivalent (IE) dose based on counts. However, only a fraction of patients transplanted with islets that meet or exceed these release criteria become insulin independent following one transplant. Measurements of islet oxygen consumption rate (OCR) have been reported as highly predictive of transplant outcome in many models. In this paper we report on the assessment of clinical islet allograft preparations using islet oxygen consumption rate (OCR) dose (or viable IE dose) and current product release assays in a series of 13 first transplant recipients. The predictive capability of each assay was examined and successful graft function was defined as 100% insulin independence within 45 days post-transplant. Results showed that OCR dose was most predictive of CTO. IE dose was also highly predictive, while GSIR and membrane integrity stains were not. In conclusion, OCR dose can predict CTO with high specificity and sensitivity and is a useful tool for evaluating islet preparations prior to clinical ITx. PMID:25131089

ALK5 inhibition maintains islet endothelial cell survival but does not enhance islet graft revascularisation or function.

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King, A J F; Clarkin, C E; Austin, A L F; Ajram, L; Dhunna, J K; Jamil, M O; Ditta, S I; Ibrahim, S; Raza, Z; Jones, P M

2015-01-01

Islet transplantation is a potential treatment for Type 1 diabetes but long term graft function is suboptimal. The rich supply of intraislet endothelial cells diminishes rapidly after islet isolation and culture, which affects the revascularisation rate of islets after transplantation. The ALK5 pathway inhibits endothelial cell proliferation and thus inhibiting ALK5 is a potential target for improving endothelial cell survival. The aim of the study was to establish whether ALK5 inhibition prevents the loss of intraislet endothelial cells during islet culture and thus improves the functional survival of transplanted islets by enhancing their subsequent revascularisation after implantation. Islets were cultured for 48 h in the absence or presence of 2 different ALK inhibitors: SB-431542 or A-83-01. Their vascular density after culture was analysed using immunohistochemistry. Islets pre-cultured with the ALK5 inhibitors were implanted into streptozotocin-diabetic mice for either 3 or 7 days and blood glucose concentrations were monitored and vascular densities of the grafts were analysed. Islets cultured with ALK5 inhibitors had higher vascular densities than control-cultured islets. Three days after implantation, endothelial cell numbers in islet grafts were minimal, irrespective of treatment during culture. Seven days after implantation, endothelial cells were evident within the islet grafts but there was no difference between control-cultured islets and islets pre-treated with an ALK5 inhibitor. Blood glucose concentrations were no different between the treatment groups. In conclusion, inhibition of ALK5 improved intraislet endothelial cell numbers after islet culture, but this effect was lost in the early post-transplantation period. © Georg Thieme Verlag KG Stuttgart · New York.

Pleckstrin homology-like domain family A, member 3 (PHLDA3 deficiency improves islets engraftment through the suppression of hypoxic damage.

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Naoaki Sakata

Full Text Available Islet transplantation is a useful cell replacement therapy that can restore the glycometabolic function of severe diabetic patients. It is known that many transplanted islets failed to engraft, and thus, new approaches for overcoming graft loss that may improve the outcome of future clinical islet transplantations are necessary. Pleckstrin homology-like domain family A, member 3 (PHLDA3 is a known suppressor of neuroendocrine tumorigenicity, yet deficiency of this gene increases islet proliferation, prevents islet apoptosis, and improves their insulin-releasing function without causing tumors. In this study, we examined the potential use of PHLDA3-deficient islets in transplantation. We observed that: 1 transplanting PHLDA3-deficient islets into diabetic mice significantly improved their glycometabolic condition, 2 the improved engraftment of PHLDA3-deficient islets resulted from increased cell survival during early transplantation, and 3 Akt activity was elevated in PHLDA3-deficient islets, especially under hypoxic conditions. Thus, we determined that PHLDA3-deficient islets are more resistant against stresses induced by islet isolation and transplantation. We conclude that use of islets with suppressed PHLDA3 expression could be a novel and promising treatment for improving engraftment and consequent glycemic control in islet transplantation.

International workshop: islet transplantation without borders enabling islet transplantation in Greece with international collaboration and innovative technology.

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Papas, Klearchos K; Karatzas, Theodore; Berney, Thierry; Minor, Thomas; Pappas, Paris; Pattou, François; Shaw, James; Toso, Christian; Schuurman, Henk-Jan

2013-01-01

Recently, initiatives have been undertaken to establish an islet transplantation program in Athens, Greece. A major hurdle is the high cost associated with the establishment and maintenance of a clinical-grade islet manufacturing center. A collaboration was established with the University Hospitals of Geneva, Switzerland, to enable remote islet cell manufacturing with an established and validated fully operational team. However, remote islet manufacturing requires shipment of the pancreas from the procurement to the islet manufacturing site (in this case from anywhere in Greece to Geneva) and then shipment of the islets from the manufacturing site to the transplant site (from Geneva to Athens). To address challenges related to cold ischemia time of the pancreas and shipment time of islets, a collaboration was initiated with the University of Arizona, Tucson, USA. An international workshop was held in Athens, December 2011, to mark the start of this collaborative project. Experts in the field presented in three main sessions: (i) islet transplantation: state-of-the-art and the "network approach"; (ii) technical aspects of clinical islet transplantation and outcomes; and (iii) islet manufacturing - from the donated pancreas to the islet product. This manuscript presents a summary of the workshop. © 2013 John Wiley & Sons A/S.

Oxygenation of the Intraportally Transplanted Pancreatic Islet.

Science.gov (United States)

Suszynski, Thomas M; Avgoustiniatos, Efstathios S; Papas, Klearchos K

2016-01-01

Intraportal islet transplantation (IT) is not widely utilized as a treatment for type 1 diabetes. Oxygenation of the intraportally transplanted islet has not been studied extensively. We present a diffusion-reaction model that predicts the presence of an anoxic core and a larger partly functional core within intraportally transplanted islets. Four variables were studied: islet diameter, islet fractional viability, external oxygen partial pressure ( P ) (in surrounding portal blood), and presence or absence of a thrombus on the islet surface. Results indicate that an islet with average size and fractional viability exhibits an anoxic volume fraction (AVF) of 14% and a function loss of 72% at a low external P . Thrombus formation increased AVF to 30% and function loss to 92%, suggesting that the effect of thrombosis may be substantial. External P and islet diameter accounted for the greatest overall impact on AVF and loss of function. At our institutions, large human alloislets (>200 μ m diameter) account for ~20% of total islet number but ~70% of total islet volume; since most of the total transplanted islet volume is accounted for by large islets, most of the intraportal islet cells are likely to be anoxic and not fully functional.

Compensatory hyperinsulinemia in high-fat diet-induced obese mice is associated with enhanced insulin translation in islets

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Kanno, Ayumi, E-mail: akanno@med.kobe-u.ac.jp [Division of Diabetes and Endocrinology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe 650-0017 (Japan); Asahara, Shun-ichiro, E-mail: asahara@med.kobe-u.ac.jp [Division of Diabetes and Endocrinology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe 650-0017 (Japan); Masuda, Katsuhisa, E-mail: katsuhisa.m.0707@gmail.com [Division of Medical Chemistry, Department of Biophysics, Kobe University Graduate School of Health Sciences, Kobe 654-0142 (Japan); Matsuda, Tomokazu, E-mail: tomokazu@med.kobe-u.ac.jp [Division of Diabetes and Endocrinology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe 650-0017 (Japan); Kimura-Koyanagi, Maki, E-mail: koyanagi@med.kobe-u.ac.jp [Division of Diabetes and Endocrinology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe 650-0017 (Japan); Seino, Susumu, E-mail: seino@med.kobe-u.ac.jp [Division of Molecular and Metabolic Medicine, Department of Physiology and Cell Biology, Kobe University Graduate School of Medicine, Kobe 650-0047 (Japan); Ogawa, Wataru, E-mail: ogawa@med.kobe-u.ac.jp [Division of Diabetes and Endocrinology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe 650-0017 (Japan); Kido, Yoshiaki, E-mail: kido@med.kobe-u.ac.jp [Division of Diabetes and Endocrinology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe 650-0017 (Japan); Division of Medical Chemistry, Department of Biophysics, Kobe University Graduate School of Health Sciences, Kobe 654-0142 (Japan)

2015-03-13

A high-fat diet (HF) is associated with obesity, insulin resistance, and hyperglycemia. Animal studies have shown compensatory mechanisms in pancreatic β-cells after high fat load, such as increased pancreatic β-cell mass, enhanced insulin secretion, and exocytosis. However, the effects of high fat intake on insulin synthesis are obscure. Here, we investigated whether insulin synthesis was altered in correlation with an HF diet, for the purpose of obtaining further understanding of the compensatory mechanisms in pancreatic β-cells. Mice fed an HF diet are obese, insulin resistant, hyperinsulinemic, and glucose intolerant. In islets of mice fed an HF diet, more storage of insulin was identified. We analyzed insulin translation in mouse islets, as well as in INS-1 cells, using non-radioisotope chemicals. We found that insulin translational levels were significantly increased in islets of mice fed an HF diet to meet systemic demand, without altering its transcriptional levels. Our data showed that not only increased pancreatic β-cell mass and insulin secretion but also elevated insulin translation is the major compensatory mechanism of pancreatic β-cells. - Highlights: • More stored insulin was recognized in islets of mice fed a high-fat diet. • Insulin translation was not enhanced by fatty acids, but by insulin demand. • Insulin transcription was not altered in islets of mice fed a high-fat diet. • Insulin translation was markedly enhanced in islets of mice fed a high-fat diet. • Non-radioisotope chemicals were used to measure insulin translation in mouse islets.

Compensatory hyperinsulinemia in high-fat diet-induced obese mice is associated with enhanced insulin translation in islets

International Nuclear Information System (INIS)

Kanno, Ayumi; Asahara, Shun-ichiro; Masuda, Katsuhisa; Matsuda, Tomokazu; Kimura-Koyanagi, Maki; Seino, Susumu; Ogawa, Wataru; Kido, Yoshiaki

2015-01-01

A high-fat diet (HF) is associated with obesity, insulin resistance, and hyperglycemia. Animal studies have shown compensatory mechanisms in pancreatic β-cells after high fat load, such as increased pancreatic β-cell mass, enhanced insulin secretion, and exocytosis. However, the effects of high fat intake on insulin synthesis are obscure. Here, we investigated whether insulin synthesis was altered in correlation with an HF diet, for the purpose of obtaining further understanding of the compensatory mechanisms in pancreatic β-cells. Mice fed an HF diet are obese, insulin resistant, hyperinsulinemic, and glucose intolerant. In islets of mice fed an HF diet, more storage of insulin was identified. We analyzed insulin translation in mouse islets, as well as in INS-1 cells, using non-radioisotope chemicals. We found that insulin translational levels were significantly increased in islets of mice fed an HF diet to meet systemic demand, without altering its transcriptional levels. Our data showed that not only increased pancreatic β-cell mass and insulin secretion but also elevated insulin translation is the major compensatory mechanism of pancreatic β-cells. - Highlights: • More stored insulin was recognized in islets of mice fed a high-fat diet. • Insulin translation was not enhanced by fatty acids, but by insulin demand. • Insulin transcription was not altered in islets of mice fed a high-fat diet. • Insulin translation was markedly enhanced in islets of mice fed a high-fat diet. • Non-radioisotope chemicals were used to measure insulin translation in mouse islets

Cellular islet autoimmunity associates with clinical outcome of islet cell transplantation.

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Volkert A L Huurman

2008-06-01

Full Text Available Islet cell transplantation can cure type 1 diabetes (T1D, but only a minority of recipients remains insulin-independent in the following years. We tested the hypothesis that allograft rejection and recurrent autoimmunity contribute to this progressive loss of islet allograft function.Twenty-one T1D patients received cultured islet cell grafts prepared from multiple donors and transplanted under anti-thymocyte globulin (ATG induction and tacrolimus plus mycophenolate mofetil (MMF maintenance immunosuppression. Immunity against auto- and alloantigens was measured before and during one year after transplantation. Cellular auto- and alloreactivity was assessed by lymphocyte stimulation tests against autoantigens and cytotoxic T lymphocyte precursor assays, respectively. Humoral reactivity was measured by auto- and alloantibodies. Clinical outcome parameters--including time until insulin independence, insulin independence at one year, and C-peptide levels over one year--remained blinded until their correlation with immunological parameters. All patients showed significant improvement of metabolic control and 13 out of 21 became insulin-independent. Multivariate analyses showed that presence of cellular autoimmunity before and after transplantation is associated with delayed insulin-independence (p = 0.001 and p = 0.01, respectively and lower circulating C-peptide levels during the first year after transplantation (p = 0.002 and p = 0.02, respectively. Seven out of eight patients without pre-existent T-cell autoreactivity became insulin-independent, versus none of the four patients reactive to both islet autoantigens GAD and IA-2 before transplantation. Autoantibody levels and cellular alloreactivity had no significant association with outcome.In this cohort study, cellular islet-specific autoimmunity associates with clinical outcome of islet cell transplantation under ATG-tacrolimus-MMF immunosuppression. Tailored immunotherapy targeting cellular

Oxygenation of the Intraportally Transplanted Pancreatic Islet

Directory of Open Access Journals (Sweden)

Thomas M. Suszynski

2016-01-01

Full Text Available Intraportal islet transplantation (IT is not widely utilized as a treatment for type 1 diabetes. Oxygenation of the intraportally transplanted islet has not been studied extensively. We present a diffusion-reaction model that predicts the presence of an anoxic core and a larger partly functional core within intraportally transplanted islets. Four variables were studied: islet diameter, islet fractional viability, external oxygen partial pressure (P (in surrounding portal blood, and presence or absence of a thrombus on the islet surface. Results indicate that an islet with average size and fractional viability exhibits an anoxic volume fraction (AVF of 14% and a function loss of 72% at a low external P. Thrombus formation increased AVF to 30% and function loss to 92%, suggesting that the effect of thrombosis may be substantial. External P and islet diameter accounted for the greatest overall impact on AVF and loss of function. At our institutions, large human alloislets (>200 μm diameter account for ~20% of total islet number but ~70% of total islet volume; since most of the total transplanted islet volume is accounted for by large islets, most of the intraportal islet cells are likely to be anoxic and not fully functional.

Application of Digital Image Analysis to Determine Pancreatic Islet Mass and Purity in Clinical Islet Isolation and Transplantation

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Wang, Ling-jia; Kissler, Hermann J; Wang, Xiaojun; Cochet, Olivia; Krzystyniak, Adam; Misawa, Ryosuke; Golab, Karolina; Tibudan, Martin; Grzanka, Jakub; Savari, Omid; Grose, Randall; Kaufman, Dixon B; Millis, Michael; Witkowski, Piotr

2015-01-01

Pancreatic islet mass, represented by islet equivalent (IEQ), is the most important parameter in decision making for clinical islet transplantation. To obtain IEQ, the sample of islets is routinely counted manually under a microscope and discarded thereafter. Islet purity, another parameter in islet processing, is routinely acquired by estimation only. In this study, we validated our digital image analysis (DIA) system developed using the software of Image Pro Plus for islet mass and purity assessment. Application of the DIA allows to better comply with current good manufacturing practice (cGMP) standards. Human islet samples were captured as calibrated digital images for the permanent record. Five trained technicians participated in determination of IEQ and purity by manual counting method and DIA. IEQ count showed statistically significant correlations between the manual method and DIA in all sample comparisons (r >0.819 and p islet particle number (IPN) and the IEQ/IPN ratio did not differ statistically between manual counting method and DIA. In conclusion, the DIA used in this study is a reliable technique in determination of IEQ and purity. Islet sample preserved as a digital image and results produced by DIA can be permanently stored for verification, technical training and islet information exchange between different islet centers. Therefore, DIA complies better with cGMP requirements than the manual counting method. We propose DIA as a quality control tool to supplement the established standard manual method for islets counting and purity estimation. PMID:24806436

Islet amyloid polypeptide and high hydrostatic pressure: towards an understanding of the fibrillization process

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Lopes, D. H. J.; Smirnovas, V.; Winter, R.

2008-07-01

Type II Diabetes Mellitus is a disease which is characterized by peripheral insulin resistance coupled with a progressive loss of insulin secretion that is associated with a decrease in pancreatic islet β-cell mass and the deposition of amyloid in the extracellular matrix of β-cells, which lead to islet cell death. The principal component of the islet amyloid is a pancreatic hormone called islet amyloid polypeptide (IAPP). High-pressure coupled with FT-IR, CD, ThT fluorescence spectroscopic and AFM studies were carried out to reveal information on the aggregation pathway as well as the aggregate structure of IAPP. Our data indicate that IAPP pre-formed fibrils exhibit a strong polymorphism with heterogeneous structures very sensitive to high hydrostatic pressure, indicating a high percentage of ionic and hydrophobic interactions being responsible for the stability the IAPP fibrils.

Human islet viability and function is maintained during high density shipment in silicone rubber membrane vessels

Science.gov (United States)

Kitzmann, Jennifer P; Pepper, Andrew R; Lopez, Boris G; Pawlick, Rena; Kin, Tatsuya; O’Gorman, Doug; Mueller, Kathryn R; Gruessner, Angelika C; Avgoustiniatos, Efstathios S; Karatzas, Theodore; Szot, Greg L; Posselt, Andrew M; Stock, Peter G; Wilson, John R; Shapiro, AM; Papas, Klearchos K

2014-01-01

The shipment of human islets from processing centers to distant laboratories is beneficial for both research and clinical applications. The maintenance of islet viability and function in transit is critically important. Gas-permeable silicone rubber membrane (SRM) vessels reduce the risk of hypoxia-induced death or dysfunction during high-density islet culture or shipment. SRM vessels may offer additional advantages: they are cost-effective (fewer flasks, less labor needed), safer (lower contamination risk), and simpler (culture vessel can also be used for shipment). Human islets(IE) were isolated from two manufacturing centers and shipped in 10cm2 surface area SRM vessels in temperature and pressure controlled containers to a distant center following at least two days of culture (n = 6). Three conditions were examined: low density (LD), high density (HD), and a micro centrifuge tube negative control (NC). LD was designed to mimic the standard culture density for human islet preparations (200 IE/cm2), while HD was designed to have a 20-fold higher tissue density, which would enable the culture of an entire human isolation in 1–3 vessels. Upon receipt, islets were assessed for viability, measured by oxygen consumption rate normalized to DNA content (OCR/DNA), and quantity, measured by DNA, and, when possible, potency and function with dynamic glucose-stimulated insulin secretion (GSIS) measurements and transplants in immunodeficient B6 rag mice. Post-shipment OCR/DNA was not reduced in HD versus LD, and was substantially reduced in the NC condition. HD islets exhibited normal function post-shipment. Based on the data we conclude that entire islet isolations (up to 400,000 IE) may be shipped using a single, larger SRM vessel with no negative effect on viability and ex vivo and in vivo function. PMID:25131090

Clinical pancreatic islet transplantation.

Science.gov (United States)

Shapiro, A M James; Pokrywczynska, Marta; Ricordi, Camillo

2017-05-01

Clinical pancreatic islet transplantation can be considered one of the safest and least invasive transplant procedures. Remarkable progress has occurred in both the technical aspects of islet cell processing and the outcomes of clinical islet transplantation. With >1,500 patients treated since 2000, this therapeutic strategy has moved from a curiosity to a realistic treatment option for selected patients with type 1 diabetes mellitus (that is, those with hypoglycaemia unawareness, severe hypoglycaemic episodes and glycaemic lability). This Review outlines the techniques required for human islet isolation, in vitro culture before the transplant and clinical islet transplantation, and discusses indications, optimization of recipient immunosuppression and management of adjunctive immunomodulatory and anti-inflammatory strategies. The potential risks, long-term outcomes and advances in treatment after the transplant are also discussed to further move this treatment towards becoming a more widely available option for patients with type 1 diabetes mellitus and eventually a potential cure.

The impact of IUGR on pancreatic islet development and β-cell function.

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Boehmer, Brit H; Limesand, Sean W; Rozance, Paul J

2017-11-01

Placental insufficiency is a primary cause of intrauterine growth restriction (IUGR). IUGR increases the risk of developing type 2 diabetes mellitus (T2DM) throughout life, which indicates that insults from placental insufficiency impair β-cell development during the perinatal period because β-cells have a central role in the regulation of glucose tolerance. The severely IUGR fetal pancreas is characterized by smaller islets, less β-cells, and lower insulin secretion. Because of the important associations among impaired islet growth, β-cell dysfunction, impaired fetal growth, and the propensity for T2DM, significant progress has been made in understanding the pathophysiology of IUGR and programing events in the fetal endocrine pancreas. Animal models of IUGR replicate many of the observations in severe cases of human IUGR and allow us to refine our understanding of the pathophysiology of developmental and functional defects in islet from IUGR fetuses. Almost all models demonstrate a phenotype of progressive loss of β-cell mass and impaired β-cell function. This review will first provide evidence of impaired human islet development and β-cell function associated with IUGR and the impact on glucose homeostasis including the development of glucose intolerance and diabetes in adulthood. We then discuss evidence for the mechanisms regulating β-cell mass and insulin secretion in the IUGR fetus, including the role of hypoxia, catecholamines, nutrients, growth factors, and pancreatic vascularity. We focus on recent evidence from experimental interventions in established models of IUGR to understand better the pathophysiological mechanisms linking placental insufficiency with impaired islet development and β-cell function. © 2017 Society for Endocrinology.

Overexpression of thioredoxin in islets transduced by a lentiviral vector prolongs graft survival in autoimmune diabetic NOD mice

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Sytwu Huey-Kang

2009-08-01

Full Text Available Abstract Pancreatic islet transplantation is considered an appropriate treatment to achieve insulin independence in type I diabetic patients. However, islet isolation and transplantation-induced oxidative stress and autoimmune-mediated destruction are still the major obstacles to the long-term survival of graft islets in this potential therapy. To protect islet grafts from inflammatory damage and prolong their survival, we transduced islets with an antioxidative gene thioredoxin (TRX using a lentiviral vector before transplantation. We hypothesized that the overexpression of TRX in islets would prolong islet graft survival when transplanted into diabetic non-obese diabetic (NOD mice. Methods Islets were isolated from NOD mice and transduced with lentivirus carrying TRX (Lt-TRX or enhanced green fluorescence protein (Lt-eGFP, respectively. Transduced islets were transplanted under the left kidney capsule of female diabetic NOD mice, and blood glucose concentration was monitored daily after transplantation. The histology of the islet graft was assessed at the end of the study. The protective effect of TRX on islets was investigated. Results The lentiviral vector effectively transduced islets without altering the glucose-stimulating insulin-secretory function of islets. Overexpression of TRX in islets reduced hydrogen peroxide-induced cytotoxicity in vitro. After transplantation into diabetic NOD mice, euglycemia was maintained for significantly longer in Lt-TRX-transduced islets than in Lt-eGFP-transduced islets; the mean graft survival was 18 vs. 6.5 days (n = 9 and 10, respectively, p Conclusion We successfully transduced the TRX gene into islets and demonstrated that these genetically modified grafts are resistant to inflammatory insult and survived longer in diabetic recipients. Our results further support the concept that the reactive oxygen species (ROS scavenger and antiapoptotic functions of TRX are critical to islet survival after

Organ culture studies for pancreatic islet transplantation

International Nuclear Information System (INIS)

Reemtsma, K.; Weber, C.J.; Pi-Sunyer, F.X.; Lerner, R.; Zimmerman, E.; Hardy, M.A.

1979-01-01

Data support the usefulness of tissue culture in isolation and preservation of islets prior to transplantation. Rodent islet viability in culture was demonstrated histologically and by functional analyses of hormone production. For reasons that remain to be defined, acinar cells disappeared rapidly in tissue culture, yielding an implant preparation relatively rich in islets and devoid of pancreatic exocrine elements. Isografts of cultured and noncultured islets were well tolerated intraperitoneally and intramuscularly; and prompt and lasting reversal of short- and long-standing experimental diabetes was observed regularly. In vitro studies of rodent islet viability after immunosuppressive treatment of donors or islet cultures showed insulin production comparable to that of control experiments, suggesting that immunologic modification of donors or islets might be feasible in eventual human islet allotransplantation

In Vivo Imaging of Transplanted Pancreatic Islets

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Donghee Kim

2018-01-01

Full Text Available The beta-cells in the islets of Langerhans in the pancreas secrete insulin and play an important role in glucose homeostasis. Diabetes, characterized by hyperglycemia, results from an absolute or a relative deficiency of the pancreatic beta-cell mass. Islet transplantation has been considered to be a useful therapeutic approach, but it is largely unsuccessful because most of the transplanted islets are lost in the early stage of transplantation. To evaluate the efficacy of intervention methods for the improvement of islet survival, monitoring of the functional islet mass is needed. Various techniques to image and track transplanted islets have been investigated to assess islets after transplantation. In this review, recent progresses in imaging methods to visualize islets are discussed.

Introducing a New Experimental Islet Transplantation Model using Biomimetic Hydrogel and a Simple High Yield Islet Isolation Technique.

Science.gov (United States)

Mohammadi Ayenehdeh, Jamal; Niknam, Bahareh; Hashemi, Seyed Mahmoud; Rahavi, Hossein; Rezaei, Nima; Soleimani, Masoud; Tajik, Nader

2017-07-01

Islet transplantation could be an ideal alternative treatment to insulin therapy for type 1 diabetes Mellitus (T1DM). This clinical and experimental field requires a model that covers problems such as requiring a large number of functional and viable islets, the optimal transplantation site, and the prevention of islet dispersion. Hence, the methods of choice for isolation of functional islets and transplantation are crucial. The present study has introduced an experimental model that overcomes some critical issues in islet transplantation, including in situ pancreas perfusion by digestive enzymes through common bile duct. In comparison with conventional methods, we inflated the pancreas in Petri dishes with only 1 ml collagenase type XI solution, which was followed by hand-picking isolation or Ficoll gradient separation to purify the islets. Then we used a hydrogel composite in which the islets were embedded and transplanted into the peritoneal cavity of the streptozotocin-induced diabetic C57BL/6 mice. As compared to the yield of the classical methods, in our modified technique, the mean yield of isolation was about 130-200 viable islets/mouse pancreas. In vitro glucose-mediated insulin secretion assay indicated an appropriate response in isolated islets. In addition, data from in vivo experiments revealed that the allograft remarkably maintained blood glucose levels under 400 mg/dl and hydrogel composite prevents the passage of immune cells. In the model presented here, the rapid islet isolation technique and the application of biomimetic hydrogel wrapping of islets could facilitate islet transplantation procedures.

Extensive Loss of Islet Mass Beyond the First Day After Intraportal Human Islet Transplantation in a Mouse Model.

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Liljebäck, Hanna; Grapensparr, Liza; Olerud, Johan; Carlsson, Per-Ola

2016-01-01

Clinical islet transplantation is characterized by a progressive deterioration of islet graft function, which renders many patients once again dependent on exogenous insulin administration within a couple of years. In this study, we aimed to investigate possible engraftment factors limiting the survival and viability of experimentally transplanted human islets beyond the first day after their transplantation to the liver. Human islets were transplanted into the liver of nude mice and characterized 1 or 30 days after transplantation by immunohistochemistry. The factors assessed were endocrine mass, cellular death, hypoxia, vascular density and amyloid formation in the transplanted islets. One day posttransplantation, necrotic cells, as well as apoptotic cells, were commonly observed. In contrast to necrotic death, apoptosis rates remained high 1 month posttransplantation, and the total islet mass was reduced by more than 50% between 1 and 30 days posttransplantation. Islet mass at 30 days posttransplantation correlated negatively to apoptotic death. Vascular density within the transplanted islets remained less than 30% of that in native human islets up to 30 days posttransplantation and was associated with prevailing hypoxia. Amyloid formation was rarely observed in the 1-day-old transplants, but was commonly observed in the 30-day-old islet transplants. We conclude that substantial islet cell death occurs beyond the immediate posttransplantation phase, particularly through apoptotic events. Concomitant low vascularization with prevailing hypoxia and progressive amyloid development was observed in the human islet grafts. Strategies to improve engraftment at the intraportal site or change of implantation site in the clinical setting are needed.

Hepatic steatosis after islet transplantation: Can ultrasound predict the clinical outcome? A longitudinal study in 108 patients.

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Venturini, Massimo; Maffi, Paola; Querques, Giulia; Agostini, Giulia; Piemonti, Lorenzo; Sironi, Sandro; De Cobelli, Francesco; Fiorina, Paolo; Secchi, Antonio; Del Maschio, Alessandro

2015-08-01

Percutaneous intra-portal islet transplantation (PIPIT) is a less invasive, safer, and repeatable therapeutic option for brittle type 1 diabetes, compared to surgical pancreas transplantation. Hepatic steatosis is a consequence of the islet engraftment but it is curiously present in a limited number of patients and its meaning is controversial. The aims of this study were to assess hepatic steatosis at ultrasound (US) after PIPIT investigating its relationship with graft function and its role in predicting the clinical outcome. From 1996 to 2012, 108 patients underwent PIPIT: 83 type-1 diabetic patients underwent allo-transplantation, 25 auto-transplantation. US was performed at baseline, 6, 12, and 24 months, recording steatosis prevalence, first detection, duration, and distribution. Contemporaneously, steatotic and non-steatotic patients were compared for the following parameters: infused islet mass, insulin independence rate, β-score, C-peptide, glycated hemoglobin, exogenous insulin requirement, and fasting plasma glucose. Steatosis at US was detected in 21/108 patients, 20/83 allo-transplanted and 1/25 auto-transplanted, mostly at 6 and 12 months. Infused islet mass was significantly higher in steatotic than non-steatotic patients (IE/kg: S=10.822; NS=6138; p=0.001). Metabolically, steatotic patients had worse basal conditions, but better islet function when steatosis was first detected, after which progressive islet exhaustion, along with steatosis disappearance, was observed. Conversely, in non-steatotic patients these parameters remained stable in time. Number of re-transplantations was significantly higher in steatotic than in non-steatotic patients (1.8 vs 1.1; p=0.001). Steatosis at US seems to be related to the islet mass and local overworking activity. It precedes metabolic alterations and can predict graft dysfunction addressing to therapeutic decisions before islet exhaustion. If steatosis does not appear, no conclusion can be drawn. Copyright �

Current issues in allogeneic islet transplantation.

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Chang, Charles A; Lawrence, Michael C; Naziruddin, Bashoo

2017-10-01

Transplantation of allogenic pancreatic islets is a minimally invasive treatment option to control severe hypoglycemia and dependence on exogenous insulin among type 1 diabetes (T1D) patients. This overview summarizes the current issues and progress in islet transplantation outcomes and research. Several clinical trials from North America and other countries have documented the safety and efficacy of clinical islet transplantation for T1D patients with impaired hypoglycemia awareness. A recently completed phase 3 clinical trial allows centres in the United States to apply for a Food and Drug Administration Biologics License for the procedure. Introduction of anti-inflammatory drugs along with T-cell depleting induction therapy has significantly improved long-term function of transplanted islets. Research into islet biomarkers, immunosuppression, extrahepatic transplant sites and potential alternative beta cell sources is driving further progress. Allogeneic islet transplantation has vastly improved over the past two decades. Success in restoration of glycemic control and hypoglycemic awareness after islet transplantation has been further highlighted by clinical trials. However, lack of effective strategies to maintain long-term islet function and insufficient sources of donor tissue still impose limitations to the widespread use of islet transplantation. In the United States, wide adoption of this technology still awaits regulatory approval and, importantly, a financial mechanism to support the use of this technology.

Effect of the Diabetic State on Islet Engraftment and Function in a Large Animal Model of Islet-Kidney Transplantation.

Science.gov (United States)

Vallabhajosyula, Prashanth; Hirakata, Atsushi; Weiss, Matthew; Griesemer, Adam; Shimizu, Akira; Hong, Hanzhou; Habertheuer, Andreas; Tchipashvili, Vaja; Yamada, Kazuhiko; Sachs, David H

2017-11-01

In islet transplantation, in addition to immunologic and ischemic factors, the diabetic/hyperglycemic state of the recipient has been proposed, although not yet validated, as a possible cause of islet toxicity, contributing to islet loss during the engraftment period. Using a miniature swine model of islet transplantation, we have now assessed the effect of a persistent state of hyperglycemia on islet engraftment and subsequent function. An islet-kidney (IK) model previously described by our laboratory was utilized. Three experimental donor animals underwent total pancreatectomy and autologous islet transplantation underneath the renal capsule to prepare an IK at a load of ≤1,000 islet equivalents (IE)/kg donor weight, leading to a chronic diabetic state during the engraftment period (fasting blood glucose >250 mg/dL). Three control donor animals underwent partial pancreatectomy (sufficient to maintain normoglycemia during islet engraftment period) and IK preparation. As in vivo functional readout for islet engraftment, the IKs were transplanted across an immunologic minor or class I mismatch barrier into diabetic, nephrectomized recipients at an islet load of ∼4,500 IE/kg recipient weight. A 12-d course of cyclosporine was administered for tolerance induction. All experimental donors became diabetic and showed signs of end organ injury, while control donors maintained normoglycemia. All recipients of IK from both experimental and control donors achieved glycemic control over long-term follow-up, with reversal of diabetic nephropathy and with similar glucose tolerance tests. In this preclinical, large animal model, neither islet engraftment nor subsequent long-term islet function after transplantation appear to be affected by the diabetic state.

Benefits of PEGylation in the early post-transplant period of intraportal islet transplantation as assessed by magnetic resonance imaging of labeled islets.

Science.gov (United States)

Jin, Sang-Man; Oh, Seung-Hoon; Oh, Bae Jun; Suh, Sunghwan; Bae, Ji Cheol; Lee, Jung Hee; Lee, Myung-Shik; Lee, Moon-Kyu; Kim, Kwang-Won; Kim, Jae Hyeon

2014-01-01

While a few studies have demonstrated the benefit of PEGylation in islet transplantation, most have employed renal subcapsular models and none have performed direct comparisons of islet mass in intraportal islet transplantation using islet magnetic resonance imaging (MRI). In this study, our aim was to demonstrate the benefit of PEGylation in the early post-transplant period of intraportal islet transplantation with a novel algorithm for islet MRI. Islets were PEGylated after ferucarbotran labeling in a rat syngeneic intraportal islet transplantation model followed by comparisons of post-transplant glycemic levels in recipient rats infused with PEGylated (n = 12) and non-PEGylated (n = 13) islets. The total area of hypointense spots and the number of hypointense spots larger than 1.758 mm(2) of PEGylated and non-PEGylated islets were quantitatively compared. The total area of hypointense spots (P islet group 7 and 14 days post translation (DPT). These results translated into better post-transplant outcomes in the PEGylated islet group 28 DPT. In validation experiments, MRI parameters obtained 1, 7, and 14 DPT predicted normoglycemia 4 wk post-transplantation. We directly demonstrated the benefit of islet PEGylation in protection against nonspecific islet destruction in the early post-transplant period of intraportal islet transplantation using a novel algorithm for islet MRI. This novel algorithm could serve as a useful tool to demonstrate such benefit in future clinical trials of islet transplantation using PEGylated islets.

An 'alpha-beta' of pancreatic islet microribonucleotides

DEFF Research Database (Denmark)

Dalgaard, Louise Torp; Eliasson, Lena

2017-01-01

. Moreover, processing of miRNAs appears to be altered by obesity, diabetes, and aging. A number of miRNAs (such as miR-7, miR-21, miR-29, miR-34a, miR-212/miR-132, miR-184, miR-200 and miR-375) are involved in mediating beta cell dysfunction and/or compensation induced by hyperglycemia, oxidative stress......, cytotoxic cytokines, and in rodent models of fetal metabolic programming prediabetes and overt diabetes. Studies of human type 2 diabetic islets underline that these miRNA families could have important roles also in human type 2 diabetes. Furthermore, there is a genuine gap of knowledge regarding mi...

Invasive assessment of coronary microvascular dysfunction in hypertrophic cardiomyopathy: the index of microvascular resistance

International Nuclear Information System (INIS)

Gutiérrez-Barrios, Alejandro; Camacho-Jurado, Francisco; Díaz-Retamino, Enrique; Gamaza-Chulián, Sergio; Agarrado-Luna, Antonio; Oneto-Otero, Jesús; Del Rio-Lechuga, Ana; Benezet-Mazuecos, Javier

2015-01-01

Summary: We present a review of microvascular dysfunction in hypertrophic cardiomyopathy (HCM) and an interesting case of a symptomatic familial HCM patient with inducible ischemia by single photon emission computed tomography. Coronary angiography revealed normal epicardial arteries. Pressure wire measurements of fractional flow reserve (FFR), coronary flow reserve (CFR) and index of microvascular resistance (IMR) demonstrated a significant microcirculatory dysfunction. This is the first such case that documents this abnormality invasively using the IMR. The measurement of IMR, a novel marker of microcirculatory dysfunction, provides novel insights into the pathophysiology of this condition. - Highlights: • Microvascular dysfunction is a common feature in hypertrophic cardiomyopathy (HCM) and represents a strong predictor of unfavorable outcome and cardiovascular mortality. • The index of microvascular resistance (IMR) is a new method for invasively assessing the state of the coronary microcirculation using a single pressure-temperature sensor-tipped coronary wire. • However assessment of IMR in HCM has not been previously reported. We report a case in which microvascular dysfunction is assessed by IMR. This index may be useful in future researches of HCM.

Invasive assessment of coronary microvascular dysfunction in hypertrophic cardiomyopathy: the index of microvascular resistance

Energy Technology Data Exchange (ETDEWEB)

Gutiérrez-Barrios, Alejandro, E-mail: aleklos@hotmail.com [Cardiology Department, Jerez Hospital, Jerez (Spain); Camacho-Jurado, Francisco [Cardiology Department, Punta Europa Hospital, Algeciras (Spain); Díaz-Retamino, Enrique; Gamaza-Chulián, Sergio; Agarrado-Luna, Antonio; Oneto-Otero, Jesús; Del Rio-Lechuga, Ana; Benezet-Mazuecos, Javier [Cardiology Department, Jerez Hospital, Jerez (Spain)

2015-10-15

Summary: We present a review of microvascular dysfunction in hypertrophic cardiomyopathy (HCM) and an interesting case of a symptomatic familial HCM patient with inducible ischemia by single photon emission computed tomography. Coronary angiography revealed normal epicardial arteries. Pressure wire measurements of fractional flow reserve (FFR), coronary flow reserve (CFR) and index of microvascular resistance (IMR) demonstrated a significant microcirculatory dysfunction. This is the first such case that documents this abnormality invasively using the IMR. The measurement of IMR, a novel marker of microcirculatory dysfunction, provides novel insights into the pathophysiology of this condition. - Highlights: • Microvascular dysfunction is a common feature in hypertrophic cardiomyopathy (HCM) and represents a strong predictor of unfavorable outcome and cardiovascular mortality. • The index of microvascular resistance (IMR) is a new method for invasively assessing the state of the coronary microcirculation using a single pressure-temperature sensor-tipped coronary wire. • However assessment of IMR in HCM has not been previously reported. We report a case in which microvascular dysfunction is assessed by IMR. This index may be useful in future researches of HCM.

Pancreas preservation for pancreas and islet transplantation

Science.gov (United States)

Iwanaga, Yasuhiro; Sutherland, David E.R.; Harmon, James V.; Papas, Klearchos K.

2010-01-01

Purpose of review To summarize advances and limitations in pancreas procurement and preservation for pancreas and islet transplantation, and review advances in islet protection and preservation. Recent findings Pancreases procured after cardiac death, with in-situ regional organ cooling, have been successfully used for islet transplantation. Colloid-free Celsior and histidine-tryptophan-ketoglutarate preservation solutions are comparable to University of Wisconsin solution when used for cold storage before pancreas transplantation. Colloid-free preservation solutions are inferior to University of Wisconsin solution for pancreas preservation prior to islet isolation and transplantation. Clinical reports on pancreas and islet transplants suggest that the two-layer method may not offer significant benefits over cold storage with the University of Wisconsin solution: improved oxygenation may depend on the graft size; benefits in experimental models may not translate to human organs. Improvements in islet yield and quality occurred from pancreases treated with inhibitors of stress-induced apoptosis during procurement, storage, isolation or culture. Pancreas perfusion may be desirable before islet isolation and transplantation and may improve islet yields and quality. Methods for real-time, noninvasive assessment of pancreas quality during preservation have been implemented and objective islet potency assays have been developed and validated. These innovations should contribute to objective evaluation and establishment of improved pancreas preservation and islet isolation strategies. Summary Cold storage may be adequate for preservation before pancreas transplants, but insufficient when pancreases are processed for islets or when expanded donors are used. Supplementation of cold storage solutions with cytoprotective agents and perfusion may improve pancreas and islet transplant outcomes. PMID:18685343

Islet transplantation using donors after cardiac death: report of the Japan Islet Transplantation Registry.

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Saito, Takuro; Gotoh, Mitsukazu; Satomi, Susumu; Uemoto, Shinji; Kenmochi, Takashi; Itoh, Toshinori; Kuroda, Yoshikazu; Yasunami, Youichi; Matsumoto, Shnichi; Teraoka, Satoshi

2010-10-15

This report summarizes outcomes of islet transplantation employing donors after cardiac death (DCD) between 2004 and 2007 as reported to the Japan Islet Transplantation Registry. Sixty-five islet isolations were performed for 34 transplantations in 18 patients with insulin-dependent diabetes mellitus, including two patients who had prior kidney transplantation. All but one donor (64/65) was DCD at the time of harvesting. Factors influencing criteria for islet release included duration of low blood pressure of the donor, cold ischemic time, and usage of Kyoto solution for preservation. Multivariate analysis selected usage of Kyoto solution as most important. Of the 18 recipients, 8, 4, and 6 recipients received 1, 2, and 3 islet infusions, respectively. Overall graft survival defined as C-peptide level more than or equal to 0.3 ng/mL was 76.5%, 47.1%, and 33.6% at 1, 2, and 3 years, respectively, whereas corresponding graft survival after multiple transplantations was 100%, 80.0%, and 57.1%, respectively. All recipients remained free of severe hypoglycemia while three achieved insulin independence for 14, 79, and 215 days. HbA1c levels and requirement of exogenous insulin were significantly improved in all patients. Islet transplantation employing DCD can ameliorate severe hypoglycemic episodes, significantly improve HbA1c levels, sustain significant levels of C-peptide, and achieve insulin independence after multiple transplantations. Thus, DCD can be an important resource for islet transplantation if used under strict releasing criteria and in multiple transplantations, particularly in countries where heart-beating donors are not readily available.

Complement activation, endothelial dysfunction, insulin resistance and chronic heart failure

DEFF Research Database (Denmark)

Bjerre, M.; Kistorp, C.; Hansen, T.K.

2010-01-01

CRP), endothelial activation (soluble E-selectin, sEsel)), endothelial damage/dysfunction (von Willebrand factor, vWf) and insulin resistance (IR) and prognosis in CHF remains unknown. Design. We investigated the association(s) between plasma sMAC, hsCRP, sEsel, vWf and IR (assessed by homeostatic model assessment...

Antigen-Encoding Bone Marrow Terminates Islet-Directed Memory CD8+ T-Cell Responses to Alleviate Islet Transplant Rejection

DEFF Research Database (Denmark)

Coleman, Miranda; Jessup, Claire F.; Bridge, Jennifer A.

2016-01-01

in islet transplantation, and this will extend to application of personalized approaches using stem cell–derived replacement β-cells. New approaches are required to limit memory autoimmune attack of transplanted islets or replacement β-cells. Here, we show that transfer of bone marrow encoding cognate......Islet-specific memory T cells arise early in type 1 diabetes (T1D), persist for long periods, perpetuate disease, and are rapidly reactivated by islet transplantation. As memory T cells are poorly controlled by “conventional” therapies, memory T cell–mediated attack is a substantial challenge......-cell responses, and this can alleviate destruction of antigen-expressing islets. This addresses a key challenge facing islet transplantation and, importantly, the clinical application of personalized β-cell replacement therapies using patient-derived stem cells....

Islets of Langerhans in the parakeet, Psittacula krameri.

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Gupta, Y K; Kumar, S

1980-01-01

The pancreatic gland of Psittacula krameri is divisible into 4 lobes i.e. dorsal, ventral, third and splenic. The endocrine part is composed of alpha 1-, alpha 2- and beta-cells. The islets are of 4 kinds viz., alpha islets (having alpha 1- and alpha 2-cells), beta islets (having beta- and alpha 1-cells), pure beta islets (consisting of beta-cells exclusively) and mixed islets (with beta-, alpha 1- and alpha 2-cells). The distribution of alpha islets is mostly restricted to the splenic and third lobes whereas the beta islets are found in all 4 lobes. Though the alpha islets are only few in the dorsal lobe, their size is best developed in the third and dorsal lobes. Sometimes beta and alpha islets are present in very close proximity but their cells never mingle. An interesting feature was the complete absence of alpha islets from the ventral lobe.A relative abundance of alpha 2- cells in this bird seems to be associated with its comparatively higher blood glucose level and frugivorous habit. Tinctorial reactions suggest that the insulin content of the endocrine pancreas is low. There were no seasonal changes in the islet tissue of P. krameri.

Pseudoislets as primary islet replacements for research: report on a symposium at King's College London, London UK.

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Persaud, Shanta J; Arden, Catherine; Bergsten, Peter; Bone, Adrian J; Brown, James; Dunmore, Simon; Harrison, Moira; Hauge-Evans, Astrid; Kelly, Catriona; King, Aileen; Maffucci, Tania; Marriott, Claire E; McClenaghan, Neville; Morgan, Noel G; Reers, Christina; Russell, Mark A; Turner, Mark D; Willoughby, Emma; Younis, Mustafa Y G; Zhi, Z L; Jones, Peter M

2010-01-01

Laboratory-based research aimed at understanding processes regulating insulin secretion and mechanisms underlying β-cell dysfunction and loss in diabetes often makes use of rodents, as these processes are in many respects similar between rats/mice and humans. Indeed, a rough calculation suggests that islets have been isolated from as many as 150,000 rodents to generate the data contained within papers published in 2009 and the first four months of 2010. Rodent use for islet isolation has been mitigated, to a certain extent, by the availability of a variety of insulin-secreting cell lines that are used by researchers world-wide. However, when maintained as monolayers the cell lines do not replicate the robust, sustained secretory responses of primary islets which limits their usefulness as islet surrogates. On the other hand, there have been several reports that configuration of MIN6 β-cells, derived from a mouse insulinoma, as three-dimensional cell clusters termed ‘pseudoislets’ largely recapitulates the function of primary islet β-cells. The Diabetes Research Group at King’s College London has been using the MIN6 pseudoislet model for over a decade and they hosted a symposium on “Pseudoislets as primary islet replacements for research”, which was funded by the UK National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs), in London on 15th and 16th April 2010. This small, focused meeting was conceived as an opportunity to consolidate information on experiences of working with pseudoislets between different UK labs, and to introduce the theory and practice of pseudoislet culture to laboratories working with islets and/or β-cell lines but who do not currently use pseudoislets. This short review summarizes the background to the development of the cell line-derived pseudoislet model, the key messages arising from the symposium and emerging themes for future pseudoislet research.

Clinical Allogeneic and Autologous Islet Cell Transplantation: Update

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Shinichi Matsumoto

2011-06-01

Full Text Available Islet cell transplantation is categorized as a β-cell replacement therapy for diabetic patients who lack the ability to secrete insulin. Allogeneic islet cell transplantation is for the treatment of type 1 diabetes, and autologous islet cell transplantation is for the prevention of surgical diabetes after a total pancreatectomy. The issues of allogeneic islet cell transplantation include poor efficacy of islet isolation, the need for multiple donor pancreata, difficulty maintaining insulin independence and undesirable side effects of immunosuppressive drugs. Those issues have been solved step by step and allogeneic islet cell transplantation is almost ready to be the standard therapy. The donor shortage will be the next issue and marginal and/or living donor islet cell transplantation might alleviate the issue. Xeno-islet cell transplantation, β-cell regeneration from human stem cells and gene induction of the naïve pancreas represent the next generation of β-cell replacement therapy. Autologous islet cell transplantation after total pancreatectomy for the treatment of chronic pancreatitis with severe abdominal pain is the standard therapy, even though only limited centers are able to perform this treatment. Remote center autologous islet cell transplantation is an attractive option for hospitals performing total pancreatectomies without the proper islet isolation facilities.

Sequential kidney/islet transplantation using prednisone-free immunosuppression.

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Kaufman, Dixon B; Baker, Marshall S; Chen, Xiaojuan; Leventhal, Joseph R; Stuart, Frank P

2002-08-01

Islet transplantation is becoming established as a treatment option for type I diabetes in select patients. Individuals with type I diabetes who have previously received a successful kidney allograft may be good candidates for islet transplantation. They have already assumed the risks of chronic immunosuppression, so the added procedural risk of a subsequent islet transplant would be minimal. Furthermore, because of the preimmunosuppressed state it is possible that islet-after-kidney transplantation may result in a more efficient early islet engraftment. Consequently, insulin independence might be achieved with significantly fewer islets than the approximately 8-10,000 islet equivalents/kg/b.w. currently required. A mass that usually demands two or more cadaveric donors. A case of successful islet-after-kidney transplantation is described using the steroid-free Edmonton immunosuppression protocol. Characteristics of the final islet product are: a) islet equivalents: 265,888 (4100 islet equivalents/kg/b.w.); b) islet purity: 75-80%; c) viability: >95% (trypan blue exclusion); and d) mean islet potency (static low-high glucose challenge): 4.16 +/- 1.91-fold increase. Post-transplant the patient's hypoglycemic episodes abated. Exogenous insulin requirements were eliminated at week 12 post-transplant as basal and Ensure (Abbott Laboratories, Abbott Park, IL, USA) oral glucose stimulated C-peptide levels peaked and stabilized. Twenty-four-hour continuous glucose monitoring confirmed moment-to-moment glycemic control, and periodic nonfasting finger stick glucose determinations over the next month confirmed glycemia was controlled. Hemoglobin A1c levels declined from a pretransplant level of 6.9% to 5.3%. Renal allograft function remained changed.

Prolongation of islet allograft survival

International Nuclear Information System (INIS)

Lacy, P.E.; Davie, J.M.; Finke, E.H.; Scharp, D.W.

1979-01-01

Pretreatment of donor rats with irradiation and silica followed by in vitro culture of the islets for 1 to 2 days prolonged survival of allografts across a minor histocompatibility barrier if hand-picked, clean islets were used for transplantation. Pretreatment of donor rats with irradiation and silica in conjunction with a single injection of antilymphocyte serum (ALS) into the recipient produced a prolongation of survival of hand-picked islets transplanted across a major histocompatibility barrier

Risk factors for islet loss during culture prior to transplantation.

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Kin, Tatsuya; Senior, Peter; O'Gorman, Doug; Richer, Brad; Salam, Abdul; Shapiro, Andrew Mark James

2008-11-01

Culturing islets can add great flexibility to a clinical islet transplant program. However, a reduction in the islet mass has been frequently observed during culture and its degree varies. The aim of this study was to identify the risk factors associated with a significant islet loss during culture. One-hundred and four islet preparations cultured in an attempt to use for transplantation constituted this study. After culture for 20 h (median), islet yield significantly decreased from 363 309 +/- 12 647 to 313 035 +/- 10 862 islet equivalent yield (IE) (mean +/- SE), accompanied by a reduction in packed tissue volume from 3.9 +/- 0.1 to 3.0 +/- 0.1 ml and islet index (IE/islet particle count) from 1.20 +/- 0.04 to 1.05 +/- 0.04. Culture did not markedly alter islet purity or percent of trapped islet. Morphology score and viability were significantly improved after culture. Of 104 islet preparations, 37 suffered a substantial islet loss (> 20%) over culture. Factors significantly associated with risk of islet loss identified by univariate analysis were longer cold ischemia time, two-layer method (TLM) preservation, lower islet purity, and higher islet index. Multivariate analysis revealed that independent predictors of islet loss were higher islet index and the use of TLM. This study provides novel information on the link between donor- isolation factors and islet loss during culture.

Islet transplantation: the quest for an ideal source

International Nuclear Information System (INIS)

Younes, Nidal A.; Nothias, J.; Garfinkel, Marc R.

2008-01-01

The progress of islet transplantation as a new therapy for patients with diabetes mellitus depends directly upon the development of efficient and practical immunoisolation methods for the supply of sufficient quantities of islet cells. Without these methods, large scale clinical application of this therapy would be impossible. Two eras of advances can be identified in the development of islet transplantation. The first was an era of experimental animal and human research that centered on islet isolation procedures and transplantation in different species as evidence that transplanted islets have the capability to reverse diabetes. The second was the era of Edmonton protocol, when the focus became the standardization of isolation procedures and introduction of new immunosuppressive drugs to maintain human allograft transplantation. The quest for an alternative source for islets (xenographs, stem cells and cell cultures) to overcome the shortage of human islets was an important issue during these eras. This paper reviews the history of islet transplantation and the current procedures in human allotransplantation, as well as different types of immunoisolation methods. It explores novel approaches to enhancing transplantation site vascularity and islet cell function, whereby future immunoisolation technology could offer additional therapeutic advantages to human islet allotransplantation. (author)

Ketosis Onset Type 2 Diabetes Had Better Islet β-Cell Function and More Serious Insulin Resistance

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Lu, Hongyun; Hu, Fang; Zeng, Yingjuan; Zou, Lingling; Luo, Shunkui; Sun, Ying; Liu, Hong; Sun, Liao

2014-01-01

Diabetic ketosis had been identified as a characteristic of type 1 diabetes mellitus (T1DM), but now emerging evidence has identified that they were diagnosed as T2DM after long time follow up. This case control study was aimed at comparing the clinical characteristic, β-cell function, and insulin resistance of ketosis and nonketotic onset T2DM and providing evidence for treatment selection. 140 cases of newly diagnosed T2DM patients were divided into ketosis (62 cases) and nonketotic onset group (78 cases). After correction of hyperglycemia and ketosis with insulin therapy, plasma C-peptide concentrations were measured at 0, 0.5, 1, 2, and 3 hours after 75 g glucose oral administration. Area under the curve (AUC) of C-peptide was calculated. Homoeostasis model assessment was used to estimate basal β-cell function (HOMA-β) and insulin resistance (HOMA-IR). Our results showed that ketosis onset group had higher prevalence of nonalcoholic fatty liver disease (NAFLD) than nonketotic group (P = 0.04). Ketosis onset group had increased plasma C-peptide levels at 0 h, 0.5 h, and 3 h and higher AUC0–0.5, AUC0–1, AUC0–3 (P ketosis onset T2DM had better islet β-cell function and more serious insulin resistance than nonketotic onset T2DM. PMID:24829925

3-D Imaging Reveals Participation of Donor Islet Schwann Cells and Pericytes in Islet Transplantation and Graft Neurovascular Regeneration.

Science.gov (United States)

Juang, Jyuhn-Huarng; Kuo, Chien-Hung; Peng, Shih-Jung; Tang, Shiue-Cheng

2015-02-01

The primary cells that participate in islet transplantation are the endocrine cells. However, in the islet microenvironment, the endocrine cells are closely associated with the neurovascular tissues consisting of the Schwann cells and pericytes, which form sheaths/barriers at the islet exterior and interior borders. The two cell types have shown their plasticity in islet injury, but their roles in transplantation remain unclear. In this research, we applied 3-dimensional neurovascular histology with cell tracing to reveal the participation of Schwann cells and pericytes in mouse islet transplantation. Longitudinal studies of the grafts under the kidney capsule identify that the donor Schwann cells and pericytes re-associate with the engrafted islets at the peri-graft and perivascular domains, respectively, indicating their adaptability in transplantation. Based on the morphological proximity and cellular reactivity, we propose that the new islet microenvironment should include the peri-graft Schwann cell sheath and perivascular pericytes as an integral part of the new tissue.

3-D Imaging Reveals Participation of Donor Islet Schwann Cells and Pericytes in Islet Transplantation and Graft Neurovascular Regeneration

Directory of Open Access Journals (Sweden)

Jyuhn-Huarng Juang

2015-02-01

Full Text Available The primary cells that participate in islet transplantation are the endocrine cells. However, in the islet microenvironment, the endocrine cells are closely associated with the neurovascular tissues consisting of the Schwann cells and pericytes, which form sheaths/barriers at the islet exterior and interior borders. The two cell types have shown their plasticity in islet injury, but their roles in transplantation remain unclear. In this research, we applied 3-dimensional neurovascular histology with cell tracing to reveal the participation of Schwann cells and pericytes in mouse islet transplantation. Longitudinal studies of the grafts under the kidney capsule identify that the donor Schwann cells and pericytes re-associate with the engrafted islets at the peri-graft and perivascular domains, respectively, indicating their adaptability in transplantation. Based on the morphological proximity and cellular reactivity, we propose that the new islet microenvironment should include the peri-graft Schwann cell sheath and perivascular pericytes as an integral part of the new tissue.

Lipotoxicity, β cell dysfunction, and gestational diabetes.

Science.gov (United States)

Nolan, Christopher J

2014-04-01

Gestational diabetes (GDM) is caused by failure of islet β cells to meet the increased insulin requirements of pregnancy. Recently, Prentice et al. (2014) discovered a 7-fold elevation of the furan fatty acid metabolite 3-carboxy-4-methyl-5-propyl-2-furanopropanoic acid (CMPF) in plasma of women with GDM and showed that CMPF directly induces β cell dysfunction. Copyright © 2014 Elsevier Inc. All rights reserved.

A Practical Guide to Rodent Islet Isolation and Assessment

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Carter Jeffrey D

2009-12-01

Full Text Available Abstract Pancreatic islets of Langerhans secrete hormones that are vital to the regulation of blood glucose and are, therefore, a key focus of diabetes research. Purifying viable and functional islets from the pancreas for study is an intricate process. This review highlights the key elements involved with mouse and rat islet isolation, including choices of collagenase, the collagenase digestion process, purification of islets using a density gradient, and islet culture conditions. In addition, this paper reviews commonly used techniques for assessing islet viability and function, including visual assessment, fluorescent markers of cell death, glucose-stimulated insulin secretion, and intracellular calcium measurements. A detailed protocol is also included that describes a common method for rodent islet isolation that our laboratory uses to obtain viable and functional mouse islets for in vitro study of islet function, beta-cell physiology, and in vivo rodent islet transplantation. The purpose of this review is to serve as a resource and foundation for successfully procuring and purifying high-quality islets for research purposes.

Delayed revascularization of islets after transplantation by IL-6 blockade in pig to non-human primate islet xenotransplantation model.

Science.gov (United States)

Min, Byoung-Hoon; Shin, Jun-Seop; Kim, Jong-Min; Kang, Seong-Jun; Kim, Hyun-Je; Yoon, Il-Hee; Park, Su-Kyoung; Choi, Ji-Won; Lee, Min-Suk; Park, Chung-Gyu

2018-01-01

Pancreatic islet transplantation is currently proven as a promising treatment for type 1 diabetes patients with labile glycemic control and severe hypoglycemia unawareness. Upon islet transplantation, revascularization is essential for proper functioning of the transplanted islets. As IL-6 is important for endothelial cell survival and systemic inflammation related to xenograft, the effect of IL-6 receptor antagonist, tocilizumab, on revascularization of the transplanted islets was examined in pig to non-human primate islet xenotransplantation model. Also, the endothelial cell origin in a new vessel of the transplanted pig islets was determined. Pig islets were isolated from designated pathogen-free (DPF) SNU miniature pigs and transplanted via portal vein into five streptozotocin-induced diabetic monkeys. One group (n = 2, basal group) was treated with anti-thymoglobulin (ATG), anti-CD40 antibody (2C10R4), sirolimus, and tacrolimus, and the other group was additionally given tocilizumab on top of basal immunosuppression (n = 3, Tocilizumab group). To confirm IL-6 blocking effect, C-reactive protein (CRP) levels and serum IL-6 concentration were measured. Scheduled biopsy of the margin of the posterior segment right lobe inferior of the liver was performed at 3 weeks after transplantation to assess the degree of revascularization of the transplanted islets. Immunohistochemical staining using anti-insulin, anti-CD31 antibodies, and lectin IB4 was conducted to find the origin of endothelial cells in the islet graft. CRP significantly increased at 1~2 days after transplantation in Basal group, but not in Tocilizumab group, and higher serum IL-6 concentration was measured in latter group, showing the biological potency of tocilizumab. In Basal group, well-developed endothelial cells were observed on the peri- and intraislet area, whereas the number of CD31 + cells in the intraislet space was significantly reduced in Tocilizumab group. Finally, new endothelial

Metabolomics applied to the pancreatic islet.

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Gooding, Jessica R; Jensen, Mette V; Newgard, Christopher B

2016-01-01

Metabolomics, the characterization of the set of small molecules in a biological system, is advancing research in multiple areas of islet biology. Measuring a breadth of metabolites simultaneously provides a broad perspective on metabolic changes as the islets respond dynamically to metabolic fuels, hormones, or environmental stressors. As a result, metabolomics has the potential to provide new mechanistic insights into islet physiology and pathophysiology. Here we summarize advances in our understanding of islet physiology and the etiologies of type-1 and type-2 diabetes gained from metabolomics studies. Copyright © 2015 Elsevier Inc. All rights reserved.

Improving Islet Engraftment by Gene Therapy

Directory of Open Access Journals (Sweden)

Xiaojie Wang

2011-01-01

Full Text Available Islet cell transplantation is currently the only feasible long-term treatment option for patients with type 1 diabetes. However, the majority of transplanted islets experience damage and apoptosis during the isolation process, a blood-mediated inflammatory microenvironment in the portal vein upon islet infusion, hypoxia induced by the low oxygenated milieu, and poor-revascularization-mediated lack of nutrients, and impaired hormone modulation in the local transplanted site. Strategies using genetic modification methods through overexpression or silencing of those proteins involved in promoting new formation of blood vessels or inhibition of apoptosis may overcome these hurdles and improve islet engraftment outcomes.

Cyclic AMP in rat pancreatic islets

International Nuclear Information System (INIS)

Grill, V.; Borglund, E.; Cerasi, E.; Uppsala Univ.

1977-01-01

The incorporation of [ 3 H]adenine into cyclic AMP was studied in rat pancreatic islets under varying conditions of labeling. Prolonging the exposure to [ 3 H]adenine progressively augmented the islet cyclic [ 3 H]AMP level. Islets labeled for different periods of time and subsequently incubated (without adenine) in the presence of D-glucose or cholera toxin showed stimulations of intra-islet cyclic [ 3 H]AMP that were proportionate to the levels of radioactive nucleotide present under non-stimulatory conditions. Labeling the islets in a high glucose concentration (27.7 mM) did not modify the nucleotide responses to glucose or cholera toxin. The specific activity of cyclic [ 3 H]AMP, determined by simultaneous assay of cyclic [ 3 H]AMP and total cyclic AMP, was not influenced by glucose or cholera toxin. Glucose had no effect on the specific activity of labeled ATP

Novel immunological strategies for islet transplantation.

Science.gov (United States)

Tezza, Sara; Ben Nasr, Moufida; Vergani, Andrea; Valderrama Vasquez, Alessandro; Maestroni, Anna; Abdi, Reza; Secchi, Antonio; Fiorina, Paolo

2015-08-01

Islet transplantation has been demonstrated to improve glycometabolic control, to reduce hypoglycemic episodes and to halt the progression of diabetic complications. However, the exhaustion of islet function and the side effects related to chronic immunosuppression limit the spread of this technique. Consequently, new immunoregulatory protocols have been developed, with the aim to avoid the use of a life-time immunosuppression. Several approaches have been tested in preclinical models, and some are now under clinical evaluation. The development of new small molecules and new monoclonal or polyclonal antibodies is continuous and raises the possibility of targeting new costimulatory pathways or depleting particular cell types. The use of stem cells and regulatory T cells is underway to take advantage of their immunological properties and to induce tolerance. Xenograft islet transplantation, although having severe problems in terms of immunological compatibility, could theoretically provide an unlimited source of donors; using pigs carrying human immune antigens has showed indeed promising results. A completely different approach, the use of encapsulated islets, has been developed; synthetic structures are used to hide islet alloantigen from the immune system, thus preserving islet endocrine function. Once one of these strategies is demonstrated safe and effective, it will be possible to establish clinical islet transplantation as a treatment for patients with type 1 diabetes long before the onset of diabetic-related complications. Copyright © 2014 Elsevier Ltd. All rights reserved.

Excessive islet NO generation in type 2 diabetic GK rats coincides with abnormal hormone secretion and is counteracted by GLP-1.

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Albert Salehi

Full Text Available BACKGROUND: A distinctive feature of type 2 diabetes is inability of insulin-secreting beta-cells to properly respond to elevated glucose eventually leading to beta-cell failure. We have hypothesized that an abnormally increased NO production in the pancreatic islets might be an important factor in the pathogenesis of beta-cell dysfunction. PRINCIPAL FINDINGS: We show now that islets of type 2 spontaneous diabetes in GK rats display excessive NO generation associated with abnormal iNOS expression in insulin and glucagon cells, increased ncNOS activity, impaired glucose-stimulated insulin release, glucagon hypersecretion, and impaired glucose-induced glucagon suppression. Pharmacological blockade of islet NO production by the NOS inhibitor N(G-nitro-L-arginine methyl ester (L-NAME greatly improved hormone secretion from GK islets suggesting islet NOS activity being an important target to inactivate for amelioration of islet cell function. The incretin hormone GLP-1, which is used in clinical practice suppressed iNOS and ncNOS expression and activity with almost full restoration of insulin release and partial restoration of glucagon release. GLP-1 suppression of iNOS expression was reversed by PKA inhibition but unaffected by the proteasome inhibitor MG132. Injection of glucose plus GLP-1 in the diabetic rats showed that GLP-1 amplified the insulin response but induced a transient increase and then a poor depression of glucagon. CONCLUSION: The results suggest that abnormally increased NO production within islet cells is a significant player in the pathogenesis of type 2 diabetes being counteracted by GLP-1 through PKA-dependent, nonproteasomal mechanisms.

3-D Imaging Reveals Participation of Donor Islet Schwann Cells and Pericytes in Islet Transplantation and Graft Neurovascular Regeneration

OpenAIRE

Juang, Jyuhn-Huarng; Kuo, Chien-Hung; Peng, Shih-Jung; Tang, Shiue-Cheng

2015-01-01

The primary cells that participate in islet transplantation are the endocrine cells. However, in the islet microenvironment, the endocrine cells are closely associated with the neurovascular tissues consisting of the Schwann cells and pericytes, which form sheaths/barriers at the islet exterior and interior borders. The two cell types have shown their plasticity in islet injury, but their roles in transplantation remain unclear. In this research, we applied 3-dimensional neurovascular histolo...

Renin-angiotensin system blockers protect pancreatic islets against diet-induced obesity and insulin resistance in mice.

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Eliete Dalla Corte Frantz

Full Text Available BACKGROUND: The associations between obesity, hypertension and diabetes are well established, and the renin-angiotensin system (RAS may provide a link among them. The effect of RAS inhibition on type 2 diabetes is still unclear; however, RAS seems to play an important role in the regulation of the pancreas and glucose intolerance of mice fed high-fat (HF diet. METHODS: C57BL/6 mice fed a HF diet (8 weeks were treated with aliskiren (50 mg/kg/day, enalapril (30 mg/kg/day or losartan (10 mg/kg/day for 6 weeks, and the protective effects were extensively compared among groups by morphometry, stereological tools, immunostaining, Western blotting and hormonal analysis. RESULTS: All RAS inhibitors significantly attenuated the increased blood pressure in mice fed a HF diet. Treatment with enalapril, but not aliskiren or losartan, significantly attenuated body mass (BM gain, glucose intolerance and insulin resistance, improved the alpha and beta cell mass and prevented the reduction of plasma adiponectin. Furthermore, enalapril treatment improved the protein expression of the pancreatic islet Pdx1, GLUT2, ACE2 and Mas receptors. Losartan treatment showed the greatest AT2R expression. CONCLUSION: Our findings indicate that ACE inhibition with enalapril attenuated several of the deleterious effects of the HF diet. In summary, enalapril appears to be responsible for the normalization of islet morphology and function, of alpha and beta cell mass and of Pdx1 and GLUT2 expression. These protective effects of enalapril were attributed, primarily, to the reduction in body mass gain and food intake and the enhancement of the ACE2/Ang (1-7 /Mas receptor axis and adiponectin levels.

Immunohistochemical detection of vimentin in pancreatic islet β- and α-cells of macrosomic infants of diabetic and nondiabetic mothers.

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Krivova, Yuliya S; Proshchina, Alexandra E; Barabanov, Valeriy M; Barinova, Irina V; Saveliev, Sergey V

2018-02-01

Expression of the intermediate filament protein vimentin has been recently observed in the pancreatic islet β- and α-cells of humans with type 2 diabetes mellitus. It was suggested that the presence of vimentin in endocrine cells may indicate islet tissue renewal, or potentially represent the dedifferentiation of endocrine cells, which could contribute to the onset of type 2 diabetes or islet cell dysfunction. To analyze the expression of vimentin in pancreatic β- and α-cells of macrosomic infants of diabetic and nondiabetic mothers. Pancreatic samples of five macrosomic infants (gestational age 34-40weeks) from three diabetic and two nondiabetic mothers were compared to six control infants (32-40weeks, weight appropriate for gestational age) from normoglycemic mothers. Pancreatic autopsy samples were examined by double immunofluorescent labeling with antibodies against vimentin and either insulin or glucagon. Alterations in the endocrine pancreas were measured using morphometric methods, then data were statistically analyzed. In the pancreatic islets of macrosomic infants from diabetic and nondiabetic mothers, we observed vimentin-positive cells, some of which simultaneously contained insulin or glucagon. We also quantitatively showed that the presence of such cells was associated with hypertrophy and hyperplasia of the islets, and with an increase in β- and α-cell density. We speculate that the appearance of vimentin-positive islet cells may reflect induction of differentiation in response to the increased insulin demand, and vimentin may serve as an early marker of endocrine pancreas disorders. Copyright © 2017 Elsevier B.V. All rights reserved.

Adipose tissue mitochondrial dysfunction triggers a lipodystrophic syndrome with insulin resistance, hepatosteatosis, and cardiovascular complications.

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Vernochet, Cecile; Damilano, Federico; Mourier, Arnaud; Bezy, Olivier; Mori, Marcelo A; Smyth, Graham; Rosenzweig, Anthony; Larsson, Nils-Göran; Kahn, C Ronald

2014-10-01

Mitochondrial dysfunction in adipose tissue occurs in obesity, type 2 diabetes, and some forms of lipodystrophy, but whether this dysfunction contributes to or is the result of these disorders is unknown. To investigate the physiological consequences of severe mitochondrial impairment in adipose tissue, we generated mice deficient in mitochondrial transcription factor A (TFAM) in adipocytes by using mice carrying adiponectin-Cre and TFAM floxed alleles. These adiponectin TFAM-knockout (adipo-TFAM-KO) mice had a 75-81% reduction in TFAM in the subcutaneous and intra-abdominal white adipose tissue (WAT) and interscapular brown adipose tissue (BAT), causing decreased expression and enzymatic activity of proteins in complexes I, III, and IV of the electron transport chain (ETC). This mitochondrial dysfunction led to adipocyte death and inflammation in WAT and a whitening of BAT. As a result, adipo-TFAM-KO mice were resistant to weight gain, but exhibited insulin resistance on both normal chow and high-fat diets. These lipodystrophic mice also developed hypertension, cardiac hypertrophy, and cardiac dysfunction. Thus, isolated mitochondrial dysfunction in adipose tissue can lead a syndrome of lipodystrophy with metabolic syndrome and cardiovascular complications. © FASEB.

Treatment of diabetic rats with encapsulated islets.

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Sweet, Ian R; Yanay, Ofer; Waldron, Lanaya; Gilbert, Merle; Fuller, Jessica M; Tupling, Terry; Lernmark, Ake; Osborne, William R A

2008-12-01

Immunoprotection of islets using bioisolator systems permits introduction of allogeneic cells to diabetic patients without the need for immunosuppression. Using TheraCyte immunoisolation devices, we investigated two rat models of type 1 diabetes mellitus (T1DM), BB rats and rats made diabetic by streptozotocin (STZ) treatment. We chose to implant islets after the onset of diabetes to mimic the probable treatment of children with T1DM as they are usually diagnosed after disease onset. We encapsulated 1000 rat islets and implanted them subcutaneously (SQ) into diabetic biobreeding (BB) rats and STZ-induced diabetic rats, defined as two or more consecutive days of blood glucose>350 mg/dl. Rats were monitored for weight and blood glucose. Untreated BB rats rapidly lost weight and were euthanized at >20% weight loss that occurred between 4 and 10 days from implantation. For period of 30-40 days following islet implantation weights of treated rats remained steady or increased. Rapid weight loss occurred after surgical removal of devices that contained insulin positive islets. STZ-treated rats that received encapsulated islets showed steady weight gain for up to 130 days, whereas untreated control rats showed steady weight loss that achieved >20% at around 55 days. Although islet implants did not normalize blood glucose, treated rats were apparently healthy and groomed normally. Autologous or allogeneic islets were equally effective in providing treatment. TheraCyte devices can sustain islets, protect allogeneic cells from immune attack and provide treatment for diabetic-mediated weight loss in both BB rats and STZ-induced diabetic rats.

Tacrolimus inhibits the revascularization of isolated pancreatic islets.

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Ryuichi Nishimura

Full Text Available AIMS: Immunosuppressive drugs could be crucial factors for a poor outcome after islet allotransplantation. Unlike rapamycin, the effects of tacrolimus, the current standard immunosuppressant used in islet transplantation, on graft revascularization remain unclear. We examined the effects of tacrolimus on islet revascularization using a highly sensitive imaging system, and analyzed the gene expression in transplanted islets by introducing laser microdissection techniques. METHODS: Islets isolated from C57BL/6-Tg (CAG-EGFP mice were transplanted into the nonmetallic dorsal skinfold chamber on the recipients. Balb/c athymic mice were used as recipients and were divided into two groups: including a control group (n = 9 and tacrolimus-treated group (n = 7. The changes in the newly-formed vessels surrounding the islet grafts were imaged and semi-quantified using multi-photon laser-scanning microscopy and a Volocity system. Gene expression in transplanted islets was analyzed by the BioMark dynamic system. RESULTS: The revascularization process was completed within 14 days after pancreatic islet transplantation at subcutaneous sites. The newly-formed vascular volume surrounding the transplanted islets in the tacrolimus-treated group was significantly less than that in the control group (p<0.05. Although the expression of Vegfa (p<0.05 and Ccnd1 (p<0.05 was significantly upregulated in the tacrolimus-treated group compared with that of the control group, no differences were observed between the groups in terms of other types of gene expression. CONCLUSIONS: The present study demonstrates that tacrolimus inhibits the revascularization of isolated pancreatic islets without affecting the characteristics of the transplanted grafts. Further refinements of this immunosuppressive regimen, especially regarding the revascularization of islet grafts, could improve the outcome of islet allotransplantation.

Periodontitis aggravated pancreatic β-cell dysfunction in diabetic mice through interleukin-12 regulation on Klotho.

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Liu, Yihua; Zhang, Qiuli

2016-05-01

Recent studies have shown that periodontitis can contribute to adipose tissue inflammation and subsequent systemic insulin resistance in the obese rat model. However, the related inflammatory mechanism is not yet clear. The present study aims to investigate the effects of periodontitis on the function of pancreatic β-cells with pro-inflammatory cytokines-related immune mechanism in a mouse model. C57BL/6-db/db and inbred C57BL/6 mice were chosen here to establish a mouse model with periodontitis, which was induced by ligatures for 8 weeks. Glucose-stimulated insulin secretion was introduced to evaluate the function of pancreatic islets and β-cells. Serum levels of pro-inflammatory cytokines and Klotho were also measured, and the correlation between immunostimulation and Klotho level was deeply investigated in vitro. Pancreatic β-cell failure, with insulin resistance, was observed in db/db mice, while periodontitis could aggravate β-cell dysfunction-related features. Serum levels of interleukin (IL)-12 and Klotho showed a negatively synergistic change, whereas the expression of Klotho was also inhibited under IL-12 treatment in MIN6 β-cells or isolated islets. Furthermore, IL-12-induced immune stimulation and also decreased insulin secretion were proven to be reversed by Klotho overexpression. Periodontitis aggravated pancreatic β-cell failure in diabetic mice. Further in vitro studies showed IL-12 regulation on Klotho, while Klotho also acted as an inhibitor on IL-12, indicating the potential of Klotho for preserving pancreatic β-cell function in diabetes.

Selective Osmotic Shock (SOS)-Based Islet Isolation for Microencapsulation.

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Enck, Kevin; McQuilling, John Patrick; Orlando, Giuseppe; Tamburrini, Riccardo; Sivanandane, Sittadjody; Opara, Emmanuel C

2017-01-01

Islet transplantation (IT) has recently been shown to be a promising alternative to pancreas transplantation for reversing diabetes. IT requires the isolation of the islets from the pancreas, and these islets can be used to fabricate a bio-artificial pancreas. Enzymatic digestion is the current gold standard procedure for islet isolation but has lingering concerns. One such concern is that it has been shown to damage the islets due to nonselective tissue digestion. This chapter provides a detailed description of a nonenzymatic method that we are exploring in our lab as an alternative to current enzymatic digestion procedures for islet isolation from human and nonhuman pancreatic tissues. This method is based on selective destruction and protection of specific cell types and has been shown to leave the extracellular matrix (ECM) of islets intact, which may thus enhance islet viability and functionality. We also show that these SOS-isolated islets can be microencapsulated for transplantation.

Evolution of Islet Transplantation for the Last 30 Years.

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Farney, Alan C; Sutherland, David E R; Opara, Emmanuel C

2016-01-01

In this article, we will review the changes that have occurred in islet transplantation at the birth of Pancreas 30 years ago. The first attempts at β-cell replacement in humans, pancreas and islet transplantation, were performed in the 1960s and 1970s. Although pancreas transplantation has been an accepted treatment for severe labile diabetes predating the emergence of the journal, allogeneic islet transplantation remains experimental. Current investigations within islet transplantation focus to improve islet function after transplantation. Improving islet viability during isolation, exploring ways to increase engraftment, and protection from the host immune system are some of the goals of these investigative efforts. The major barriers to clinical islet transplantation are shortage of human pancreas, the need for immunosuppression, and the inadequacy of the islet isolation process. It is generally accepted that islet encapsulation is an immunoisolation tool with good potential to address the first 2 of those barriers. We have therefore devoted a major part of this review to the critical factors needed to make it a clinical reality. With improved islet isolation techniques and determination of the best site of engraftment as well as improved encapsulation techniques, we hope that islet transplantation could someday achieve routine clinical use.

Redox-Dependent Inflammation in Islet Transplantation Rejection

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Jessie M. Barra

2018-04-01

Full Text Available Type 1 diabetes is an autoimmune disease that results in the progressive destruction of insulin-producing pancreatic β-cells inside the islets of Langerhans. The loss of this vital population leaves patients with a lifelong dependency on exogenous insulin and puts them at risk for life-threatening complications. One method being investigated to help restore insulin independence in these patients is islet cell transplantation. However, challenges associated with transplant rejection and islet viability have prevented long-term β-cell function. Redox signaling and the production of reactive oxygen species (ROS by recipient immune cells and transplanted islets themselves are key players in graft rejection. Therefore, dissipation of ROS generation is a viable intervention that can protect transplanted islets from immune-mediated destruction. Here, we will discuss the newly appreciated role of redox signaling and ROS synthesis during graft rejection as well as new strategies being tested for their efficacy in redox modulation during islet cell transplantation.

Redox-Dependent Inflammation in Islet Transplantation Rejection

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Barra, Jessie M.; Tse, Hubert M.

2018-01-01

Type 1 diabetes is an autoimmune disease that results in the progressive destruction of insulin-producing pancreatic β-cells inside the islets of Langerhans. The loss of this vital population leaves patients with a lifelong dependency on exogenous insulin and puts them at risk for life-threatening complications. One method being investigated to help restore insulin independence in these patients is islet cell transplantation. However, challenges associated with transplant rejection and islet viability have prevented long-term β-cell function. Redox signaling and the production of reactive oxygen species (ROS) by recipient immune cells and transplanted islets themselves are key players in graft rejection. Therefore, dissipation of ROS generation is a viable intervention that can protect transplanted islets from immune-mediated destruction. Here, we will discuss the newly appreciated role of redox signaling and ROS synthesis during graft rejection as well as new strategies being tested for their efficacy in redox modulation during islet cell transplantation. PMID:29740396

Pancreatic Islet Cell Transplantation

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Warnock, Garth L.; Rajotte, Ray V.

1992-01-01

Transplantation of insulin-producing tissue offers a physiologic approach to restoration of glycemic control. Whereas transplantation of vascularized pancreatic grafts has recently achieved encouraging results, pancreatic islet cell transplantation holds the promise of low morbidity and reduced requirements for agressive immunosuppression for recipients. Islet cell transplantation was recently demonstrated to induce euglycemia with insulin independence. Imagesp1656-a PMID:21221366

Human pancreatic islet transplantation: an update and description of the establishment of a pancreatic islet isolation laboratory.

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Rheinheimer, Jakeline; Bauer, Andrea C; Silveiro, Sandra P; Estivalet, Aline A F; Bouças, Ana P; Rosa, Annelise R; Souza, Bianca M de; Oliveira, Fernanda S de; Cruz, Lavínia A; Brondani, Letícia A; Azevedo, Mirela J; Lemos, Natália E; Carlessi, Rodrigo; Assmann, Taís S; Gross, Jorge L; Leitão, Cristiane B; Crispim, Daisy

2015-04-01

Type 1 diabetes mellitus (T1DM) is associated with chronic complications that lead to high morbidity and mortality rates in young adults of productive age. Intensive insulin therapy has been able to reduce the likelihood of the development of chronic diabetes complications. However, this treatment is still associated with an increased incidence of hypoglycemia. In patients with "brittle T1DM", who have severe hypoglycemia without adrenergic symptoms (hypoglycemia unawareness), islet transplantation may be a therapeutic option to restore both insulin secretion and hypoglycemic perception. The Edmonton group demonstrated that most patients who received islet infusions from more than one donor and were treated with steroid-free immunosuppressive drugs displayed a considerable decline in the initial insulin independence rates at eight years following the transplantation, but showed permanent C-peptide secretion, which facilitated glycemic control and protected patients against hypoglycemic episodes. Recently, data published by the Collaborative Islet Transplant Registry (CITR) has revealed that approximately 50% of the patients who undergo islet transplantation are insulin independent after a 3-year follow-up. Therefore, islet transplantation is able to successfully decrease plasma glucose and HbA1c levels, the occurrence of severe hypoglycemia, and improve patient quality of life. The goal of this paper was to review the human islet isolation and transplantation processes, and to describe the establishment of a human islet isolation laboratory at the Endocrine Division of the Hospital de Clínicas de Porto Alegre - Rio Grande do Sul, Brazil.

Preimplantation of an immunoprotective device can lower the curative dose of islets to that of free islet transplantation: studies in a rodent model.

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Sörenby, Anne K; Kumagai-Braesch, Makiko; Sharma, Amit; Hultenby, Kjell R; Wernerson, Annika M; Tibell, Annika B

2008-07-27

Islet graft survival inside macroencapsulation devices is suboptimal. We hypothesized that induction of neovascularization by preimplantation of devices would improve the physiological conditions, thereby lowering the number of islets required for cure. Several rat islets were transplanted to TheraCyte immunoprotective devices implanted subcutaneously in diabetic athymic mice. Cure rates in the groups with preimplanted devices were significantly better than in those with freshly implanted devices (375 islets: 8/8 vs. 1/6, P=0.003; 125 islets: 6/6 vs. 0/7, P=0.001). Morphometric evaluations of the 125 islet groups showed higher fractional and absolute volumes of endocrine tissue in the group with preimplanted devices (P<0.001 and P=0.035, respectively). In the following dose titration study, using preimplanted devices, as low as 50 islets cured diabetic mice (100% cure, n=6). We conclude that preimplantation significantly lowers the curative dose of macroencapsulated islets to levels resembling those of free islets transplanted under the renal capsule.

Pancreatic islet transplantation. Experimental and clinical aspects

DEFF Research Database (Denmark)

Yderstræde, Knud Bonnet

1987-01-01

interest has been shown in transplantation of isolated islets either directly, introduced intraportally, intramuscularly, inter alia, or encapsulated in artificial devices providing an immuno-isolation. Clinical application has revealed promising results concerning the immunological aspects. However......, quantitative assessment points to a difficulty in achieving satisfactory amounts of islets to attain normoglycaemia. Work with fetal pancreata has shown these to possess a growth potential in vitro thus, possibly, aiding the quantification of islets in transplantation models. In the field of pancreatic islet...... transplantation, future models include microencapsulation and hybrid artificial devices, both of which provide immuno-isolation - thus the ability of allo- as well as xeno-transplantation. The obvious advantage of immuno-isolated islet transplant, as opposed to segmentally engrafted pancreas, is stressed...

Islet Microencapsulation: Strategies and Clinical Status in Diabetes.

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Omami, Mustafa; McGarrigle, James J; Reedy, Mick; Isa, Douglas; Ghani, Sofia; Marchese, Enza; Bochenek, Matthew A; Longi, Maha; Xing, Yuan; Joshi, Ira; Wang, Yong; Oberholzer, José

2017-07-01

Type 1 diabetes mellitus (T1DM) is an autoimmune disease that results from the destruction of insulin-producing pancreatic β cells in the islets of Langerhans. Islet cell transplantation has become a successful therapy for specific patients with T1DM with hypoglycemic unawareness. The reversal of T1DM by islet transplantation is now performed at many major medical facilities throughout the world. However, many challenges must still be overcome in order to achieve continuous, long-term successful transplant outcomes. Two major obstacles to this therapy are a lack of islet cells for transplantation and the need for life-long immunosuppressive treatment. Microencapsulation is seen as a technology that can overcome both these limitations of islet cell transplantation. This review depicts the present state of microencapsulated islet transplantation. Microencapsulation can play a significant role in overcoming the need for immunosuppression and lack of donor islet cells. This review focuses on microencapsulation and the clinical status of the technology in combating T1DM.

Islet transplantation in multicenter networks: the GRAGIL example

International Nuclear Information System (INIS)

Thierry Berney; Pierre-Yves Benhamou; Laurence Kessler; Philippe Morel

2006-01-01

Purpose of review: The enthusiasm generated by the results of the Edmonton protocol of islet transplantation is inciting a great number of institutions to start such programs. However, the procedure of islet isolation and purification is costly, complex and technically challenging. In order to share costs and to avoid facing the steep learning curve of the procedure, many centers interested in islet transplantation have looked into collaborating with experienced groups serving as core islet isolation facilities. Recent findings: The proof of principle that remote islet processing and shipment could be successfully implemented with obtainng the Portland/Minneapolis, Huddinge/Giessen and Houston/Miami partnerships. Moreover, in order to increase both the donor pool and the number of patients gaining access to islet transplantation, multicenter networks, such as the Swiss-French GRAGIL consortium and the 4-country Nordic Network in Scandinavia have been built. The GRAGIL group has been fully operational since 1999, allowing the transplantation of 27 islet preparations processed in Geneva, Switzerland into 20 recipients in France over the course of 4.5 years. Organizational issues in the design of such networks are discussed based on the example of the GRAGIL experience. Summary: The feasibility and the efficiency of islet transplantation in multicenter networks have been demonstrated. This strategy allows to increase the donor pool and the accessibility to islet transplantation in an extended population area. (authors)

Current status and outlook of pancreatic islets transplantation research

International Nuclear Information System (INIS)

Wang Wei; Ye Bin

2006-01-01

Diabetes is a common disease, severely harmful to the human's health and life quality. The pancreatic islets transplantation can correct the patient's hyperglycemia, stop or even reverse the progress of the complication and thus decrease the mortality of diabetic patients. It is the most safe and efficient therapy for diabetes. Since the Edmonton Protocol got success in pancreatic islet transplantation in 2000, it has been more and more interested because of its great clinical curative effect. Research strategy of islet transplantation is now focussed on increasing the acquired islets with normal viability, selecting the best transplantation pathway, and improving the immunosuppression protocol. The shortage of human pancreatic donor is an ever unsolved problem in clinical application. The potential resolutions may include acquisition from xenogenic-islets; islets originated from stem cells, and islets from the living-donor human pancreas. The islets transplantation will open a new application field for interventional radiology. (authors)

IKKβ inhibition prevents fat-induced beta cell dysfunction in vitro and in vivo in rodents.

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Ivovic, Aleksandar; Oprescu, Andrei I; Koulajian, Khajag; Mori, Yusaku; Eversley, Judith A; Zhang, Liling; Nino-Fong, Rodolfo; Lewis, Gary F; Donath, Marc Y; Karin, Michael; Wheeler, Michael B; Ehses, Jan; Volchuk, Allen; Chan, Catherine B; Giacca, Adria

2017-10-01

We have previously shown that oxidative stress plays a causal role in beta cell dysfunction induced by fat. Here, we address whether the proinflammatory kinase inhibitor of (nuclear factor) κB kinase β (IKKβ), which is activated by oxidative stress, is also implicated. Fat (oleate or olive oil) was infused intravenously in Wistar rats for 48 h with or without the IKKβ inhibitor salicylate. Thereafter, beta cell function was evaluated in vivo using hyperglycaemic clamps or ex vivo in islets isolated from fat-treated rats. We also exposed rat islets to oleate in culture, with or without salicylate and 4(2'-aminoethyl)amino-1,8-dimethylimidazo(1,2-a)quinoxaline; BMS-345541 (BMS, another inhibitor of IKKβ) and evaluated beta cell function in vitro. Furthermore, oleate was infused in mice treated with BMS and in beta cell-specific Ikkb-null mice. 48 h infusion of fat impaired beta-cell function in vivo, assessed using the disposition index (DI), in rats (saline: 1.41 ± 0.13; oleate: 0.95 ± 0.11; olive oil [OLO]: 0.87 ± 0.15; p < 0.01 for both fats vs saline) and in mice (saline: 2.51 ± 0.39; oleate: 1.20 ± 0.19; p < 0.01 vs saline) and ex vivo (i.e., insulin secretion, units are pmol insulin islet -1  h -1 ) in rat islets (saline: 1.51 ± 0.13; oleate: 1.03 ± 0.10; OLO: 0.91 ± 0.13; p < 0.001 for both fats vs saline) and the dysfunction was prevented by co-infusion of salicylate in rats (oleate + salicylate: 1.30 ± 0.09; OLO + salicylate: 1.33 ± 0.23) or BMS in mice (oleate + BMS: 2.25 ± 0.42) in vivo and by salicylate in rat islets ex vivo (oleate + salicylate: 1.74 ± 0.31; OLO + salicylate: 1.54 ± 0.29). In cultured islets, 48 h exposure to oleate impaired beta-cell function ([in pmol insulin islet -1  h -1 ] control: 0.66 ± 0.12; oleate: 0.23 ± 0.03; p < 0.01 vs saline), an effect prevented by both inhibitors (oleate + salicylate: 0.98 ± 0.08; oleate + BMS: 0.50 ± 0.02). Genetic

Entrapment of cultured pancreas islets in three-dimensional collagen matrices.

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Chao, S H; Peshwa, M V; Sutherland, D E; Hu, W S

1992-01-01

In vitro culture of islets of Langerhans decreases their immunogenicity, presumably by eliminating passenger leukocytes and other Ia+ presenting cells within the islets. Islets cultivated in petri dishes either at 37 degrees C or at 25 degrees C gradually disintegrate during culture in a time-dependent manner which is related to the free-floating condition of the islets. Also, a fraction of the islets disperse as single cells and beta-cell aggregates or adhere to the bottom of the culture dishes. Thus, the retrieval rate of transplantable islets is dampened due to their disintegration and spontaneous dispersion in conventional petri dish cultures. Entrapment of freshly harvested islets of Langerhans in a three-dimensional collagen matrix was studied as an alternative method for islet cultivation. The contraction of collagen fibrils during in vitro culture counteracts the dispersion of islets and helps in maintaining their integrity while in culture. It was observed that the entrapped islets maintain satisfactory morphology, viability, and capability of glucose-dependent insulin secretion for over 2 wk. The oxygen consumption rate and glucose metabolism of these islets was not deranged when entrapped in collagen. Also, the retrieval of islets is easier and more efficient than that observed in conventional culture systems. Our results indicate that culture of islets in three-dimensional collagen gels can potentially develop into an ideal system applicable to clinical transplantation of cultured islets or beta-cell aggregates.

JANEX-1, a JAK3 inhibitor, protects pancreatic islets from cytokine toxicity through downregulation of NF-{kappa}B activation and the JAK/STAT pathway

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Lv, Na; Kim, Eun-Kyung; Song, Mi-Young [Department of Biochemistry, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of); Choi, Ha-Na; Moon, Woo Sung [Department of Pathology, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of); Park, Sung-Joo [Department of Herbology, School of Oriental Medicine, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Park, Jin-Woo [Department of Biochemistry, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of); Kwon, Kang-Beom, E-mail: desson@wonkwang.ac.kr [Department of Physiology, School of Oriental Medicine, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Park, Byung-Hyun, E-mail: bhpark@chonbuk.ac.kr [Department of Biochemistry, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of)

2009-07-15

JANEX-1/WHI-P131, a selective Janus kinase 3 (JAK3) inhibitor, has been shown to delay the onset of diabetes in the NOD mouse model. However, the molecular mechanism by which JANEX-1 protects pancreatic {beta}-cells is unknown. In the current study, we investigated the role of JANEX-1 on interleukin (IL)-1{beta} and interferon (IFN)-{gamma}-induced {beta}-cell damage using isolated islets. JANEX-1-pretreated islets showed resistance to cytokine toxicity, namely suppressed nitric oxide (NO) production, reduced inducible form of NO synthase (iNOS) expression, and decreased islet destruction. The molecular mechanism by which JANEX-1 inhibits iNOS expression was mediated through suppression of the nuclear factor {kappa}B (NF-{kappa}B) and JAK/signal transducer and activator of transcription (STAT) pathways. Islets treated with the cytokines downregulated the protein levels of suppressor of cytokine signaling (SOCS)-1 and SOCS-3, but pretreatment with JANEX-1 attenuated these decreases. Additionally, islets from JAK3{sup -/-} mice were more resistant to cytokine toxicity than islets from control mice. These results demonstrate that JANEX-1 protects {beta}-cells from cytokine toxicity through suppression of the NF-{kappa}B and JAK/STAT pathways and upregulation of SOCS proteins, suggesting that JANEX-1 may be used to preserve functional {beta}-cell mass.

Intra- and Inter-islet Synchronization of Metabolically Driven Insulin Secretion

DEFF Research Database (Denmark)

Pedersen, Morten Gram; Bertram, Richard; Sherman, Arthur

2005-01-01

mechanisms for intra-islet and inter-islet synchronization. We show that electrical coupling is sufficient to synchronize both electrical bursting activity and metabolic oscillations. We also demonstrate that islets can synchronize by mutually entraining each other by their effects on a simple model "liver......,'' which responds to the level of insulin secretion by adjusting the blood glucose concentration in an appropriate way. Since all islets are exposed to the blood, the distributed islet-liver system can synchronize the individual islet insulin oscillations. Thus, we demonstrate how intra-islet and inter...

Roles of Toll-like receptors in allogeneic islet transplantation.

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Ro, Han; Hong, Juho; Kim, Beom Seok; Lee, Eun Won; Kim, Myung-Gyu; Han, Kyu Hyun; Yeom, Hye-Jung; Lee, Eun Mi; Jeong, Jong Cheol; Oh, Kook-Hwan; Ahn, Curie; Yang, Jaeseok

2012-11-27

Toll-like receptors (TLRs) are involved in the rejection of solid organ allografts. However, the roles of TLRs in islets are still controversial. We investigated the roles of TLRs in donor islets together with those in recipients in allogeneic islet transplantation. To assess the roles of TLRs in either donor islets or recipients, allogeneic islet transplantation was performed using myeloid differentiation factor 88 (MyD88)-knockout (KO), TLR4-KO, or Toll/interleukin-1 receptor domain-containing adaptor-inducing interferon-β (TRIF)-KO mice. Both polyriboinosinic polyribocytidylic acid and lipopolysaccharide (LPS) stimulation induced the mRNA expression of regulated and normal T cell expressed and secreted, interferon-γ-inducible protein-10, monocyte chemotactic protein-1, interleukin-8, and inducible nitric oxide synthase in murine islets, whereas the induction was attenuated in TRIF-KO, interferon-β promoter stimulator-1-KO, and TLR4-KO mice. When islets from MyD88-KO, TLR4-KO, or TRIF-KO C57BL/6 mice were transplanted to BALB/c recipients, graft survival was not better than that of wild-type (WT) islets. However, the survival of the MyD88-KO islet allograft was significantly prolonged when combined with anti-CD40L. In parallel, LPS stimulation in donor islets interfered with anti-CD40L blockade-mediated long-term survival of islet allografts in TLR4-KO recipients. LPS stimulation increased the perigraft infiltration of both T cells and macrophages. Then again, when islets from WT BALB/c mice were transplanted to MyD88-KO, TRIF-KO, or WT C57BL/6 mice, there was no difference in graft survival, although some of the MyD88-KO recipients obtained long-term graft survival. However, anti-CD40L prolonged graft survival significantly in MyD88-KO recipients. The absence of MyD88 in either donors or recipients decreased the perigraft infiltration of inflammatory cells when combined with anti-CD40L. TLRs in both donor islets and recipients are involved in islet allograft

Impact of anti-insulin antibodies on islet transplantation outcome: data from the GRAGIL Network.

Science.gov (United States)

Lablanche, Sandrine; Borot, Sophie; Thaunat, Olivier; Bayle, Francois; Badet, Lionel; Morelon, Emmanuel; Thivolet, Charles; Wojtusciszyn, Anne; Frimat, Luc; Kessler, Laurence; Penfornis, Alfred; Brault, Coralie; Colin, Cyrille; Bosco, Domenico; Berney, Thierry; Benhamou, Pierre Y

2014-08-27

In patients with type 1 diabetes, insulin antibodies (IA), altering the pharmacokinetics of circulating insulin, might be associated with high glucose concentration, prolonged hypoglycemia, and higher insulin requirement. The impact of IA on islet transplantation has never been explored. Our aim was to evaluate islet transplantation results at 1 year according to the presence of IA. Our work is a retrospective, case-control study, comparing IA-negative and IA-positive patients among the cohort of patients with type 1 diabetes transplanted within the Swiss-French GRAGIL network between 2003 and 2010. Data about IA were available for 17 patients. Before islet transplantation, 10 patients (59%) were screened positive for IA. At 12 months after transplantation, IA-positive patients reached insulin independence less frequently than IA-negative patients (cumulative incidence of insulin independence, 22.2% vs. 71.4%; P=0.02); β score was ≥7 in 43% of IA-negative patients versus 0% in IA-positive patients (P=0.022). When comparing IA-positive patients with IA-negative patients, insulin dose was 0.15 U/kg (0.10-0.18 U/kg) versus 0.01 U/kg (0-0.09 U/kg) (P=0.2); HbA1c was 6.1% (5.8%-6.3%) versus 6.1% (5.9%-6.8%) (P=0.16); basal C-peptide level was 460 ρmol/L (350-510 ρmol/L) versus 265 ρmol/L (177-405 ρmol/L) (P=0.28); occurrence of hypoglycemia was 12.5% versus 16.5% (P=0.9); and homeostatic model assessment insulin resistance was 1.25 (1-2.4) versus 0.7 (0.52-0.92) (P=0.01). After islet transplantation, IA-positive patients achieved insulin independence less frequently, exhibiting lower β score and higher homeostatic model assessment insulin resistance compared with IA-negative patients. However, in both groups, islet transplantation restored good glycemic control and drastically reduced hypoglycemia and insulin requirements.

Long-term effects of islet transplantation.

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Holmes-Walker, D Jane; Kay, Thomas W H

2016-10-01

Islet transplantation has made great progress in recent years. This is a remarkable technical feat but raises the question of what the long-term benefits and risks are for type I diabetes recipients. Graft survival continues to improve, and recent multicenter studies show that islet transplantation is particularly effective to prevent hypoglycemic events even in those who do not become insulin-independent and to achieve excellent glycemic control. Concerns include histocompatability leucocyte antigen (HLA) sensitization and other risks including from immunosuppression that islet transplantation shares with other forms of allotransplantation. Reversal of hypoglycemia unawareness and protection from severe hypoglycemia events are two of the main benefits of islet transplantation and they persist for the duration of graft function. Islet transplantation compares favorably with other therapies for those with hypoglycemia unawareness, although new technologies have not been tested head-to-head with transplantation. HLA sensitization increases with time after transplantation especially if immunosuppression is ceased and is a risk for those who may require future transplantation as well as being associated with loss of graft function.

Regulatory challenges in manufacturing of pancreatic islets.

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Linetsky, E; Ricordi, C

2008-03-01

At the present time, transplantation of pancreatic islet cells is considered an experimental therapy for a selected cohort of patients with type 1 diabetes, and is conducted under an Investigational New Drug (IND) application. Encouraging results of the Edmonton Protocol published in the year 2000 sparked a renewed interest in clinical transplantation of allogeneic islets, triggering a large number of IND applications for phase I clinical trials. Promising results reported by a number of centers since then prompted the Food and Drug Administration (FDA) to consider the possibility of licensing allogeneic islets as a therapeutic treatment for patients with type 1 diabetes. However, prior to licensure, issues such as safety, purity, efficacy, and potency of the islet product must be addressed. This is complicated by the intricate nature of pancreatic islets and limited characterization prior to transplantation. In this context, control of the manufacturing process plays a critical role in the definition of the final product. Despite significant progress made in standardization of the donor organ preservation methods, reagents used, and characterization assays performed to qualify an islet cell product, control of the isolation process remains a challenge. Within the scope of the FDA regulations, islet cells meet the definition of a biologic product, somatic cell therapy, and a drug. In addition, AABB standards that address cellular therapy products apply to manufacturing facilities accredited by this organization. Control of the source material, isolation process, and final product are critical issues that must be addressed in the context of FDA and other relevant regulations applicable to islet cell products.

Effect of liver histopathology on islet cell engraftment in the model mimicking autologous islet cell transplantation.

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Desai, Chirag S; Khan, Khalid M; Ma, Xiaobo; Li, Henghong; Wang, Juan; Fan, Lijuan; Chen, Guoling; Smith, Jill P; Cui, Wanxing

2017-11-02

The inflammatory milieu in the liver as determined by histopathology is different in individual patients undergoing autologous islet cell transplantation. We hypothesized that inflammation related to fatty-liver adversely impacts islet survival. To test this hypothesis, we used a mouse model of fatty-liver to determine the outcome of syngeneic islet transplantation after chemical pancreatectomy. Mice (C57BL/6) were fed a high-fat-diet from 6 weeks of age until attaining a weight of ≥28 grams (6-8 weeks) to produce a fatty liver (histologically > 30% fat);steatosis was confirmed with lipidomic profile of liver tissue. Islets were infused via the intra-portal route in fatty-liver and control mice after streptozotocin induction of diabetes. Outcomes were assessed by the rate of euglycemia, liver histopathology, evaluation of liver inflammation by measuring tissue cytokines IL-1β and TNF-α by RT-PCR and CD31 expression by immunohistochemistry. The difference in the euglycemic fraction between the normal liver group (90%, 9/10) and the fatty-liver group (37.5%, 3/8) was statistically significant at the 18 th day post- transplant and was maintained to the end of the study (day 28) (p = 0.019, X 2 = 5.51). Levels of TNF-α and IL-1β were elevated in fatty-liver mice (p = 0.042, p = 0.037). Compared to controls cytokine levels were elevated after islet cell transplantation and in transplanted fatty-liver mice as compared to either fatty- or islet transplant group alone (p = NS). A difference in the histochemical pattern of CD31 could not be determined. Fatty-liver creates an inflammatory state which adversely affects the outcome of autologous islet cell transplantation.

Glucose metabolism, islet architecture, and genetic homogeneity in imprinting of [Ca2+](i and insulin rhythms in mouse islets.

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Craig S Nunemaker

2009-12-01

Full Text Available We reported previously that islets isolated from individual, outbred Swiss-Webster mice displayed oscillations in intracellular calcium ([Ca2+](i that varied little between islets of a single mouse but considerably between mice, a phenomenon we termed "islet imprinting." We have now confirmed and extended these findings in several respects. First, imprinting occurs in both inbred (C57BL/6J as well as outbred mouse strains (Swiss-Webster; CD1. Second, imprinting was observed in NAD(PH oscillations, indicating a metabolic component. Further, short-term exposure to a glucose-free solution, which transiently silenced [Ca2+](i oscillations, reset the oscillatory patterns to a higher frequency. This suggests a key role for glucose metabolism in maintaining imprinting, as transiently suppressing the oscillations with diazoxide, a K(ATP-channel opener that blocks [Ca2+](i influx downstream of glucose metabolism, did not change the imprinted patterns. Third, imprinting was not as readily observed at the level of single beta cells, as the [Ca2+](i oscillations of single cells isolated from imprinted islets exhibited highly variable, and typically slower [Ca2+](i oscillations. Lastly, to test whether the imprinted [Ca2+](i patterns were of functional significance, a novel microchip platform was used to monitor insulin release from multiple islets in real time. Insulin release patterns correlated closely with [Ca2+](i oscillations and showed significant mouse-to-mouse differences, indicating imprinting. These results indicate that islet imprinting is a general feature of islets and is likely to be of physiological significance. While islet imprinting did not depend on the genetic background of the mice, glucose metabolism and intact islet architecture may be important for the imprinting phenomenon.

A novel insulinotropic mechanism of whole grain-derived γ-oryzanol via the suppression of local dopamine D2 receptor signalling in mouse islet.

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Kozuka, Chisayo; Sunagawa, Sumito; Ueda, Rei; Higa, Moritake; Ohshiro, Yuzuru; Tanaka, Hideaki; Shimizu-Okabe, Chigusa; Takayama, Chitoshi; Matsushita, Masayuki; Tsutsui, Masato; Ishiuchi, Shogo; Nakata, Masanori; Yada, Toshihiko; Miyazaki, Jun-Ichi; Oyadomari, Seiichi; Shimabukuro, Michio; Masuzaki, Hiroaki

2015-07-03

γ-Oryzanol, derived from unrefined rice, attenuated the preference for dietary fat in mice, by decreasing hypothalamic endoplasmic reticulum stress. However, no peripheral mechanisms, whereby γ-oryzanol could ameliorate glucose dyshomeostasis were explored. Dopamine D 2 receptor signalling locally attenuates insulin secretion in pancreatic islets, presumably via decreased levels of intracellular cAMP. We therefore hypothesized that γ-oryzanol would improve high-fat diet (HFD)-induced dysfunction of islets through the suppression of local D 2 receptor signalling. Glucose metabolism and regulation of molecules involved in D 2 receptor signalling in pancreatic islets were investigated in male C57BL/6J mice, fed HFD and treated with γ-oryzanol . In isolated murine islets and the beta cell line, MIN6 , the effects of γ-oryzanol on glucose-stimulated insulin secretion (GSIS) was analysed using siRNA for D 2 receptors and a variety of compounds which alter D 2 receptor signalling. In islets, γ-oryzanol enhanced GSIS via the activation of the cAMP/PKA pathway. Expression of molecules involved in D 2 receptor signalling was increased in islets from HFD-fed mice, which were reciprocally decreased by γ-oryzanol. Experiments with siRNA for D 2 receptors and D 2 receptor ligands in vitro suggest that γ-oryzanol suppressed D 2 receptor signalling and augmented GSIS. γ-Oryzanol exhibited unique anti-diabetic properties. The unexpected effects of γ-oryzanol on D 2 receptor signalling in islets may provide a novel; natural food-based, approach to anti-diabetic therapy. © 2015 The British Pharmacological Society.

Isolation of Human Islets for Autologous Islet Transplantation in Children and Adolescents with Chronic Pancreatitis

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Rita Bottino

2012-01-01

Full Text Available Chronic pancreatitis is an inflammatory disease of the pancreas that causes permanent changes in the function and structure of the pancreas. It is most commonly a complication of cystic fibrosis or due to a genetic predisposition. Chronic pancreatitis generally presents symptomatically as recurrent abdominal pain, which becomes persistent over time. The pain eventually becomes disabling. Once specific medical treatments and endoscopic interventions are no longer efficacious, total pancreatectomy is the alternative of choice for helping the patient achieve pain control. While daily administrations of digestive enzymes cannot be avoided, insulin-dependent diabetes can be prevented by transplanting the isolated pancreatic islets back to the patient. The greater the number of islets infused, the greater the chance to prevent or at least control the effects of surgical diabetes. We present here a technical approach for the isolation and preservation of the islets proven to be efficient to obtain high numbers of islets, favoring the successful treatment of young patients.

Autologous Pancreatic Islet Transplantation in Human Bone Marrow

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Maffi, Paola; Balzano, Gianpaolo; Ponzoni, Maurilio; Nano, Rita; Sordi, Valeria; Melzi, Raffaella; Mercalli, Alessia; Scavini, Marina; Esposito, Antonio; Peccatori, Jacopo; Cantarelli, Elisa; Messina, Carlo; Bernardi, Massimo; Del Maschio, Alessandro; Staudacher, Carlo; Doglioni, Claudio; Ciceri, Fabio; Secchi, Antonio; Piemonti, Lorenzo

2013-01-01

The liver is the current site of choice for pancreatic islet transplantation, even though it is far from being ideal. We recently have shown in mice that the bone marrow (BM) may be a valid alternative to the liver, and here we report a pilot study to test feasibility and safety of BM as a site for islet transplantation in humans. Four patients who developed diabetes after total pancreatectomy were candidates for the autologous transplantation of pancreatic islet. Because the patients had contraindications for intraportal infusion, islets were infused in the BM. In all recipients, islets engrafted successfully as shown by measurable posttransplantation C-peptide levels and histopathological evidence of insulin-producing cells or molecular markers of endocrine tissue in BM biopsy samples analyzed during follow-up. Thus far, we have recorded no adverse events related to the infusion procedure or the presence of islets in the BM. Islet function was sustained for the maximum follow-up of 944 days. The encouraging results of this pilot study provide new perspectives in identifying alternative sites for islet infusion in patients with type 1 diabetes. Moreover, this is the first unequivocal example of successful engraftment of endocrine tissue in the BM in humans. PMID:23733196

Vagotomy ameliorates islet morphofunction and body metabolic homeostasis in MSG-obese rats

International Nuclear Information System (INIS)

Lubaczeuski, C.; Balbo, S.L.; Ribeiro, R.A.; Vettorazzi, J.F.; Santos-Silva, J.C.; Carneiro, E.M.; Bonfleur, M.L.

2015-01-01

The parasympathetic nervous system is important for β-cell secretion and mass regulation. Here, we characterized involvement of the vagus nerve in pancreatic β-cell morphofunctional regulation and body nutrient homeostasis in 90-day-old monosodium glutamate (MSG)-obese rats. Male newborn Wistar rats received MSG (4 g/kg body weight) or saline [control (CTL) group] during the first 5 days of life. At 30 days of age, both groups of rats were submitted to sham-surgery (CTL and MSG groups) or subdiaphragmatic vagotomy (Cvag and Mvag groups). The 90-day-old MSG rats presented obesity, hyperinsulinemia, insulin resistance, and hypertriglyceridemia. Their pancreatic islets hypersecreted insulin in response to glucose but did not increase insulin release upon carbachol (Cch) stimulus, despite a higher intracellular Ca 2+ mobilization. Furthermore, while the pancreas weight was 34% lower in MSG rats, no alteration in islet and β-cell mass was observed. However, in the MSG pancreas, increases of 51% and 55% were observed in the total islet and β-cell area/pancreas section, respectively. Also, the β-cell number per β-cell area was 19% higher in MSG rat pancreas than in CTL pancreas. Vagotomy prevented obesity, reducing 25% of body fat stores and ameliorated glucose homeostasis in Mvag rats. Mvag islets demonstrated partially reduced insulin secretion in response to 11.1 mM glucose and presented normalization of Cch-induced Ca 2+ mobilization and insulin release. All morphometric parameters were similar among Mvag and CTL rat pancreases. Therefore, the higher insulin release in MSG rats was associated with greater β-cell/islet numbers and not due to hypertrophy. Vagotomy improved whole body nutrient homeostasis and endocrine pancreatic morphofunction in Mvag rats

Alterations of pancreatic islet structure, metabolism and gene expression in diet-induced obese C57BL/6J mice.

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Regan Roat

Full Text Available The reduction of functional β cell mass is a key feature of type 2 diabetes. Here, we studied metabolic functions and islet gene expression profiles of C57BL/6J mice with naturally occurring nicotinamide nucleotide transhydrogenase (NNT deletion mutation, a widely used model of diet-induced obesity and diabetes. On high fat diet (HF, the mice developed obesity and hyperinsulinemia, while blood glucose levels were only mildly elevated indicating a substantial capacity to compensate for insulin resistance. The basal serum insulin levels were elevated in HF mice, but insulin secretion in response to glucose load was significantly blunted. Hyperinsulinemia in HF fed mice was associated with an increase in islet mass and size along with higher BrdU incorporation to β cells. The temporal profiles of glucose-stimulated insulin secretion (GSIS of isolated islets were comparable in HF and normal chow fed mice. Islets isolated from HF fed mice had elevated basal oxygen consumption per islet but failed to increase oxygen consumption further in response to glucose or carbonyl cyanide-4-trifluoromethoxyphenylhydrazone (FCCP. To obtain an unbiased assessment of metabolic pathways in islets, we performed microarray analysis comparing gene expression in islets from HF to normal chow-fed mice. A few genes, for example, those genes involved in the protection against oxidative stress (hypoxia upregulated protein 1 and Pgc1α were up-regulated in HF islets. In contrast, several genes in extracellular matrix and other pathways were suppressed in HF islets. These results indicate that islets from C57BL/6J mice with NNT deletion mutation develop structural, metabolic and gene expression features consistent with compensation and decompensation in response to HF diet.

The journey of islet cell transplantation and future development.

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Gamble, Anissa; Pepper, Andrew R; Bruni, Antonio; Shapiro, A M James

2018-03-04

Intraportal islet transplantation has proven to be efficacious in preventing severe hypoglycemia and restoring insulin independence in selected patients with type 1 diabetes. Multiple islet infusions are often required to achieve and maintain insulin independence. Many challenges remain in clinical islet transplantation, including substantial islet cell loss early and late after islet infusion. Contributions to graft loss include the instant blood-mediated inflammatory reaction, potent host auto- and alloimmune responses, and beta cell toxicity from immunosuppressive agents. Protective strategies are being tested to circumvent several of these events including exploration of alternative transplantation sites, stem cell-derived insulin producing cell therapies, co-transplantation with mesenchymal stem cells or exploration of novel immune protective agents. Herein, we provide a brief introduction and history of islet cell transplantation, limitations associated with this procedure and methods to alleviate islet cell loss as a means to improve engraftment outcomes.

Striated Muscle as Implantation Site for Transplanted Pancreatic Islets

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Daniel Espes

2011-01-01

Full Text Available Islet transplantation is an attractive treatment for selected patients with brittle type 1 diabetes. In the clinical setting, intraportal transplantation predominates. However, due to extensive early islet cell death, the quantity of islets needed to restore glucose homeostasis requires in general a minimum of two donors. Moreover, the deterioration of islet function over time results in few insulin-independent patients after five-year followup. Specific obstacles to the success of islet transplantation include site-specific concerns for the liver such as the instant blood mediated inflammatory reaction, islet lipotoxicity, low oxygen tension, and poor revascularization, impediments that have led to the developing interest for alternative implantation sites over recent years. Within preclinical settings, several alternative sites have now been investigated and proven favorable in various aspects. Muscle is considered a very promising site and has physiologically properties and technical advantages that could make it optimal for islet transplantation.

Ketosis Onset Type 2 Diabetes Had Better Islet β-Cell Function and More Serious Insulin Resistance

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Hongyun Lu

2014-01-01

Full Text Available Diabetic ketosis had been identified as a characteristic of type 1 diabetes mellitus (T1DM, but now emerging evidence has identified that they were diagnosed as T2DM after long time follow up. This case control study was aimed at comparing the clinical characteristic, β-cell function, and insulin resistance of ketosis and nonketotic onset T2DM and providing evidence for treatment selection. 140 cases of newly diagnosed T2DM patients were divided into ketosis (62 cases and nonketotic onset group (78 cases. After correction of hyperglycemia and ketosis with insulin therapy, plasma C-peptide concentrations were measured at 0, 0.5, 1, 2, and 3 hours after 75 g glucose oral administration. Area under the curve (AUC of C-peptide was calculated. Homoeostasis model assessment was used to estimate basal β-cell function (HOMA-β and insulin resistance (HOMA-IR. Our results showed that ketosis onset group had higher prevalence of nonalcoholic fatty liver disease (NAFLD than nonketotic group (P=0.04. Ketosis onset group had increased plasma C-peptide levels at 0 h, 0.5 h, and 3 h and higher AUC0–0.5, AUC0–1, AUC0–3 (P<0.05. Moreover, this group also had higher HOMA-β and HOMA-IR than nonketotic group (P<0.05. From these data, we concluded that ketosis onset T2DM had better islet β-cell function and more serious insulin resistance than nonketotic onset T2DM.

Mitochondrial dysfunction precedes depression of AMPK/AKT signaling in insulin resistance induced by high glucose in primary cortical neurons.

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Peng, Yunhua; Liu, Jing; Shi, Le; Tang, Ying; Gao, Dan; Long, Jiangang; Liu, Jiankang

2016-06-01

Recent studies have demonstrated brain insulin signaling impairment and mitochondrial dysfunction in diabetes. Hyperinsulinemia and hyperlipidemia arising from diabetes have been linked to neuronal insulin resistance, and hyperglycemia induces peripheral sensory neuronal impairment and mitochondrial dysfunction. However, how brain glucose at diabetic conditions elicits cortical neuronal insulin signaling impairment and mitochondrial dysfunction remains unknown. In the present study, we cultured primary cortical neurons with high glucose levels and investigated the neuronal mitochondrial function and insulin response. We found that mitochondrial function was declined in presence of 10 mmol/L glucose, prior to the depression of AKT signaling in primary cortical neurons. We further demonstrated that the cerebral cortex of db/db mice exhibited both insulin resistance and loss of mitochondrial complex components. Moreover, we found that adenosine monophosphate-activated protein kinase (AMPK) inactivation is involved in high glucose-induced mitochondrial dysfunction and insulin resistance in primary cortical neurons and neuroblastoma cells, as well as in cerebral cortex of db/db mice, and all these impairments can be rescued by mitochondrial activator, resveratrol. Taken together, our results extend the finding that high glucose (≥10 mmol/L) comparable to diabetic brain extracellular glucose level leads to neuronal mitochondrial dysfunction and resultant insulin resistance, and targeting mitochondria-AMPK signaling might be a promising strategy to protect against diabetes-related neuronal impairment in central nerves system. We found that high glucose (≥10 mmol/L), comparable to diabetic brain extracellular glucose level, leads to neuronal mitochondrial dysfunction and resultant insulin resistance in an AMPK-dependent manner, and targeting mitochondria-AMPK signaling might be a promising strategy to protect against diabetes-related neuronal impairment in central

Mechanisms of pancreatic islet cell destruction. Dose-dependent cytotoxic effect of soluble blood mononuclear cell mediators on isolated islets of Langerhans

DEFF Research Database (Denmark)

Mandrup-Poulsen, T; Bendtzen, K; Nerup, J

1986-01-01

Supernatants of peripheral blood mononuclear cells from healthy human donors stimulated with recall antigen (purified protein derivative of tuberculin) or lectin (phytohaemagglutinin) markedly inhibited the insulin release from isolated human and rat islets of Langerhans, and decreased rat islet...... reconstituted with tuberculin or phytohaemagglutinin did not impair islet function. Electron microscopy demonstrated that supernatants were cytotoxic to islet cells. The cytotoxic mononuclear cell mediator(s) was non-dialysable, sensitive to heating to 56 degrees C, labile even when stored at -70 degrees C...

Rat pancreatic islet size standardization by the "hanging drop" technique.

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Cavallari, G; Zuellig, R A; Lehmann, R; Weber, M; Moritz, W

2007-01-01

Rejection and hypoxia are the main factors that limit islet engraftment in the recipient liver in the immediate posttransplant period. Recently authors have reported a negative relationship of graft function and islet size, concluding that small islets are superior to large islets. Islets can be dissociated into single cells and reaggregated into so called "pseudoislets," which are functionally equivalent to intact islets but exhibit reduced immunogenicity. The aim of our study was develop a technique that enabled one to obtain pseudoislets of defined, preferably small, dimensions. Islets were harvested from Lewis rats by the collagenase digestion procedure. After purification, the isolated islets were dissociated into single cells by trypsin digestion. Fractions with different cell numbers were seeded into single drops onto cell culture dishes, which were inverted and incubated for 5 to 8 days under cell culture conditions. Newly formed pseudoislets were analyzed for dimension, morphology, and cellular composition. The volume of reaggregated pseudoislets strongly correlated with the cell number (r(2) = .995). The average diameter of a 250-cell aggregate was 95 +/- 8 microm (mean +/- SD) compared with 122 +/- 46 microm of freshly isolated islets. Islet cell loss may be minimized by performing reaggregation in the presence of medium glucose (11 mmol/L) and the GLP-1 analogue Exendin-4. Morphology, cellular composition, and architecture of reaggregated islets were comparable to intact islets. The "hanging drop" culture method allowed us to obtain pseudoislets of standardized size and regular shape, which did not differ from intact islets in terms of cellular composition or architecture. Further investigations are required to minimize cell loss and test in vivo function of transplanted pseudoislets.

A novel subcutaneous site of islet transplantation superior to the liver.

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Yasunami, Yohichi; Nakafusa, Yuki; Nitta, Naoyoshi; Nakamura, Masafumi; Goto, Masafumi; Ono, Junko; Taniguchi, Masaru

2018-03-08

Islet transplantation is an attractive treatment for patients with insulin-dependent diabetes mellitus, and currently the liver is the favored transplantation site. However, an alternative site is desirable because of the low efficiency of hepatic transplantation, requiring 2-3 donors for a single recipient, and because the transplanted islets cannot be accessed or retrieved. We developed a novel procedure of islet transplantation to the inguinal subcutaneous white adipose tissue (ISWAT) of mice and described functional and morphological characteristics of transplanted syngeneic islets. Also, it was determined whether islet allograft rejection in the ISWAT can be prevented by immunosuppressive agents. Furthermore, it was examined whether human islets function when grafted in this particular site of immune-deficient mice. In this site, transplanted islets are engrafted as clusters and function to reverse STZ-induced diabetes in mice. Importantly, transplanted islets can be visualized by CT and are easily retrievable, and allograft rejection is preventable by blockade of co-stimulatory signals. Of much importance, the efficiency of islet transplantation in this site is superior to the liver, in which hyperglycemia of diabetic recipient mice is ameliorated after transplantation of 200 syngeneic islets (the islet number yielded from 1 mouse pancreas) to the ISWAT but not to the liver. Furthermore, human islets transplanted in this particular site function to reverse diabetes in immune-deficient mice. Thus, the ISWAT is superior to the liver as the site of islet transplantation, which may lead to improved outcome of clinical islet transplantation.

Exploiting mitochondrial dysfunction for effective elimination of imatinib-resistant leukemic cells.

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Jérome Kluza

Full Text Available Challenges today concern chronic myeloid leukemia (CML patients resistant to imatinib. There is growing evidence that imatinib-resistant leukemic cells present abnormal glucose metabolism but the impact on mitochondria has been neglected. Our work aimed to better understand and exploit the metabolic alterations of imatinib-resistant leukemic cells. Imatinib-resistant cells presented high glycolysis as compared to sensitive cells. Consistently, expression of key glycolytic enzymes, at least partly mediated by HIF-1α, was modified in imatinib-resistant cells suggesting that imatinib-resistant cells uncouple glycolytic flux from pyruvate oxidation. Interestingly, mitochondria of imatinib-resistant cells exhibited accumulation of TCA cycle intermediates, increased NADH and low oxygen consumption. These mitochondrial alterations due to the partial failure of ETC were further confirmed in leukemic cells isolated from some imatinib-resistant CML patients. As a consequence, mitochondria generated more ROS than those of imatinib-sensitive cells. This, in turn, resulted in increased death of imatinib-resistant leukemic cells following in vitro or in vivo treatment with the pro-oxidants, PEITC and Trisenox, in a syngeneic mouse tumor model. Conversely, inhibition of glycolysis caused derepression of respiration leading to lower cellular ROS. In conclusion, these findings indicate that imatinib-resistant leukemic cells have an unexpected mitochondrial dysfunction that could be exploited for selective therapeutic intervention.

A review of piscine islet xenotransplantation using wild-type tilapia donors and the production of transgenic tilapia expressing a "humanized" tilapia insulin.

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Wright, James R; Yang, Hua; Hyrtsenko, Olga; Xu, Bao-You; Yu, Weiming; Pohajdak, Bill

2014-01-01

Most islet xenotransplantation laboratories have focused on porcine islets, which are both costly and difficult to isolate. Teleost (bony) fish, such as tilapia, possess macroscopically visible distinct islet organs called Brockmann bodies which can be inexpensively harvested. When transplanted into diabetic nude mice, tilapia islets maintain long-term normoglycemia and provide human-like glucose tolerance profiles. Like porcine islets, when transplanted into euthymic mice, they are rejected in a CD4 T-cell-dependent manner. However, unlike pigs, tilapia are so phylogenetically primitive that their cells do not express α(1,3)Gal and, because tilapia are highly evolved to live in warm stagnant waters nearly devoid of dissolved oxygen, their islet cells are exceedingly resistant to hypoxia, making them ideal for transplantation within encapsulation devices. Encapsulation, especially when combined with co-stimulatory blockade, markedly prolongs tilapia islet xenograft survival in small animal recipients, and a collaborator has shown function in diabetic cynomolgus monkeys. In anticipation of preclinical xenotransplantation studies, we have extensively characterized tilapia islets (morphology, embryologic development, cell biology, peptides, etc.) and their regulation of glucose homeostasis. Because tilapia insulin differs structurally from human insulin by 17 amino acids, we have produced transgenic tilapia whose islets stably express physiological levels of humanized insulin and have now bred these to homozygosity. These transgenic fish can serve as a platform for further development into a cell therapy product for diabetes. © 2014 The Authors. Xenotransplantation Published by John Wiley & Sons Ltd.

The effect of curcumin on insulin release in rat-isolated pancreatic islets.

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Abdel Aziz, Mohamed T; El-Asmar, Mohamed F; El Nadi, Essam G; Wassef, Mohamed A; Ahmed, Hanan H; Rashed, Laila A; Obaia, Eman M; Sabry, Dina; Hassouna, Amira A; Abdel Aziz, Ahmed T

2010-08-01

Curcumin exerts a hypoglycemic action and induces heme-oxygenase-1 (HO-1). We evaluated the effect of curcumin on isolated islets of Langerhans and studied whether its action on insulin secretion is mediated by inducible HO-1. Islets were isolated from rats and divided into control islets, islets incubated in different curcumin concentrations, islets incubated in hemin, islets incubated in curcumin and HO inhibitor, stannous mesoporphyrin (SnMP), islets incubated in hemin and SnMP, islets incubated in SnMP only, and islets incubated in 16.7 mmol/L glucose. Heme-oxygenase activity, HO-1 expression, and insulin estimation was assessed. Insulin secretion, HO-1 gene expression and HO activity were significantly increased in islets incubated in curcumin, hemin, and glucose compared with controls. This increase in insulin secretion was significantly decreased by incubation of islets in SnMP. The action of curcumin on insulin secretion from the isolated islets may be, in part, mediated through increased HO-1 gene expression.

Vagotomy ameliorates islet morphofunction and body metabolic homeostasis in MSG-obese rats

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Lubaczeuski, C.; Balbo, S.L. [Laboratório de Fisiologia Endócrina e Metabolismo, Centro de Ciências Biológicas e da Saúde, Universidade Estadual do Oeste do Paraná, Cascavel, PR (Brazil); Ribeiro, R.A. [Universidade Federal do Rio de Janeiro, Macaé, RJ (Brazil); Vettorazzi, J.F.; Santos-Silva, J.C.; Carneiro, E.M. [Laboratório de Pâncreas Endócrino e Metabolismo, Departamento de Biologia Estrutural e Funcional, Instituto de Biologia, Universidade Estadual de Campinas, Campinas, SP (Brazil); Bonfleur, M.L. [Laboratório de Fisiologia Endócrina e Metabolismo, Centro de Ciências Biológicas e da Saúde, Universidade Estadual do Oeste do Paraná, Cascavel, PR (Brazil)

2015-02-24

The parasympathetic nervous system is important for β-cell secretion and mass regulation. Here, we characterized involvement of the vagus nerve in pancreatic β-cell morphofunctional regulation and body nutrient homeostasis in 90-day-old monosodium glutamate (MSG)-obese rats. Male newborn Wistar rats received MSG (4 g/kg body weight) or saline [control (CTL) group] during the first 5 days of life. At 30 days of age, both groups of rats were submitted to sham-surgery (CTL and MSG groups) or subdiaphragmatic vagotomy (Cvag and Mvag groups). The 90-day-old MSG rats presented obesity, hyperinsulinemia, insulin resistance, and hypertriglyceridemia. Their pancreatic islets hypersecreted insulin in response to glucose but did not increase insulin release upon carbachol (Cch) stimulus, despite a higher intracellular Ca{sup 2+} mobilization. Furthermore, while the pancreas weight was 34% lower in MSG rats, no alteration in islet and β-cell mass was observed. However, in the MSG pancreas, increases of 51% and 55% were observed in the total islet and β-cell area/pancreas section, respectively. Also, the β-cell number per β-cell area was 19% higher in MSG rat pancreas than in CTL pancreas. Vagotomy prevented obesity, reducing 25% of body fat stores and ameliorated glucose homeostasis in Mvag rats. Mvag islets demonstrated partially reduced insulin secretion in response to 11.1 mM glucose and presented normalization of Cch-induced Ca{sup 2+} mobilization and insulin release. All morphometric parameters were similar among Mvag and CTL rat pancreases. Therefore, the higher insulin release in MSG rats was associated with greater β-cell/islet numbers and not due to hypertrophy. Vagotomy improved whole body nutrient homeostasis and endocrine pancreatic morphofunction in Mvag rats.

Separation of empty microcapsules after microencapsulation of porcine neonatal islets.

Science.gov (United States)

Shin, Soojeong; Yoo, Young Je

2013-12-01

Pancreatic islet transplantation is used to treat diabetes mellitus that has minimal complications and avoids hypoglycemic shock. Conformal microencapsulation of pancreatic islets improves their function by blocking immunogenic molecules while protecting fragile islets. However, production of empty alginate capsules during microencapsulation causes enlargement of the transplantation volume of the encapsulated islets and interferes with efficient transfer of nutrients and insulin. In this study, empty alginate capsules were separated after microencapsulation of neonatal porcine islet-like cell clusters (NPCC) using density-gradient centrifugation. Densities of NPCC and alginate capsules were determined using Percoll. Encapsulation products following alginate removal were 97 % of products, with less than 10 % of the capsules remaining empty. The viability of this process compared with manually-selected encapsulated islets indicates the separation process does not harm islets.

Compromised Photosynthetic Electron Flow And H2O2 Generation Correlate with Genotype-Specific Stomatal Dysfunctions During Resistance Against Powdery Mildew In Oats.

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Javier Sánchez-Martín

2016-11-01

Full Text Available Stomatal dysfunction known as locking has been linked to the elicitation of a hypersensitive response (HR following attack of fungal pathogens in cereals. We here assess how spatial and temporal patterns of different resistance mechanisms, such as HR and penetration resistance influence stomatal and photosynthetic parameters in oat (Avena sativa and the possible involvement of hydrogen peroxide (H2O2 in the dysfunctions observed. Four oat cultivars with differential resistance responses (i.e. penetration resistance, early and late HR to powdery mildew (Blumeria graminis f. sp. avenae, Bga were used. Results demonstrated that stomatal dysfunctions were genotype but not response-type dependent since genotypes with similar resistance responses when assessed histologically showed very different locking patterns. Maximum quantum yield (Fv/Fm of photosystem II were compromised in most Bga–oat interactions and photoinhibition increased. However, the extent of the photosynthetic alterations was not directly related to the extent of HR. H2O2 generation is triggered during the execution of resistance responses and can influence stomatal function. Artificially increasing H2O2 by exposing plants to increased light intensity further reduced Fv/Fm ratios and augmented the patterns of stomatal dysfunctions previously observed. The latter results suggest that the observed dysfunctions and hence a cost of resistance may be linked with oxidative stress occurring during defence induced photosynthetic disruption.

Molecular Imaging: A Promising Tool to Monitor Islet Transplantation

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Ping Wang

2011-01-01

Full Text Available Replacement of insulin production by pancreatic islet transplantation has great potential as a therapy for type 1 diabetes mellitus. At present, the lack of an effective approach to islet grafts assessment limits the success of this treatment. The development of molecular imaging techniques has the potential to fulfill the goal of real-time noninvasive monitoring of the functional status and viability of the islet grafts. We review the application of a variety of imaging modalities for detecting endogenous and transplanted beta-cell mass. The review also explores the various molecular imaging strategies for assessing islet delivery, the metabolic effects on the islet grafts as well as detection of immunorejection. Here, we highlight the use of combined imaging and therapeutic interventions in islet transplantation and the in vivo monitoring of stem cells differentiation into insulin-producing cells.

Islet and Stem Cell Encapsulation for Clinical Transplantation

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Krishnan, Rahul; Alexander, Michael; Robles, Lourdes; Foster 3rd, Clarence E.; Lakey, Jonathan R.T.

2014-01-01

Over the last decade, improvements in islet isolation techniques have made islet transplantation an option for a certain subset of patients with long-standing diabetes. Although islet transplants have shown improved graft function, adequate function beyond the second year has not yet been demonstrated, and patients still require immunosuppression to prevent rejection. Since allogeneic islet transplants have experienced some success, the next step is to improve graft function while eliminating the need for systemic immunosuppressive therapy. Biomaterial encapsulation offers a strategy to avoid the need for toxic immunosuppression while increasing the chances of graft function and survival. Encapsulation entails coating cells or tissue in a semipermeable biocompatible material that allows for the passage of nutrients, oxygen, and hormones while blocking immune cells and regulatory substances from recognizing and destroying the cell, thus avoiding the need for systemic immunosuppressive therapy. Despite advances in encapsulation technology, these developments have not yet been meaningfully translated into clinical islet transplantation, for which several factors are to blame, including graft hypoxia, host inflammatory response, fibrosis, improper choice of biomaterial type, lack of standard guidelines, and post-transplantation device failure. Several new approaches, such as the use of porcine islets, stem cells, development of prevascularized implants, islet nanocoating, and multilayer encapsulation, continue to generate intense scientific interest in this rapidly expanding field. This review provides a comprehensive update on islet and stem cell encapsulation as a treatment modality in type 1 diabetes, including a historical outlook as well as current and future research avenues. PMID:25148368

A review of piscine islet xenotransplantation using wild-type tilapia donors and the production of transgenic tilapia expressing a “humanized” tilapia insulin

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Wright, James R; Yang, Hua; Hyrtsenko, Olga; Xu, Bao-You; Yu, Weiming; Pohajdak, Bill

2014-01-01

Most islet xenotransplantation laboratories have focused on porcine islets, which are both costly and difficult to isolate. Teleost (bony) fish, such as tilapia, possess macroscopically visible distinct islet organs called Brockmann bodies which can be inexpensively harvested. When transplanted into diabetic nude mice, tilapia islets maintain long-term normoglycemia and provide human-like glucose tolerance profiles. Like porcine islets, when transplanted into euthymic mice, they are rejected in a CD4 T-cell-dependent manner. However, unlike pigs, tilapia are so phylogenetically primitive that their cells do not express α(1,3)Gal and, because tilapia are highly evolved to live in warm stagnant waters nearly devoid of dissolved oxygen, their islet cells are exceedingly resistant to hypoxia, making them ideal for transplantation within encapsulation devices. Encapsulation, especially when combined with co-stimulatory blockade, markedly prolongs tilapia islet xenograft survival in small animal recipients, and a collaborator has shown function in diabetic cynomolgus monkeys. In anticipation of preclinical xenotransplantation studies, we have extensively characterized tilapia islets (morphology, embryologic development, cell biology, peptides, etc.) and their regulation of glucose homeostasis. Because tilapia insulin differs structurally from human insulin by 17 amino acids, we have produced transgenic tilapia whose islets stably express physiological levels of humanized insulin and have now bred these to homozygosity. These transgenic fish can serve as a platform for further development into a cell therapy product for diabetes. PMID:25040337

Effect of gamma-irradiation on mouse pancreatic islet-allograft survival

International Nuclear Information System (INIS)

Kanai, T.; Porter, J.; Gotoh, M.; Monaco, A.P.; Maki, T.

1989-01-01

Elimination or inactivation of lymphoid tissue in the pancreatic islet preparation achieves prolongation of islet-allograft survival. In this study we examined the effect of gamma-irradiation on mouse islet-allograft survival. In a B6AF1 isograft model, irradiation up to 2400 rad did not induce deterioration of islet function over 200 days, but greater doses caused cessation of graft function between 83 and 186 days. When DBA/2 crude islets were transplanted into B6AF1 recipients, all nonirradiated allografts were acutely rejected. Marked prolongation of allograft survival was achieved by islet irradiation with doses between 800 and 12,000 rad. With higher doses, significant numbers of allografts survived beyond the controls, but many lost function between 78 and 180 days, with none surviving greater than 200 days. Irradiation with 16,000 rad caused acute radiation damage. Because most secondary islet allografts in recipient mice that lost primary islet-graft function between 84 and 195 days survived greater than 100 days, late functional loss was probably due to the radiation injury. Combined use of recipient treatment with cyclosporin A and graft irradiation (2400 rad) achieved prolongation of DBA/2 islets in B6AF1 mice

Serotonin- and Dopamine-Related Gene Expression in db/db Mice Islets and in MIN6 β-Cells Treated with Palmitate and Oleate

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L. R. Cataldo

2016-01-01

Full Text Available High circulating nonesterified fatty acids (NEFAs concentration, often reported in diabetes, leads to impaired glucose-stimulated insulin secretion (GSIS through not yet well-defined mechanisms. Serotonin and dopamine might contribute to NEFA-dependent β-cell dysfunction, since extracellular signal of these monoamines decreases GSIS. Moreover, palmitate-treated β-cells may enhance the expression of the serotonin receptor Htr2c, affecting insulin secretion. Additionally, the expression of monoamine-oxidase type B (Maob seems to be lower in islets from humans and mice with diabetes compared to nondiabetic islets, which may lead to increased monoamine concentrations. We assessed the expression of serotonin- and dopamine-related genes in islets from db/db and wild-type (WT mice. In addition, the effect of palmitate and oleate on the expression of such genes, 5HT content, and GSIS in MIN6 β-cell was determined. Lower Maob expression was found in islets from db/db versus WT mice and in MIN6 β-cells in response to palmitate and oleate treatment compared to vehicle. Reduced 5HT content and impaired GSIS in response to palmitate (−25%; p<0.0001 and oleate (−43%; p<0.0001 were detected in MIN6 β-cells. In conclusion, known defects of GSIS in islets from db/db mice and MIN6 β-cells treated with NEFAs are accompanied by reduced Maob expression and reduced 5HT content.

Islet grafting and imaging in a bioengineered intramuscular space†

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Witkowski, Piotr; Sondermeijer, Hugo; Hardy, Mark A.; Woodland, David C.; Lee, Keagan; Bhagat, Govind; Witkowski, Kajetan; See, Fiona; Rana, Abbas; Maffei, Antonella; Itescu, Silviu; Harris, Paul E.

2011-01-01

Background Since the hepatic portal system may not be the optimal site for islet transplantation, several extrahepatic sites have been studied. Here we examine an intramuscular transplantation site, bioengineered to better support islet neovascularization, engraftment, and survival, and demonstrate that at this novel site, grafted beta cell mass may be quantitated in a real time non-invasive manner by PET imaging. Methods Streptozotocin induced rats were pretreated intramuscularly with a biocompatible angiogenic scaffold received syngeneic islet transplants 2 weeks later. The recipients were monitored serially by blood glucose and glucose tolerance measurements and by PET imaging of the transplant site with [11C] dihydrotetrabenazine. Parallel histopathologic evaluation of the grafts was done using insulin staining and evaluation of microvasularity. Results Reversal of hyperglycemia by islet transplantation was most successful in recipients pretreated with bioscaffolds containing angiogenic factors as compared to those who received no bioscaffolds or bioscaffolds not treated with angiogenic factors. PET imaging with [11C] dihydrotetrabenazine, insulin staining and microvascular density patterns were consistent with islet survival, increased levels of angiogenesis, and with reversal of hyperglycemia. Conclusions Induction of increased neovascularization at an intramuscular site significantly improves islet transplant engraftment and survival compared to controls. The use of a non hepatic transplant site may avoid intrahepatic complications and permit the use of PET imaging to measure and follow transplanted beta-cell mass in real time. These findings have important implications for effective islet implantation outside of the liver, and offer promising possibilities for improving islet survival, monitoring, and even prevention of islet loss. PMID:19898201

Islet grafting and imaging in a bioengineered intramuscular space.

Science.gov (United States)

Witkowski, Piotr; Sondermeijer, Hugo; Hardy, Mark A; Woodland, David C; Lee, Keagan; Bhagat, Govind; Witkowski, Kajetan; See, Fiona; Rana, Abbas; Maffei, Antonella; Itescu, Silviu; Harris, Paul E

2009-11-15

Because the hepatic portal system may not be the optimal site for islet transplantation, several extrahepatic sites have been studied. Here, we examine an intramuscular transplantation site, bioengineered to better support islet neovascularization, engraftment, and survival, and we demonstrate that at this novel site, grafted beta cell mass may be quantitated in a real-time noninvasive manner by positron emission tomography (PET) imaging. Streptozotocin-induced rats were pretreated intramuscularly with a biocompatible angiogenic scaffold received syngeneic islet transplants 2 weeks later. The recipients were monitored serially by blood glucose and glucose tolerance measurements and by PET imaging of the transplant site with [11C] dihydrotetrabenazine. Parallel histopathologic evaluation of the grafts was performed using insulin staining and evaluation of microvasularity. Reversal of hyperglycemia by islet transplantation was most successful in recipients pretreated with bioscaffolds containing angiogenic factors when compared with those who received no bioscaffolds or bioscaffolds not treated with angiogenic factors. PET imaging with [11C] dihydrotetrabenazine, insulin staining, and microvascular density patterns were consistent with islet survival, increased levels of angiogenesis, and with reversal of hyperglycemia. Induction of increased neovascularization at an intramuscular site significantly improves islet transplant engraftment and survival compared with controls. The use of a nonhepatic transplant site may avoid intrahepatic complications and permit the use of PET imaging to measure and follow transplanted beta cell mass in real time. These findings have important implications for effective islet implantation outside of the liver and offer promising possibilities for improving islet survival, monitoring, and even prevention of islet loss.

Who Should Be Considered for Islet Transplantation Alone?

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Othonos, Nantia; Choudhary, Pratik

2017-04-01

Episodic hypoglycemia is an almost inevitable consequence of exogenous insulin treatment of type 1 diabetes, and in up to 30% of patients, this can lead to impaired awareness of hypoglycemia. This predisposes to recurrent severe hypoglycemia and has a huge impact on quality of life. Although many patients can get resolution of severe hypoglycemia through novel education and technology, some patients continue to have ongoing life-threatening hypoglycemia. Islet transplantation offers an alternative therapeutic option for these patients, in whom these conventional approaches have been unsuccessful. This review discusses the selection process of identifying suitable candidates based on recent clinical data. Results from studies of islet transplantation suggest the optimal recipient characteristics for successful islet transplantation include age >35 years, insulin requirements 85 kg. Islet transplantation can completely resolve hypoglycemia and near-normalize glucose levels, achieving insulin independence for a limited period of time in up to 40% of patients. The selection of appropriate candidates, optimizing donor selection, the use of an optimized protocol for islet cell extraction, and immunosuppression therapy have been proved to be the key criteria for a favorable outcome in islet transplantation.

Homogenization of heterogeneously coupled bistable ODE's - applied to excitation waves in pancreatic islets of Langerhans

DEFF Research Database (Denmark)

Pedersen, Morten Gram

2004-01-01

We consider a lattice of coupled identical differential equations. The coupling is between nearest neighbors and of resistance type, but the strength of coupling varies from site to site. Such a lattice can, for example, model an islet of Langerhans, where the sites in the lattice model individua...

Hemiptera community and species responses to grassland sward islets

OpenAIRE

Helden, Alvin J.; Dittrich, Alex D. K.

2016-01-01

Sward islet is a term that has been used to describe a patch of longer vegetation in a pasture produced by a reduction in cattle grazing around their dung. They are known to affect the abundance and distribution of grassland arthropods. Hemiptera, like other groups, are found in higher densities within islets than the surrounding sward. Does this modify the community composition or is there just a density effect? Evidence from a paired (islets, non-islets) study at an Irish cattle-grazed site...

Experimental treatment of diabetic mice with microencapsulated rat islet cells transplantation

International Nuclear Information System (INIS)

Luo Yun; Xue Yilong; Li Yanling; Li Xinjian

2006-01-01

To observe treatment effects of diabetic mice with microcapsulated and non-microcapsulated rat islet cell transplantation, pancreas of SD rat was perfused with collagenase through cloledchus, and then the pancreatic tissues were isolated and digested. Histopaque-1077 was used to purify the digested pancreas. Islet cells were collected and implanted into the peritoneal cavity of diabetic mice. The isolated islets had a response upon glucose stimulation. When the microcapsulated islets and non- microcapsulated islets were transplanted into diabetic mices the high blood glucose level could be decreased to normal. The normal blood glucose level in the diabetic mice transpanted with microcapsulated islets could be maintained for over 30 days,but it could be mainlained only for 2-3 days in the diabetic mice transplanted with non-microcapsulated islets. Thus it is believed that microcapsulated islet cell transplantation exerts good effect on diabetic mice and the microcapsules possessed good immunoisolating function. (authors)

The F-actin modifier villin regulates insulin granule dynamics and exocytosis downstream of islet cell autoantigen 512

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Hassan Mziaut

2016-08-01

Full Text Available Objective: Insulin release from pancreatic islet β cells should be tightly controlled to avoid hypoglycemia and insulin resistance. The cortical actin cytoskeleton is a gate for regulated exocytosis of insulin secretory granules (SGs by restricting their mobility and access to the plasma membrane. Prior studies suggest that SGs interact with F-actin through their transmembrane cargo islet cell autoantigen 512 (Ica512 (also known as islet antigen 2/Ptprn. Here we investigated how Ica512 modulates SG trafficking and exocytosis. Methods: Transcriptomic changes in Ica512−/− mouse islets were analyzed. Imaging as well as biophysical and biochemical methods were used to validate if and how the Ica512-regulated gene villin modulates insulin secretion in mouse islets and insulinoma cells. Results: The F-actin modifier villin was consistently downregulated in Ica512−/− mouse islets and in Ica512-depleted insulinoma cells. Villin was enriched at the cell cortex of β cells and dispersed villin−/− islet cells were less round and less deformable. Basal mobility of SGs in villin-depleted cells was enhanced. Moreover, in cells depleted either of villin or Ica512 F-actin cages restraining cortical SGs were enlarged, basal secretion was increased while glucose-stimulated insulin release was blunted. The latter changes were reverted by overexpressing villin in Ica512-depleted cells, but not vice versa. Conclusion: Our findings show that villin controls the size of the F-actin cages restricting SGs and, thus, regulates their dynamics and availability for exocytosis. Evidence that villin acts downstream of Ica512 also indicates that SGs directly influence the remodeling properties of the cortical actin cytoskeleton for tight control of insulin secretion. Keywords: F-actin, Granules, Ica512, Insulin, Secretion, Villin

Spontaneous Hypoglycemia After Islet Autotransplantation for Chronic Pancreatitis.

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Lin, Yu Kuei; Faiman, Charles; Johnston, Philip C; Walsh, R Matthew; Stevens, Tyler; Bottino, Rita; Hatipoglu, Betul A

2016-10-01

Spontaneous hypoglycemia has been reported in patients after total pancreatectomy (TP) and islet autotransplantation (IAT) with maintained insulin independence. Details surrounding these events have not been well described. The objective of the study was to determine the frequency and characteristics of spontaneous hypoglycemia in patients undergoing TP-IAT and/or to ascertain predictive or protective factors of its development. This was an observational cohort study in 40 patients who underwent TP-IAT from August 2008 to May 2014, with a median follow-up of 34 months. The study was conducted at a single institution (Cleveland Clinic). Patients included recipients of TP-IAT. The intervention included small, frequent meals in those patients who developed spontaneous hypoglycemia. Incidence of spontaneous hypoglycemia development, characteristics of the patients developing hypoglycemia, and their response to small, frequent meals were measured. Six of 12 patients, who maintained insulin independence, developed spontaneous hypoglycemia. The episodes could be fasting, postprandial, and/or exercise associated, with the frequency ranging from two to three times daily to once every 1-2 weeks. All patients experienced at least one episode that required external assistance, glucagon administration, and/or emergent medical attention. Patients who developed hypoglycemia had a lower median age and tended to have a lower median islet equivalent/kg body weight but a higher median total islet equivalent, body mass index, and homeostatic model assessment for insulin resistance score. All patients who received small, frequent meal intervention had improvement in severity and/or frequency of the hypoglycemic episodes. Spontaneous hypoglycemia is prevalent after TP-IAT. Although the underlying pathophysiology responsible for these hypoglycemia events remains to be elucidated, small, frequent meal intervention is helpful in ameliorating this condition.

Improvement in Outcomes of Clinical Islet Transplantation: 1999–2010

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Barton, Franca B.; Rickels, Michael R.; Alejandro, Rodolfo; Hering, Bernhard J.; Wease, Stephen; Naziruddin, Bashoo; Oberholzer, Jose; Odorico, Jon S.; Garfinkel, Marc R.; Levy, Marlon; Pattou, Francois; Berney, Thierry; Secchi, Antonio; Messinger, Shari; Senior, Peter A.; Maffi, Paola; Posselt, Andrew; Stock, Peter G.; Kaufman, Dixon B.; Luo, Xunrong; Kandeel, Fouad; Cagliero, Enrico; Turgeon, Nicole A.; Witkowski, Piotr; Naji, Ali; O’Connell, Philip J.; Greenbaum, Carla; Kudva, Yogish C.; Brayman, Kenneth L.; Aull, Meredith J.; Larsen, Christian; Kay, Tom W.H.; Fernandez, Luis A.; Vantyghem, Marie-Christine; Bellin, Melena; Shapiro, A.M. James

2012-01-01

OBJECTIVE To describe trends of primary efficacy and safety outcomes of islet transplantation in type 1 diabetes recipients with severe hypoglycemia from the Collaborative Islet Transplant Registry (CITR) from 1999 to 2010. RESEARCH DESIGN AND METHODS A total of 677 islet transplant-alone or islet-after-kidney recipients with type 1 diabetes in the CITR were analyzed for five primary efficacy outcomes and overall safety to identify any differences by early (1999–2002), mid (2003–2006), or recent (2007–2010) transplant era based on annual follow-up to 5 years. RESULTS Insulin independence at 3 years after transplant improved from 27% in the early era (1999–2002, n = 214) to 37% in the mid (2003–2006, n = 255) and to 44% in the most recent era (2007–2010, n = 208; P = 0.006 for years-by-era; P = 0.01 for era alone). C-peptide ≥0.3 ng/mL, indicative of islet graft function, was retained longer in the most recent era (P islet reinfusion rate was lower: 48% by 1 year in 2007–2010 vs. 60–65% in 1999–2006 (P islet graft function (P islet transplantation in recipients who received transplants in 2007–2010 compared with those in 1999–2006, with fewer islet infusions and adverse events per recipient. PMID:22723582

PD-L1 Deficiency within Islets Reduces Allograft Survival in Mice.

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Dongxia Ma

Full Text Available Islet transplantation may potentially cure type 1 diabetes mellitus (T1DM. However, immune rejection, especially that induced by the alloreactive T-cell response, remains a restraining factor for the long-term survival of grafted islets. Programmed death ligand-1 (PD-L1 is a negative costimulatory molecule. PD-L1 deficiency within the donor heart accelerates allograft rejection. Here, we investigate whether PD-L1 deficiency in donor islets reduces allograft survival time.Glucose Stimulation Assays were performed to evaluate whether PD-L1 deficiency has detrimental effects on islet function. Islets isolated from PDL1-deficient mice or wild- type (WT mice (C57BL/6j were implanted beneath the renal capsule of streptozotocin (STZ-induced diabetic BALB/c mice. Blood glucose levels and graft survival time after transplantation were monitored. Moreover, we analyzed the residual islets, infiltrating immune cells and alloreactive cells from the recipients.PD-L1 deficiency within islets does not affect islet function. However, islet PD-L1 deficiency increased allograft rejection and was associated with enhanced inflammatory cell infiltration and recipient T-cell alloreactivity.This is the first report to demonstrate that PD-L1 deficiency accelerated islet allograft rejection and regulated recipient alloimmune responses.

Total pancreatectomy and islet autotransplantation for chronic pancreatitis.

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Sutherland, David E R; Radosevich, David M; Bellin, Melena D; Hering, Bernard J; Beilman, Gregory J; Dunn, Ty B; Chinnakotla, Srinath; Vickers, Selwyn M; Bland, Barbara; Balamurugan, A N; Freeman, Martin L; Pruett, Timothy L

2012-04-01

Total pancreatectomy (TP) with intraportal islet autotransplantation (IAT) can relieve pain and preserve β-cell mass in patients with chronic pancreatitis (CP) when other therapies fail. We report on a >30-year single-center series. Four hundred and nine patients (including 53 children, 5 to 18 years) with CP underwent TP-IAT from February 1977 to September 2011 (etiology: idiopathic, 41%; Sphincter of Oddi dysfunction/biliary, 9%; genetic, 14%; divisum, 17%; alcohol, 7%; and other, 12%; mean age was 35.3 years, 74% were female; 21% has earlier operations, including 9% Puestow procedure, 6% Whipple, 7% distal pancreatectomy, and 2% other). Islet function was classified as insulin independent for those on no insulin; partial, if known C-peptide positive or euglycemic on once-daily insulin; and insulin dependent if on standard basal-bolus diabetic regimen. A 36-item Short Form (SF-36) survey for quality of life was completed by patients before and in serial follow-up since 2007, with an integrated survey that was added in 2008. Actuarial patient survival post TP-IAT was 96% in adults and 98% in children (1 year) and 89% and 98% (5 years). Complications requiring relaparotomy occurred in 15.9% and bleeding (9.5%) was the most common complication. IAT function was achieved in 90% (C-peptide >0.6 ng/mL). At 3 years, 30% were insulin independent (25% in adults, 55% in children) and 33% had partial function. Mean hemoglobin A1c was 5,000/kg [24%]) correlated with degree of function with insulin-independent rates at 3 years of 12%, 22%, and 72%, and rates of partial function 33%, 62%, and 24%. All patients had pain before TP-IAT and nearly all were on daily narcotics. After TP-IAT, 85% had pain improvement. By 2 years, 59% had ceased narcotics. All children were on narcotics before, 39% at follow-up; pain improved in 94%; and 67% became pain-free. In the SF-36 survey, there was significant improvement from baseline in all dimensions, including the Physical and Mental

Prediction of Marginal Mass Required for Successful Islet Transplantation

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Papas, Klearchos K.; Colton, Clark K.; Qipo, Andi; Wu, Haiyan; Nelson, Rebecca A.; Hering, Bernhard J.; Weir, Gordon C.; Koulmanda, Maria

2013-01-01

Islet quality assessment methods for predicting diabetes reversal (DR) following transplantation are needed. We investigated two islet parameters, oxygen consumption rate (OCR) and OCR per DNA content, to predict transplantation outcome and explored the impact of islet quality on marginal islet mass for DR. Outcomes in immunosuppressed diabetic mice were evaluated by transplanting mixtures of healthy and purposely damaged rat islets for systematic variation of OCR/DNA over a wide range. The probability of DR increased with increasing transplanted OCR and OCR/DNA. On coordinates of OCR versus OCR/DNA, data fell into regions in which DR occurred in all, some, or none of the animals with a sharp threshold of around 150-nmol/min mg DNA. A model incorporating both parameters predicted transplantation outcome with sensitivity and specificity of 93% and 94%, respectively. Marginal mass was not constant, depended on OCR/DNA, and increased from 2,800 to over 100,000 islet equivalents/kg body weight as OCR/DNA decreased. We conclude that measurements of OCR and OCR/DNA are useful for predicting transplantation outcome in this model system, and OCR/DNA can be used to estimate the marginal mass required for reversing diabetes. Because human clinical islet preparations in a previous study had OCR/DNA values in the range of 100–150-nmol/min mg DNA, our findings suggest that substantial improvement in transplantation outcome may accompany increasedOCR/DNAin clinical islet preparations. PMID:20233002

Distinct cell clusters touching islet cells induce islet cell replication in association with over-expression of Regenerating Gene (REG protein in fulminant type 1 diabetes.

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Kaoru Aida

Full Text Available BACKGROUND: Pancreatic islet endocrine cell-supporting architectures, including islet encapsulating basement membranes (BMs, extracellular matrix (ECM, and possible cell clusters, are unclear. PROCEDURES: The architectures around islet cell clusters, including BMs, ECM, and pancreatic acinar-like cell clusters, were studied in the non-diabetic state and in the inflamed milieu of fulminant type 1 diabetes in humans. RESULT: Immunohistochemical and electron microscopy analyses demonstrated that human islet cell clusters and acinar-like cell clusters adhere directly to each other with desmosomal structures and coated-pit-like structures between the two cell clusters. The two cell-clusters are encapsulated by a continuous capsule composed of common BMs/ECM. The acinar-like cell clusters have vesicles containing regenerating (REG Iα protein. The vesicles containing REG Iα protein are directly secreted to islet cells. In the inflamed milieu of fulminant type 1 diabetes, the acinar-like cell clusters over-expressed REG Iα protein. Islet endocrine cells, including beta-cells and non-beta cells, which were packed with the acinar-like cell clusters, show self-replication with a markedly increased number of Ki67-positive cells. CONCLUSION: The acinar-like cell clusters touching islet endocrine cells are distinct, because the cell clusters are packed with pancreatic islet clusters and surrounded by common BMs/ECM. Furthermore, the acinar-like cell clusters express REG Iα protein and secrete directly to neighboring islet endocrine cells in the non-diabetic state, and the cell clusters over-express REG Iα in the inflamed milieu of fulminant type 1 diabetes with marked self-replication of islet cells.

Biobreeding rat islets exhibit reduced antioxidative defense and N-acetyl cysteine treatment delays type 1 diabetes

Science.gov (United States)

Bogdani, Marika; Henschel, Angela M.; Kansra, Sanjay; Fuller, Jessica M.; Geoffrey, Rhonda; Jia, Shuang; Kaldunski, Mary L.; Pavletich, Scott; Prosser, Simon; Chen, Yi-Guang; Lernmark, Åke; Hessner, Martin J.

2014-01-01

Islet-level oxidative stress has been proposed as a trigger for type 1 diabetes (T1D), and release of cytokines by infiltrating immune cells further elevates reactive oxygen species (ROS), exacerbating β cell duress. To identify genes/mechanisms involved with diabeto-genesis at the β cell level, gene expression profiling and targeted follow-up studies were used to investigate islet activity in the biobreeding (BB) rat. Forty-day-old spontaneously diabetic lymphopenic BB DRlyp/lyp rats (before T cell insulitis) as well as nondiabetic BB DR+/+ rats, nondiabetic but lymphopenic F344lyp/lyp rats, and healthy Fischer (F344) rats were examined. Gene expression profiles of BB rat islets were highly distinct from F344 islets and under-expressed numerous genes involved in ROS metabolism, including glutathione S-transferase (GST) family members (Gstm2, Gstm4, Gstm7, Gstt1, Gstp1, and Gstk1), superoxide dismutases (Sod2 and Sod3), peroxidases, and peroxiredoxins. This pattern of under-expression was not observed in brain, liver, or muscle. Compared with F344 rats, BB rat pancreata exhibited lower GST protein levels, while plasma GST activity was found significantly lower in BB rats. Systemic administration of the antioxidant N-acetyl cysteine to DRlyp/lyp rats altered abundances of peripheral eosinophils, reduced severity of insulitis, and significantly delayed but did not prevent diabetes onset. We find evidence of β cell dysfunction in BB rats independent of T1D progression, which includes lower expression of genes related to antioxidative defense mechanisms during the pre-onset period that may contribute to overall T1D susceptibility. PMID:23111281

Photoacoustic imaging of angiogenesis in subdermal islet transplant sites

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Shi, Wei; Pawlick, Rena; Bruni, Antonio; Rafiei, Yasmin; Pepper, Andrew R.; Gala-Lopez, Boris; Choi, Min; Malcolm, Andrew; Zemp, Roger J.; Shapiro, A. M. James

2016-03-01

Exogenous insulin administration is the mainstay treatment therapy for patients with Type-1 diabetes mellitus (T1DM). However, for select patients, clinical islet transplantation is an alternative therapeutic treatment. In this procedure, islets are transplanted into the hepatic portal vein, and despite improved success within the last decade, obstacles are still associated with this approach. It has been discovered that the subcutaneous space may be an effective alternative site for islet transplantation, and may provide advantages of easy access and potential for simple monitoring. The ability to monitor islet viability and the transplant microenvironment may be key to future success in islet transplantation. A subcutaneous device-less technique has been developed to facilitate angiogenesis in the islet transplant site, however, a method for monitoring the potential engraftment site have yet to be explored fully. Here we demonstrate the ability to track angiogenesis in mice with 1, 2, 3 and 4 weeks post-catheter implant on both sides of the abdomen using a FujiFilm VisualSonics Vevo-LAZR system. Quantitative analysis on vessel densities exhibited gradual vessel growth successfully induced by catheter implantation. Our study demonstrates the ability of employing photoacoustic and micro-ultrasound imaging to track angiogenesis around the catheter site prior to islet transplantation.

Magnetic separation of encapsulated islet cells labeled with superparamagnetic iron oxide nano particles.

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Mettler, Esther; Trenkler, Anja; Feilen, Peter J; Wiegand, Frederik; Fottner, Christian; Ehrhart, Friederike; Zimmermann, Heiko; Hwang, Yong Hwa; Lee, Dong Yun; Fischer, Stefan; Schreiber, Laura M; Weber, Matthias M

2013-01-01

Islet cell transplantation is a promising option for the restoration of normal glucose homeostasis in patients with type 1 diabetes. Because graft volume is a crucial issue in islet transplantations for patients with diabetes, we evaluated a new method for increasing functional tissue yield in xenogeneic grafts of encapsulated islets. Islets were labeled with three different superparamagnetic iron oxide nano particles (SPIONs; dextran-coated SPION, siloxane-coated SPION, and heparin-coated SPION). Magnetic separation was performed to separate encapsulated islets from the empty capsules, and cell viability and function were tested. Islets labeled with 1000 μg Fe/ml dextran-coated SPIONs experienced a 69.9% reduction in graft volume, with a 33.2% loss of islet-containing capsules. Islets labeled with 100 μg Fe/ml heparin-coated SPIONs showed a 46.4% reduction in graft volume, with a 4.5% loss of capsules containing islets. No purification could be achieved using siloxane-coated SPIONs due to its toxicity to the primary islets. SPION labeling of islets is useful for transplant purification during islet separation as well as in vivo imaging after transplantation. Furthermore, purification of encapsulated islets can also reduce the volume of the encapsulated islets without impairing their function by removing empty capsules. © 2013 John Wiley & Sons A/S.

Phosphatidylcholine (PC) biosynthesis in pancreatic islets of Langerhans

International Nuclear Information System (INIS)

Hoffman, J.M.; Laychock, S.G.

1986-01-01

Islets of Langerhans isolated from rat pancreata were incubated with [ 14 C]choline to determine the biosynthesis of PC by the CDP choline to determine the biosynthesis of PC by the CDPcholine pathway. Recovery of [ 14 C]PC in islet membranes was time-related, and stimulated by glucose (17mM) during 60 min. The rate of PC synthesis was constant during 60 min with glucose stimulation. In contrast, the sulfonylurea tolbutamide (2 mM) reduced the recovery of [ 14 C]choline in PC, and 8-bromo-cyclic AMP (5 mM) did not significantly affect [ 14 C]PC recovery. Incubation of islets in Ca 2+ -free medium enhanced glucose-stimulated recovery of [ 14 C]choline-labeled PC due to the inhibition of phospholipase and phospholipid hydrolysis. Inhibition of CTP:phosphocholine cytidylyltransferase with 5-deoxy-5'-isobutylthioadenosine (SIBA) reduced [ 14 C]PC levels and insulin release in a concentration dependent manner. Treatment with SIBA also reduced Mg 2+ -dependent Ca 2+ -ATPase activity in islet microsomes. Quantitation of membrane PC showed that glucose stimulation did not alter islet P levels. Thus, islet PC biosynthesis is linked to glucose stimulation and contributes to the maintenance of PC levels in membranes undergoing exocytosis and phospholipid hydrolysis. Adequate PC levels support Ca 2+ pump activity and secretory mechanisms

Is islet transplantation a realistic approach to curing diabetes?

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Jin, Sang-Man; Kim, Kwang-Won

2017-01-01

Since the report of type 1 diabetes reversal in seven consecutive patients by the Edmonton protocol in 2000, pancreatic islet transplantation has been reappraised based on accumulated clinical evidence. Although initially expected to therapeutically target long-term insulin independence, islet transplantation is now indicated for more specific clinical benefits. With the long-awaited report of the first phase 3 clinical trial in 2016, allogeneic islet transplantation is now transitioning from an experimental to a proven therapy for type 1 diabetes with problematic hypoglycemia. Islet autotransplantation has already been therapeutically proven in chronic pancreatitis with severe abdominal pain refractory to conventional treatments, and it holds promise for preventing diabetes after partial pancreatectomy due to benign pancreatic tumors. Based on current evidence, this review focuses on islet transplantation as a realistic approach to treating diabetes.

Extracellular Matrix and Growth Factors Improve the Efficacy of Intramuscular Islet Transplantation.

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Tsuchiya, Haruyuki; Sakata, Naoaki; Yoshimatsu, Gumpei; Fukase, Masahiko; Aoki, Takeshi; Ishida, Masaharu; Katayose, Yu; Egawa, Shinichi; Unno, Michiaki

2015-01-01

The efficacy of intramuscular islet transplantation is poor despite being technically simple, safe, and associated with reduced rates of severe complications. We evaluated the efficacy of combined treatment with extracellular matrix (ECM) and growth factors in intramuscular islet transplantation. Male BALB/C mice were used for the in vitro and transplantation studies. The following three groups were evaluated: islets without treatment (islets-only group), islets embedded in ECM with growth factors (Matrigel group), and islets embedded in ECM without growth factors [growth factor-reduced (GFR) Matrigel group]. The viability and insulin-releasing function of islets cultured for 96 h were significantly improved in Matrigel and GFR Matrigel groups compared with the islets-only group. Blood glucose and serum insulin levels immediately following transplantation were significantly improved in the Matrigel and GFR Matrigel groups and remained significantly improved in the Matrigel group at postoperative day (POD) 28. On histological examination, significantly decreased numbers of TdT-mediated deoxyuridine triphosphate-biotin nick end labeling-positive islet cells and significantly increased numbers of Ki67-positive cells were observed in the Matrigel and GFR Matrigel groups at POD 3. Peri-islet revascularization was most prominent in the Matrigel group at POD 14. The efficacy of intramuscular islet transplantation was improved by combination treatment with ECM and growth factors through the inhibition of apoptosis, increased proliferation of islet cells, and promotion of revascularization.

Prolonged survival of isolated rat islet allografts pre-irradiated with X-rays

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Kimura, Toshihisa; Note, Masayuki; Nakagawara, Gizo (Fukui Medical School, Matsuoka (Japan)); Kojima, Yasuhiko

1994-04-01

Prior to transplantation of islets, pre-incubation, or pre-irradiation may suppress the immunogenicity of islet cells without suppressing islet function. In the presently described experiments we investigated the use of X-ray irradiation prior to transplantation to reduce islet immunogenicity. To determine whether or not the islet function was reduced after irradiation, ACI rat islets were transplanted into the subrenal capsules of isogeneic rats which had been diabetic and examined the blood glucose level over a period of 40 days. The results indicated that irradiation injury was dose-dependent and that islets irradiated with over 80 Gy lost their function. Next, allogeneic transplantation was performed using the model of ACI rats to Lewis rats without the use of any immunosuppressive agent. Non-irradiated islets were rejected within 7 days. However 20 Gy or 40 Gy irradiated islets prolonged survival (18.7[+-]5.8 days (n=6) and 26.7[+-]10.0 days (n=6), respectively). To determine the basis for this effect, MHC expression of islets was examined by the immunoperoxidase technique. Immunohistologic studies showed that 40 Gy-irradiated islets were depleted of Class II antigen positive cells while Class I antigen expression was unchanged. These results suggest that the prolongation of islets survival by X-ray irradiation may possibly be due to, in part, the depletion of donor Class II antigen positive cells. (author).

Autologous Mesenchymal Stem Cell and Islet Cotransplantation: Safety and Efficacy.

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Wang, Hongjun; Strange, Charlie; Nietert, Paul J; Wang, Jingjing; Turnbull, Taylor L; Cloud, Colleen; Owczarski, Stefanie; Shuford, Betsy; Duke, Tara; Gilkeson, Gary; Luttrell, Louis; Hermayer, Kathie; Fernandes, Jyotika; Adams, David B; Morgan, Katherine A

2018-01-01

Islet engraftment after transplantation is impaired by high rates of islet/β cell death caused by cellular stressors and poor graft vascularization. We studied whether cotransplantation of ex vivo expanded autologous bone marrow-derived mesenchymal stem cells (MSCs) with islets is safe and beneficial in chronic pancreatitis patients undergoing total pancreatectomy with islet autotransplantation. MSCs were harvested from the bone marrow of three islet autotransplantation patients and expanded at our current Good Manufacturing Practices (cGMP) facility. On the day of islet transplantation, an average dose of 20.0 ± 2.6 ×10 6 MSCs was infused with islets via the portal vein. Adverse events and glycemic control at baseline, 6, and 12 months after transplantation were compared with data from 101 historical control patients. No adverse events directly related to the MSC infusions were observed. MSC patients required lower amounts of insulin during the peritransplantation period (p = .02 vs. controls) and had lower 12-month fasting blood glucose levels (p = .02 vs. controls), smaller C-peptide declines over 6 months (p = .01 vs. controls), and better quality of life compared with controls. In conclusion, our pilot study demonstrates that autologous MSC and islet cotransplantation may be a safe and potential strategy to improve islet engraftment after transplantation. (Clinicaltrials.gov registration number: NCT02384018). Stem Cells Translational Medicine 2018;7:11-19. © 2017 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Induction of Protective Genes Leads to Islet Survival and Function

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Hongjun Wang

2011-01-01

Full Text Available Islet transplantation is the most valid approach to the treatment of type 1 diabetes. However, the function of transplanted islets is often compromised since a large number of β cells undergo apoptosis induced by stress and the immune rejection response elicited by the recipient after transplantation. Conventional treatment for islet transplantation is to administer immunosuppressive drugs to the recipient to suppress the immune rejection response mounted against transplanted islets. Induction of protective genes in the recipient (e.g., heme oxygenase-1 (HO-1, A20/tumor necrosis factor alpha inducible protein3 (tnfaip3, biliverdin reductase (BVR, Bcl2, and others or administration of one or more of the products of HO-1 to the donor, the islets themselves, and/or the recipient offers an alternative or synergistic approach to improve islet graft survival and function. In this perspective, we summarize studies describing the protective effects of these genes on islet survival and function in rodent allogeneic and xenogeneic transplantation models and the prevention of onset of diabetes, with emphasis on HO-1, A20, and BVR. Such approaches are also appealing to islet autotransplantation in patients with chronic pancreatitis after total pancreatectomy, a procedure that currently only leads to 1/3 of transplanted patients being diabetes-free.

Fibroblasts accelerate islet revascularization and improve long-term graft survival in a mouse model of subcutaneous islet transplantation.

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Marcos Perez-Basterrechea

Full Text Available Pancreatic islet transplantation has been considered for many years a promising therapy for beta-cell replacement in patients with type-1 diabetes despite that long-term clinical results are not as satisfactory. This fact points to the necessity of designing strategies to improve and accelerate islets engraftment, paying special attention to events assuring their revascularization. Fibroblasts constitute a cell population that collaborates on tissue homeostasis, keeping the equilibrium between production and degradation of structural components as well as maintaining the required amount of survival factors. Our group has developed a model for subcutaneous islet transplantation using a plasma-based scaffold containing fibroblasts as accessory cells that allowed achieving glycemic control in diabetic mice. Transplanted tissue engraftment is critical during the first days after transplantation, thus we have gone in depth into the graft-supporting role of fibroblasts during the first ten days after islet transplantation. All mice transplanted with islets embedded in the plasma-based scaffold reversed hyperglycemia, although long-term glycemic control was maintained only in the group transplanted with the fibroblasts-containing scaffold. By gene expression analysis and histology examination during the first days we could conclude that these differences might be explained by overexpression of genes involved in vessel development as well as in β-cell regeneration that were detected when fibroblasts were present in the graft. Furthermore, fibroblasts presence correlated with a faster graft re-vascularization, a higher insulin-positive area and a lower cell death. Therefore, this work underlines the importance of fibroblasts as accessory cells in islet transplantation, and suggests its possible use in other graft-supporting strategies.

Fibroblasts accelerate islet revascularization and improve long-term graft survival in a mouse model of subcutaneous islet transplantation.

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Perez-Basterrechea, Marcos; Esteban, Manuel Martinez; Alvarez-Viejo, Maria; Fontanil, Tania; Cal, Santiago; Sanchez Pitiot, Marta; Otero, Jesus; Obaya, Alvaro Jesus

2017-01-01

Pancreatic islet transplantation has been considered for many years a promising therapy for beta-cell replacement in patients with type-1 diabetes despite that long-term clinical results are not as satisfactory. This fact points to the necessity of designing strategies to improve and accelerate islets engraftment, paying special attention to events assuring their revascularization. Fibroblasts constitute a cell population that collaborates on tissue homeostasis, keeping the equilibrium between production and degradation of structural components as well as maintaining the required amount of survival factors. Our group has developed a model for subcutaneous islet transplantation using a plasma-based scaffold containing fibroblasts as accessory cells that allowed achieving glycemic control in diabetic mice. Transplanted tissue engraftment is critical during the first days after transplantation, thus we have gone in depth into the graft-supporting role of fibroblasts during the first ten days after islet transplantation. All mice transplanted with islets embedded in the plasma-based scaffold reversed hyperglycemia, although long-term glycemic control was maintained only in the group transplanted with the fibroblasts-containing scaffold. By gene expression analysis and histology examination during the first days we could conclude that these differences might be explained by overexpression of genes involved in vessel development as well as in β-cell regeneration that were detected when fibroblasts were present in the graft. Furthermore, fibroblasts presence correlated with a faster graft re-vascularization, a higher insulin-positive area and a lower cell death. Therefore, this work underlines the importance of fibroblasts as accessory cells in islet transplantation, and suggests its possible use in other graft-supporting strategies.

A preclinical evaluation of alternative site for islet allotransplantation.

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Chengshi Wang

Full Text Available The bone marrow cavity (BMC has recently been identified as an alternative site to the liver for islet transplantation. This study aimed to compare the BMC with the liver as an islet allotransplantation site in diabetic monkeys. Diabetes was induced in Rhesus monkeys using streptozocin, and the monkeys were then divided into the following three groups: Group1 (islets transplanted in the liver with immunosuppressant, Group 2 (islets transplanted in the tibial BMC, and Group 3 (islets transplanted in the tibial BMC with immunosuppressant. The C-peptide and blood glucose levels were preoperatively measured. An intravenous glucose tolerance test (IVGTT was conducted to assess graft function, and complete blood cell counts were performed to assess cell population changes. Cytokine expression was measured using an enzyme-linked immune sorbent assay (ELISA and MILLIPLEX. Five monkeys in Group 3 exhibited a significantly increased insulin-independent time compared with the other groups (Group 1: 78.2 ± 19.0 days; Group 2: 58.8 ± 17.0 days; Group 3: 189.6 ± 26.2 days and demonstrated increases in plasma C-peptide 4 months after transplantation. The infusion procedure was not associated with adverse effects. Functional islets in the BMC were observed 225 days after transplantation using the dithizone (DTZ and insulin/glucagon stains. Our results showed that allogeneic islets transplanted in the BMC of diabetic Rhesus monkeys remained alive and functional for a longer time than those transplanted in the liver. This study was the first successful demonstration of allogeneic islet engraftment in the BMC of non-human primates (NHPs.

Regulation of the JNK3 signaling pathway during islet isolation: JNK3 and c-fos as new markers of islet quality for transplantation.

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Saida Abdelli

Full Text Available Stress conditions generated throughout pancreatic islet processing initiate the activation of pro-inflammatory pathways and beta-cell destruction. Our goal is to identify relevant and preferably beta-specific markers to assess the activation of beta-cell stress and apoptotic mechanisms, and therefore the general quality of the islet preparation prior to transplantation. Protein expression and activation were analyzed by Western blotting and kinase assays. ATP measurements were performed by a luminescence-based assay. Oxygen consumption rate (OCR was measured based on standard protocols using fiber optic sensors. Total RNA was used for gene expression analyzes. Our results indicate that pancreas digestion initiates a potent stress response in the islets by activating two stress kinases, c-Jun N-terminal Kinase (JNK and p38. JNK1 protein levels remained unchanged between different islet preparations and following culture. In contrast, levels of JNK3 increased after islet culture, but varied markedly, with a subset of preparations bearing low JNK3 expression. The observed changes in JNK3 protein content strongly correlated with OCR measurements as determined by the Spearman's rank correlation coefficient rho [Formula: see text] in the matching islet samples, while inversely correlating with c-fos mRNA expression [Formula: see text]. In conclusion, pancreas digestion recruits JNK and p38 kinases that are known to participate to beta-cell apoptosis. Concomitantly, the islet isolation alters JNK3 and c-fos expression, both strongly correlating with OCR. Thus, a comparative analysis of JNK3 and c-fos expression before and after culture may provide for novel markers to assess islet quality prior to transplantation. JNK3 has the advantage over all other proposed markers to be islet-specific, and thus to provide for a marker independent of non-beta cell contamination.

Factors influencing insulin secretion from encapsulated islets

NARCIS (Netherlands)

de Haan, BJ; Faas, MM; de Vos, P

2003-01-01

Adequate regulation of glucose levels by a microencapsulated pancreatic islet graft requires a minute-to-minute regulation of blood glucose. To design such a transplant, it is mandatory to have sufficient insight in factors influencing the kinetics of insulin secretion by encapsulated islets. The

Assimilating Dokdo: The Islets in Korean Everyday Life

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Brandon Palmer

2016-03-01

Full Text Available Sovereignty over the Tokto Islets is heatedly contested between South Korea and Japan. The Korean government and citizenry have responded to this dispute by inserting the islets into their national collective memory in multifarious ways in an attempt to strengthen their nation’s claim to Tokto. The islets are included in the material culture and public memory of the nation in ways that make them part of everyday life for millions of Koreans. Korea’s claim to Tokto is currently taught in schools, presented in museums, found in popular songs, and exploited by businesses for profit. The deeper Tokto becomes entrenched in Korean society, the less likely a compromise can be reached with Japan over the islets.

Protein-Mediated Interactions of Pancreatic Islet Cells

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Paolo Meda

2013-01-01

Full Text Available The islets of Langerhans collectively form the endocrine pancreas, the organ that is soley responsible for insulin secretion in mammals, and which plays a prominent role in the control of circulating glucose and metabolism. Normal function of these islets implies the coordination of different types of endocrine cells, noticeably of the beta cells which produce insulin. Given that an appropriate secretion of this hormone is vital to the organism, a number of mechanisms have been selected during evolution, which now converge to coordinate beta cell functions. Among these, several mechanisms depend on different families of integral membrane proteins, which ensure direct (cadherins, N-CAM, occludin, and claudins and paracrine communications (pannexins between beta cells, and between these cells and the other islet cell types. Also, other proteins (integrins provide communication of the different islet cell types with the materials that form the islet basal laminae and extracellular matrix. Here, we review what is known about these proteins and their signaling in pancreatic β-cells, with particular emphasis on the signaling provided by Cx36, given that this is the integral membrane protein involved in cell-to-cell communication, which has so far been mostly investigated for effects on beta cell functions.

A novel method for murine intrahepatic islet transplantation via cecal vein.

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Byun, Nari; Kim, Hyun-Je; Min, Byoung-Hoon; Shin, Jun-Seop; Yoon, Il-Hee; Kim, Jong-Min; Kim, Yong-Hee; Park, Chung-Gyu

2015-12-01

Islet transplantation is one of the most beneficial treatment modality to treat type 1 diabetic patients with frequent hypoglycemic unawareness. In clinical setting, human islets are infused via portal vein and are settled in the end-portal venules in the liver. However, mouse islets are transplanted into kidney subcapsule or liver through direct portal vein. These conventional transplantation methods have several drawbacks such as different physiological environments around the transplanted islets in kidney subcapsule from the liver and high mortality rate in direct portal vein approach. In this study, we introduced murine intrahepatic islet transplantation method via cecal vein to have the same surgical operation route in humans as well as guaranteeing low mortality rate after islet transplantation. With this protocol, consistent normoglycemia can be obtained in diabetic mice, while keeping operation-related mortality extremely low. This approach with easier accessibility and low mortality will make murine intrahepatic islet transplantation a useful model for studying immunological mechanisms such as strong innate and adaptive immune responses that occur in human islet transplantation. Copyright © 2015 Elsevier B.V. All rights reserved.

Nonhuman Primate Models of Type 1 Diabetes Mellitus for Islet Transplantation

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Haitao Zhu

2014-01-01

Full Text Available Islet transplantation is an attractive treatment of type 1 diabetes mellitus (T1DM. Animal models of diabetes mellitus (DM contribute a lot to the experimental studies of islet transplantation and to evaluations of isolated islet grafts for future clinical applications. Diabetic nonhuman primates (NHPs represent the suitable models of DMs to better evaluate the effectiveness of islet transplantation, to assess new strategies for controlling blood glucose (BG, relieving immune rejection, or prolonging islet survival, and eventually to translate the preclinical data into tangible clinical practice. This review introduces some NHP models of DM, clarifies why and how the models should be used, and elucidates the usefulness and limitations of the models in islet transplantation.

High fat programming of beta cell compensation, exhaustion, death and dysfunction.

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Cerf, Marlon E

2015-03-01

Programming refers to events during critical developmental windows that shape progeny health outcomes. Fetal programming refers to the effects of intrauterine (in utero) events. Lactational programming refers to the effects of events during suckling (weaning). Developmental programming refers to the effects of events during both fetal and lactational life. Postnatal programming refers to the effects of events either from birth (lactational life) to adolescence or from weaning (end of lactation) to adolescence. Islets are most plastic during the early life course; hence programming during fetal and lactational life is most potent. High fat (HF) programming is the maintenance on a HF diet (HFD) during critical developmental life stages that alters progeny metabolism and physiology. HF programming induces variable diabetogenic phenotypes dependent on the timing and duration of the dietary insult. Maternal obesity reinforces HF programming effects in progeny. HF programming, through acute hyperglycemia, initiates beta cell compensation. However, HF programming eventually leads to chronic hyperglycemia that triggers beta cell exhaustion, death and dysfunction. In HF programming, beta cell dysfunction often co-presents with insulin resistance. Balanced, healthy nutrition during developmental windows is critical for preserving beta cell structure and function. Thus early positive nutritional interventions that coincide with the development of beta cells may reduce the overwhelming burden of diabetes and metabolic disease. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Mitochondrial dysfunction enhances cisplatin resistance in human gastric cancer cells via the ROS-activated GCN2-eIF2α-ATF4-xCT pathway.

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Wang, Sheng-Fan; Chen, Meng-Shian; Chou, Yueh-Ching; Ueng, Yune-Fang; Yin, Pen-Hui; Yeh, Tien-Shun; Lee, Hsin-Chen

2016-11-08

Mitochondrial DNA mutations and defects in mitochondrial enzymes have been identified in gastric cancers, and they might contribute to cancer progression. In previous studies, mitochondrial dysfunction was induced by oligomycin-enhanced chemoresistance to cisplatin. Herein, we dissected the regulatory mechanism for mitochondrial dysfunction-enhanced cisplatin resistance in human gastric cancer cells. Repeated cisplatin treatment-induced cisplatin-resistant cells exhibited high SLC7A11 (xCT) expression, and xCT inhibitors (sulfasalazine or erastin), xCT siRNA, or a GSH synthesis inhibitor (buthionine sulphoximine, BSO) could sensitize these cells to cisplatin. Clinically, the high expression of xCT was associated with a poorer prognosis for gastric cancer patients under adjuvant chemotherapy. Moreover, we found that mitochondrial dysfunction enhanced cisplatin resistance and up-regulated xCT expression, as well as intracellular glutathione (GSH). The xCT inhibitors, siRNA against xCT or BSO decreased mitochondrial dysfunction-enhanced cisplatin resistance. We further demonstrated that the upregulation of the eIF2α-ATF4 pathway contributed to mitochondrial dysfunction-induced xCT expression, and activated eIF2α kinase GCN2, but not PERK, stimulated the eIF2α-ATF4-xCT pathway in response to mitochondrial dysfunction-increased reactive oxygen species (ROS) levels. In conclusion, our results suggested that the ROS-activated GCN2-eIF2α-ATF4-xCT pathway might contribute to mitochondrial dysfunction-enhanced cisplatin resistance and could be a potential target for gastric cancer therapy.

Extracellular Matrix and Growth Factors Improve the Efficacy of Intramuscular Islet Transplantation.

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Haruyuki Tsuchiya

Full Text Available The efficacy of intramuscular islet transplantation is poor despite being technically simple, safe, and associated with reduced rates of severe complications. We evaluated the efficacy of combined treatment with extracellular matrix (ECM and growth factors in intramuscular islet transplantation.Male BALB/C mice were used for the in vitro and transplantation studies. The following three groups were evaluated: islets without treatment (islets-only group, islets embedded in ECM with growth factors (Matrigel group, and islets embedded in ECM without growth factors [growth factor-reduced (GFR Matrigel group]. The viability and insulin-releasing function of islets cultured for 96 h were significantly improved in Matrigel and GFR Matrigel groups compared with the islets-only group.Blood glucose and serum insulin levels immediately following transplantation were significantly improved in the Matrigel and GFR Matrigel groups and remained significantly improved in the Matrigel group at postoperative day (POD 28. On histological examination, significantly decreased numbers of TdT-mediated deoxyuridine triphosphate-biotin nick end labeling-positive islet cells and significantly increased numbers of Ki67-positive cells were observed in the Matrigel and GFR Matrigel groups at POD 3. Peri-islet revascularization was most prominent in the Matrigel group at POD 14.The efficacy of intramuscular islet transplantation was improved by combination treatment with ECM and growth factors through the inhibition of apoptosis, increased proliferation of islet cells, and promotion of revascularization.

Impact of Pancreatic Rat Islet Density on Cell Survival during Hypoxia

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A. Rodriguez-Brotons

2016-01-01

Full Text Available In bioartificial pancreases (BP, the number of islets needed to restore normoglycaemia in the diabetic patient is critical. However, the confinement of a high quantity of islets in a limited space may impact islet survival, particularly in regard to the low oxygen partial pressure (PO2 in such environments. The aim of the present study was to evaluate the impact of islet number in a confined space under hypoxia on cell survival. Rat islets were seeded at three different concentrations (150, 300, and 600 Islet Equivalents (IEQ/cm2 and cultured in normal atmospheric pressure (160 mmHg as well as hypoxic conditions (15 mmHg for 24 hours. Cell viability, function, hypoxia-induced changes in gene expression, and cytokine secretion were then assessed. Notably, hypoxia appeared to induce a decrease in viability and increasing islet density exacerbated the observed increase in cellular apoptosis as well as the loss of function. These changes were also associated with an increase in inflammatory gene transcription. Taken together, these data indicate that when a high number of islets are confined to a small space under hypoxia, cell viability and function are significantly impacted. Thus, in order to improve islet survival in this environment during transplantation, oxygenation is of critical importance.

Effect of oxygenated perfluorocarbon on isolated islets during transportation.

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Terai, Sachio; Tsujimura, Toshiaki; Li, Shiri; Hori, Yuichi; Toyama, Hirochika; Shinzeki, Makoto; Matsumoto, Ippei; Kuroda, Yoshikazu; Ku, Yonson

2010-08-01

Previous studies demonstrated the efficacy of the two-layer method (TLM) using oxygenated perfluorochemicals (PFC) for pancreas preservation. The current study investigated the effect of oxygenated PFC on isolated islets during transportation. Purified rat islets were stored in an airtight conical tube for 24h in RPMI culture medium at 22 degrees C or University of Wisconsin solution (UW) at 4 degrees C, either with or without oxygenated PFC. After storage, the islets were assessed for in vitro viability by static incubation (SI), FDA/PI staining, and energy status (ATP, energy charge, and ADP/ATP ratio) and for in vivo viability by a transplantation study. UW at 4 degrees C and RPMI medium at 22 degrees C maintained islet quality almost equally in both in vitro and in vivo assessments. The ATP levels and energy status in the groups with PFC were significantly lower than those without PFC. The groups with PFC showed a significantly higher ADP/ATP ratio than those without PFC. In the transplantation study, blood glucose levels and AUC in the UW+PFC group were significantly higher than those in UW group. UW at 4 degrees C and RPMI medium at 22 degrees C maintained islet quality equally under the conditions for islet transportation. The addition of oxygenated PFC, while advantageous for pancreas preservation, is not useful for islet transportation. Copyright 2010 Elsevier Inc. All rights reserved.

Possible modulatory effect of endogenous islet catecholamines on insulin secretion

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Gagliardino Juan J

2001-10-01

Full Text Available Abstract Background The possible participation of endogenous islet catecholamines (CAs in the control of insulin secretion was tested. Methods Glucose-induced insulin secretion was measured in the presence of 3-Iodo-L-Tyrosine (MIT, a specific inhibitor of tyrosine-hydroxylase activity, in fresh and precultured islets isolated from normal rats. Incubated islets were also used to measure CAs release in the presence of low and high glucose, and the effect of α2-(yohimbine [Y] and idazoxan [I] and α1-adrenergic antagonists (prazosin [P] and terazosin [T] upon insulin secretion elicited by high glucose. Results Fresh islets incubated with 16.7 mM glucose released significantly more insulin in the presence of 1 μM MIT (6.66 ± 0.39 vs 5.01 ± 0.43 ng/islet/h, p Conclusion Our results suggest that islet-originated CAs directly modulate insulin release in a paracrine manner.

Metabolic Profile of Pancreatic Acinar and Islet Tissue in Culture

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Suszynski, Thomas M.; Mueller, Kathryn; Gruessner, Angelika C.; Papas, Klearchos K.

2016-01-01

The amount and condition of exocrine impurities may affect the quality of islet preparations especially during culture. In this study, the objective was to determine the oxygen demandand viability of islet and acinar tissue post-isolation and whether they change disproportionately while in culture. We compare the OCR normalized to DNA (OCR/DNA, a measure of fractional viability in units nmol/min/mg DNA), and percent change in OCR and DNA recoveries between adult porcine islet and acinar tissue from the same preparation (paired) over a 6-9 days of standard culture. Paired comparisons were done to quantify differences in OCR/DNA between islet and acinar tissue from the same preparation, at specified time points during culture; the mean (± standard error) OCR/DNA was 74.0 (±11.7) units higher for acinar (vs. islet) tissue on the day of isolation (n=16, p<0.0001), but 25.7 (±9.4) units lower after 1 day (n=8, p=0.03), 56.6 (±11.5) units lower after 2 days (n=12, p=0.0004), and 65.9 (±28.7) units lower after 8 days (n=4, p=0.2) in culture. DNA and OCR recoveries decreased at different rates for acinar versus islet tissue over 6-9 days in culture (n=6). DNA recovery decreased to 24±7% for acinar and 75±8% for islets (p=0.002). Similarly, OCR recovery decreased to 16±3% for acinar and remained virtually constant for islets (p=0.005). Differences in the metabolic profile of acinarand islet tissue should be considered when culturing impure islet preparations. OCR-based measurements may help optimize pre-IT culture protocols. PMID:25131082

Modular tissue engineering for the vascularization of subcutaneously transplanted pancreatic islets.

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Vlahos, Alexander E; Cober, Nicholas; Sefton, Michael V

2017-08-29

The transplantation of pancreatic islets, following the Edmonton Protocol, is a promising treatment for type I diabetics. However, the need for multiple donors to achieve insulin independence reflects the large loss of islets that occurs when islets are infused into the portal vein. Finding a less hostile transplantation site that is both minimally invasive and able to support a large transplant volume is necessary to advance this approach. Although the s.c. site satisfies both these criteria, the site is poorly vascularized, precluding its utility. To address this problem, we demonstrate that modular tissue engineering results in an s.c. vascularized bed that enables the transplantation of pancreatic islets. In streptozotocin-induced diabetic SCID/beige mice, the injection of 750 rat islet equivalents embedded in endothelialized collagen modules was sufficient to restore and maintain normoglycemia for 21 days; the same number of free islets was unable to affect glucose levels. Furthermore, using CLARITY, we showed that embedded islets became revascularized and integrated with the host's vasculature, a feature not seen in other s.c. Collagen-embedded islets drove a small (albeit not significant) shift toward a proangiogenic CD206 + MHCII - (M2-like) macrophage response, which was a feature of module-associated vascularization. While these results open the potential for using s.c. islet delivery as a treatment option for type I diabetes, the more immediate benefit may be for the exploration of revascularized islet biology.

Beta-Cell Replacement: Pancreas and Islet Cell Transplantation.

Science.gov (United States)

Niclauss, Nadja; Meier, Raphael; Bédat, Benoît; Berishvili, Ekaterine; Berney, Thierry

2016-01-01

Pancreas and islet transplantation are 2 types of beta-cell replacement therapies for type 1 diabetes mellitus. Since 1966, when pancreas transplantation was first performed, it has evolved to become a highly efficient procedure with high success rates, thanks to advances in surgical technique and immunosuppression. Pancreas transplantation is mostly performed as simultaneous pancreas-kidney transplantation in patients with end-stage nephropathy secondary to diabetes. In spite of its efficiency, pancreas transplantation is still a major surgical procedure burdened by high morbidity, which called for the development of less invasive and hazardous ways of replacing beta-cell function in the past. Islet transplantation was developed in the 1970s as a minimally invasive procedure with initially poor outcomes. However, since the report of the 'Edmonton protocol' in 2000, the functional results of islet transplantation have substantially and constantly improved and are about to match those of whole pancreas transplantation. Islet transplantation is primarily performed alone in nonuremic patients with severe hypoglycemia. Both pancreas transplantation and islet transplantation are able to abolish hypoglycemia and to prevent or slow down the development of secondary complications of diabetes. Pancreas transplantation and islet transplantation should be seen as two complementary, rather than competing, therapeutic approaches for beta-cell replacement that are able to optimize organ donor use and patient care. © 2016 S. Karger AG, Basel.

Effect of alcohol on insulin secretion and viability of human pancreatic islets

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Nikolić Dragan

2017-01-01

Full Text Available Introduction/Objective. There are controversial data in the literature on the topic of effects of alcohol on insulin secretion, apoptosis, and necrosis of the endocrine and exocrine pancreas. The goal of this research was to determine how alcohol affects the insulin secretion and viability of human adult pancreatic islets in vitro during a seven-day incubation. Methods. Human pancreatic tissue was digested with Collagenase XI, using a non-automated method. Cultures were incubated in Roswell Park Memorial Institute (RPMI medium containing alcohol (10 μl of alcohol in 100 ml of medium. Insulin stimulation index (SI and viability of the islets were determined on the first, third, and seventh day of cultivation. Results. Analysis of the viability of the islets showed that there wasn’t significant difference between the control and the test group. In the test group, viability of the cultures declined with the time of incubation. SI of the test group was higher compared to the control group, by 50% and 25% on the first and third day of cultivation, respectively. On the seventh day, insulin secretion was reduced by 25%. The difference was not statistically significant (p > 0.05. In the test group, significant decline in insulin secretion was found on the third and seventh day of incubation (p ≤ 0.05. Conclusion. Alcohol can increase or decrease insulin secretion of islets cultures, which may result in an inadequate response of pancreatic β-cells to blood glucose, leading to insulin resistance, and increased risk of developing type 2 diabetes. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. 41002

Effects of Acute Cytomegalovirus Infection on Rat Islet Allograft Survival

NARCIS (Netherlands)

Smelt, M. J.; Faas, M. M.; Melgert, B. N.; de Vos, P.; de Haan, Bart; de Haan, Aalzen

2011-01-01

Transplantation of pancreatic islets is a promising therapy for the treatment of type 1 diabetes mellitus. However, long-term islet graft survival rates are still unsatisfactory low. In this study we investigated the role of cytomegalovirus (CMV) in islet allograft failure. STZ-diabetic rats

New stepwise cooling system for short-term porcine islet preservation.

Science.gov (United States)

Ikemoto, Tetsuya; Noguchi, Hirofumi; Fujita, Yasutaka; Takita, Morihito; Shimoda, Masayuki; Sugimoto, Koji; Jackson, Andrew; Naziruddin, Bashoo; Shimada, Mitsuo; Levy, Marlon F; Matsumoto, Shinichi

2010-10-01

Porcine islets are the most suitable for xeno-islet transplantation. However, it is necessary to establish an effective preservation method against its fragility. Recently, we developed a new cooling and preservation (Keep and Fresh [KFC]; FUJIYA Co, Tokushima, Japan) system, which can maintain viability of hepatocyte. In this study, we examined the KFC for porcine islet preservation. Isolated porcine islets were preserved in CMRL 1066 culture media with bovine serum at 37°C, 22°C, and 4°C and KFC for 24, 48, and 72 hours. Islet recovery rate, purity, and viability were evaluated. After 24-hour preservation, the recovery rate was the highest in the KFC, but no significant difference was found. After 48-hour preservation, the recovery rate by the KFC was 73.9% ± 17.3%, which was significantly higher than the other groups (48.7% ± 28.6% at 37°C, P KFC group, purities and viabilities were the highest among the groups after 24-, 48-, and 72-hour preservation. The KFC system significantly improved porcine islet preservation; therefore, the KFC might be useful for porcine islet preservation.

Automated digital image analysis of islet cell mass using Nikon's inverted eclipse Ti microscope and software to improve engraftment may help to advance the therapeutic efficacy and accessibility of islet transplantation across centers.

Science.gov (United States)

Gmyr, Valery; Bonner, Caroline; Lukowiak, Bruno; Pawlowski, Valerie; Dellaleau, Nathalie; Belaich, Sandrine; Aluka, Isanga; Moermann, Ericka; Thevenet, Julien; Ezzouaoui, Rimed; Queniat, Gurvan; Pattou, Francois; Kerr-Conte, Julie

2015-01-01

Reliable assessment of islet viability, mass, and purity must be met prior to transplanting an islet preparation into patients with type 1 diabetes. The standard method for quantifying human islet preparations is by direct microscopic analysis of dithizone-stained islet samples, but this technique may be susceptible to inter-/intraobserver variability, which may induce false positive/negative islet counts. Here we describe a simple, reliable, automated digital image analysis (ADIA) technique for accurately quantifying islets into total islet number, islet equivalent number (IEQ), and islet purity before islet transplantation. Islets were isolated and purified from n = 42 human pancreata according to the automated method of Ricordi et al. For each preparation, three islet samples were stained with dithizone and expressed as IEQ number. Islets were analyzed manually by microscopy or automatically quantified using Nikon's inverted Eclipse Ti microscope with built-in NIS-Elements Advanced Research (AR) software. The AIDA method significantly enhanced the number of islet preparations eligible for engraftment compared to the standard manual method (p image analysis utilizing the Nikon Instruments software is an unbiased, simple, and reliable teaching tool to comprehensively assess the individual size of each islet cell preparation prior to transplantation. Implementation of this technology to improve engraftment may help to advance the therapeutic efficacy and accessibility of islet transplantation across centers.

Altered Expression of Somatostatin Receptors in Pancreatic Islets from NOD Mice Cultured at Different Glucose Concentrations In Vitro and in Islets Transplanted to Diabetic NOD Mice In Vivo

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Eva Ludvigsen

2011-01-01

Full Text Available Somatostatin acts via five receptors (sst1-5. We investigated if the changes in pancreatic islet sst expression in diabetic NOD mice compared to normoglycemic mice are a consequence of hyperglycemia or the ongoing immune reaction in the pancreas. Pancreatic islets were isolated from NOD mice precultured for 5 days and further cultured for 3 days at high or low glucose before examined. Islets were also isolated from NOD mice and transplanted to normal or diabetic mice in a number not sufficient to cure hyperglycemia. After three days, the transplants were removed and stained for sst1-5 and islet hormones. Overall, changes in sst islet cell expression were more common in islets cultured in high glucose concentration in vitro as compared to the islet transplantation in vivo to diabetic mice. The beta and PP cells exhibited more frequent changes in sst expression, while the alpha and delta cells were relatively unaffected by the high glucose condition. Our findings suggest that the glucose level may alter sst expressed in islets cells; however, immune mechanisms may counteract such changes in islet sst expression.

Intraportal islet transplantation: the impact of the liver microenvironment.

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Delaune, Vaihere; Berney, Thierry; Lacotte, Stéphanie; Toso, Christian

2017-03-01

The portal vein remains the preferred site for pancreatic islet transplantation due to its easy access and low morbidity. However, despite great progress in isolation and transplantation protocols over the past few years, it is still associated with the early loss of some 50-70% of transplanted islets. The complex liver microenvironment itself presumably plays an important role in this loss. The present review focuses on the specifics of the liver microenvironment, notably the localized hepatic ischemia/reperfusion injury following transplantation, the low oxygenation of the portal vein, the instant blood-mediated inflammatory reaction, the endogenous liver immune system, and the gut-liver axis, and how they can each have an impact on the transplanted islets. It identifies the potential, or already applied, clinical interventions for improving intraportal islet survival, and pinpoints those promising areas still lacking preclinical research. Future interventions on clinical intraportal islet transplantation need to take into account the global context of the liver microenvironment, with multi-point interventions being most likely to improve early islet survival and engraftment. © 2017 The Authors. Transplant International published by John Wiley & Sons Ltd on behalf of Steunstichting ESOT.

Affinity-purified human interleukin I is cytotoxic to isolated islets of Langerhans

DEFF Research Database (Denmark)

Mandrup-Poulsen, T; Bendtzen, K; Nerup, J

1986-01-01

Addition of highly purified human Interleukin-1 to the culture medium of isolated rat islets of Langerhans for 6 days led to 88% inhibition of glucose-induced insulin-release, reduction of islet contents of insulin and glucagon to 31% and 8% respectively, and disintegration of the islets. These e......Addition of highly purified human Interleukin-1 to the culture medium of isolated rat islets of Langerhans for 6 days led to 88% inhibition of glucose-induced insulin-release, reduction of islet contents of insulin and glucagon to 31% and 8% respectively, and disintegration of the islets...

Alginate Microencapsulation of Human Islets Does Not Increase Susceptibility to Acute Hypoxia

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I. K. Hals

2013-01-01

Full Text Available Islet transplantation in diabetes is hampered by the need of life-long immunosuppression. Encapsulation provides partial immunoprotection but could possibly limit oxygen supply, a factor that may enhance hypoxia-induced beta cell death in the early posttransplantation period. Here we tested susceptibility of alginate microencapsulated human islets to experimental hypoxia (0.1–0.3% O2 for 8 h, followed by reoxygenation on viability and functional parameters. Hypoxia reduced viability as measured by MTT by 33.8±3.5% in encapsulated and 42.9±5.2% in nonencapsulated islets (P<0.2. Nonencapsulated islets released 37.7% (median more HMGB1 compared to encapsulated islets after hypoxic culture conditions (P<0.001. Glucose-induced insulin release was marginally affected by hypoxia. Basal oxygen consumption was equally reduced in encapsulated and nonencapsulated islets, by 22.0±6.1% versus 24.8±5.7%. Among 27 tested cytokines/chemokines, hypoxia increased the secretion of IL-6 and IL-8/CXCL8 in both groups of islets, whereas an increase of MCP-1/CCL2 was seen only with nonencapsulated islets. Conclusion. Alginate microencapsulation of human islets does not increase susceptibility to acute hypoxia. This is a positive finding in relation to potential use of encapsulation for islet transplantation.

Impact of Procedure-Related Complications on Long-term Islet Transplantation Outcome.

Science.gov (United States)

Caiazzo, Robert; Vantyghem, Marie-Christine; Raverdi, Violeta; Bonner, Caroline; Gmyr, Valery; Defrance, Frederique; Leroy, Clara; Sergent, Geraldine; Hubert, Thomas; Ernst, Oliver; Noel, Christian; Kerr-Conte, Julie; Pattou, François

2015-05-01

Pancreatic islet transplantation offers a promising biotherapy for the treatment of type 1 diabetes, but this procedure has met significant challenges over the years. One such challenge is to address why primary graft function still remains inconsistent after islet transplantation. Several variables have been shown to affect graft function, but the impact of procedure-related complications on primary and long-term graft functions has not yet been explored. Twenty-six patients with established type 1 diabetes were included in this study. Each patient had two to three intraportal islet infusions to obtain 10,000 islet equivalent (IEQ)/kg in body weight, equaling a total of 68 islet infusions. Islet transplantation consisted of three sequential fresh islet infusions within 3 months. Islet infusions were performed surgically or under ultrasound guidance, depending on patient morphology, availability of the radiology suite, and patient medical history. Prospective assessment of adverse events was recorded and graded using "Common Terminology Criteria for adverse events in Trials of Adult Pancreatic Islet Transplantation." There were no deaths or patients dropouts. Early complications occurred in nine of 68 procedures. β score 1 month after the last graft and optimal graft function (β score ≥7) rate were significantly lower in cases of procedure-related complications (P = 0.02, P = 0.03). Procedure-related complications negatively impacted graft function (P = 0.009) and was an independent predictive factor of long-term graft survival (P = 0.033) in multivariate analysis. Complications occurring during radiologic or surgical intraportal islet transplantation significantly impair primary graft function and graft survival regardless of their severity.

Oxygenated thawing and rewarming alleviate rewarming injury of cryopreserved pancreatic islets.

Science.gov (United States)

Komatsu, Hirotake; Barriga, Alyssa; Medrano, Leonard; Omori, Keiko; Kandeel, Fouad; Mullen, Yoko

2017-05-06

Pancreatic islet transplantation is an effective treatment for Type 1 diabetic patients to eliminate insulin injections; however, a shortage of donor organs hinders the widespread use. Although long-term islet storage, such as cryopreservation, is considered one of the key solutions, transplantation of cryopreserved islets is still not practical due to the extensive loss during the cryopreservation-rewarming process. We have previously reported that culturing islets in a hyperoxic environment is an effective treatment to prevent islet death from the hypoxic injury during culture. In this study, we explored the effectiveness of thawing and rewarming cryopreserved islets in a hyperoxic environment. Following cryopreservation of isolated human islets, the thawing solution and culture media were prepared with or without pre-equilibration to 50% oxygen. Thawing/rewarming and the pursuant two-day culture were performed with or without oxygenation. Short-term recovery rate, defined as the volume change during cryopreservation and thawing/rewarming, was assessed. Ischemia-associated and inflammation-associated gene expressions were examined using qPCR after the initial rewarming period. Long-term recovery rate, defined as the volume change during the two-day culture after the thawing/rewarming, was also examined. Islet metabolism and function were assessed by basal oxygen consumption rate and glucose stimulated insulin secretion after long-term recovery. Oxygenated thawing/rewarming did not alter the short-term recovery rate. Inflammation-associated gene expressions were elevated by the conventional thawing/rewarming method and suppressed by the oxygenated thawing/rewarming, whereas ischemia-associated gene expressions did not change between the thawing/rewarming methods. Long-term recovery rate experiments revealed that only the combination therapy of oxygenated thawing/rewarming and oxygenated culture alleviated islet volume loss. These islets showed higher metabolism

Can pancreatic duct-derived progenitors be a source of islet regeneration?

International Nuclear Information System (INIS)

Xia, Bing; Zhan, Xiao-Rong; Yi, Ran; Yang, Baofeng

2009-01-01

The regenerative process of the pancreas is of interest because the main pathogenesis of diabetes mellitus is an inadequate number of insulin-producing β-cells. The functional mass of β-cells is decreased in type 1 diabetes, so replacing missing β-cells or triggering their regeneration may allow for improved type 1 diabetes treatment. Therefore, expansion of the β-cell mass from endogenous sources, either in vivo or in vitro, represents an area of increasing interest. The mechanism of islet regeneration remains poorly understood, but the identification of islet progenitor sources is critical for understanding β-cell regeneration. One potential source is the islet proper, via the dedifferentiation, proliferation, and redifferentiation of facultative progenitors residing within the islet. Neogenesis, or that the new pancreatic islets can derive from progenitor cells present within the ducts has been reported, but the existence and identity of the progenitor cells have been debated. In this review, we focus on pancreatic ductal cells, which are islet progenitors capable of differentiating into islet β-cells. Islet neogenesis, seen as budding of hormone-positive cells from the ductal epithelium, is considered to be one mechanism for normal islet growth after birth and in regeneration, and has suggested the presence of pancreatic stem cells. Numerous results support the neogenesis hypothesis, the evidence for the hypothesis in the adult comes primarily from morphological studies that have in common the production of damage to all or part of the pancreas, with consequent inflammation and repair. Although numerous studies support a ductal origin for new islets after birth, lineage-tracing experiments are considered the 'gold standard' of proof. Lineage-tracing experiments show that pancreatic duct cells act as progenitors, giving rise to new islets after birth and after injury. The identification of differentiated pancreatic ductal cells as an in vivo progenitor for

Can pancreatic duct-derived progenitors be a source of islet regeneration?

Energy Technology Data Exchange (ETDEWEB)

Xia, Bing [Department of Endocrinology, First Hospital of Harbin Medical University, Harbin, Hei Long Jiang Province 150001 (China); Zhan, Xiao-Rong, E-mail: xiaorongzhan@sina.com [Department of Endocrinology, First Hospital of Harbin Medical University, Harbin, Hei Long Jiang Province 150001 (China); Yi, Ran [Department of Endocrinology, First Hospital of Harbin Medical University, Harbin, Hei Long Jiang Province 150001 (China); Yang, Baofeng [Department of Pharmacology, State Key Laboratory of Biomedicine and Pharmacology, Harbin Medical University, Harbin, Hei Long Jiang Province 150001 (China)

2009-06-12

The regenerative process of the pancreas is of interest because the main pathogenesis of diabetes mellitus is an inadequate number of insulin-producing {beta}-cells. The functional mass of {beta}-cells is decreased in type 1 diabetes, so replacing missing {beta}-cells or triggering their regeneration may allow for improved type 1 diabetes treatment. Therefore, expansion of the {beta}-cell mass from endogenous sources, either in vivo or in vitro, represents an area of increasing interest. The mechanism of islet regeneration remains poorly understood, but the identification of islet progenitor sources is critical for understanding {beta}-cell regeneration. One potential source is the islet proper, via the dedifferentiation, proliferation, and redifferentiation of facultative progenitors residing within the islet. Neogenesis, or that the new pancreatic islets can derive from progenitor cells present within the ducts has been reported, but the existence and identity of the progenitor cells have been debated. In this review, we focus on pancreatic ductal cells, which are islet progenitors capable of differentiating into islet {beta}-cells. Islet neogenesis, seen as budding of hormone-positive cells from the ductal epithelium, is considered to be one mechanism for normal islet growth after birth and in regeneration, and has suggested the presence of pancreatic stem cells. Numerous results support the neogenesis hypothesis, the evidence for the hypothesis in the adult comes primarily from morphological studies that have in common the production of damage to all or part of the pancreas, with consequent inflammation and repair. Although numerous studies support a ductal origin for new islets after birth, lineage-tracing experiments are considered the 'gold standard' of proof. Lineage-tracing experiments show that pancreatic duct cells act as progenitors, giving rise to new islets after birth and after injury. The identification of differentiated pancreatic ductal

Has the gap between pancreas and islet transplantation closed?

Science.gov (United States)

Niclauss, Nadja; Morel, Philippe; Berney, Thierry

2014-09-27

Both pancreas and islet transplantations are therapeutic options for complicated type 1 diabetes. Until recent years, outcomes of islet transplantation have been significantly inferior to those of whole pancreas. Islet transplantation is primarily performed alone in patients with severe hypoglycemia, and recent registry reports have suggested that results of islet transplantation alone in this indication may be about to match those of pancreas transplant alone in insulin independence. Figures of 50% insulin independence at 5 years for either procedure have been cited. In this article, we address the question whether islet transplantation has indeed bridged the gap with whole pancreas. Looking at the evidence to answer this question, we propose that although pancreas may still be more efficient in taking recipients off insulin than islets, there are in fact numerous "gaps" separating both procedures that must be taken into the equation. These "gaps" relate to organ utilization, organ allocation, indication for transplantation, and morbidity. In-depth analysis reveals that islet transplantation, in fact, has an edge on whole pancreas in some of these aspects. Accordingly, attempts should be made to bridge these gaps from both sides to achieve the same level of success with either procedure. More realistically, it is likely that some of these gaps will remain and that both procedures will coexist and complement each other, to ensure that β cell replacement can be successfully implemented in the greatest possible number of patients with type 1 diabetes.

A 3D map of the islet routes throughout the healthy human pancreas

Science.gov (United States)

Ionescu-Tirgoviste, Constantin; Gagniuc, Paul A.; Gubceac, Elvira; Mardare, Liliana; Popescu, Irinel; Dima, Simona; Militaru, Manuella

2015-01-01

Islets of Langerhans are fundamental in understanding diabetes. A healthy human pancreas from a donor has been used to asses various islet parameters and their three-dimensional distribution. Here we show that islets are spread gradually from the head up to the tail section of the pancreas in the form of contracted or dilated islet routes. We also report a particular anatomical structure, namely the cluster of islets. Our observations revealed a total of 11 islet clusters which comprise of small islets that surround large blood vessels. Additional observations in the peripancreatic adipose tissue have shown lymphoid-like nodes and blood vessels captured in a local inflammatory process. Our observations are based on regional slice maps of the pancreas, comprising of 5,423 islets. We also devised an index of sphericity which briefly indicates various islet shapes that are dominant throughout the pancreas. PMID:26417671

Update on Islet Transplantation

Science.gov (United States)

McCall, Michael; James Shapiro, A.M.

2012-01-01

Clinical islet transplantation has progressed considerably over the past 12 years, and >750 patients with type 1 diabetes have received islet transplants internationally over this time. Many countries are beginning to accept the transition from research to accepted and funded clinical care, especially for patients with brittle control that cannot be stabilized by more conventional means. Major challenges remain, including the need for more than one donor, and the requirement for potent, chronic immunosuppression. Combining immunological tolerance both to allo- and autoantigens, and a limitless expandable source of stem cell- or xenograft-derived insulin-secreting cells represent remaining hurdles in moving this effective treatment to a potential cure for all those with type 1 or 2 diabetes. PMID:22762022

The hypothalamic satiety peptide CART is expressed in anorectic and non-anorectic pancreatic islet tumors and in the normal islet of Langerhans.

Science.gov (United States)

Jensen, P B; Kristensen, P; Clausen, J T; Judge, M E; Hastrup, S; Thim, L; Wulff, B S; Foged, C; Jensen, J; Holst, J J; Madsen, O D

1999-03-26

The hypothalamic satiety peptide CART (cocaine and amphetamine regulated transcript) is expressed at high levels in anorectic rat glucagonomas but not in hypoglycemic insulinomas. However, a non-anorectic metastasis derived from the glucagonoma retained high CART expression levels and produced circulating CART levels comparable to that of the anorectic tumors. Moreover, distinct glucagonoma lines derived by stable HES-1 transfection of the insulinoma caused severe anorexia but retained low circulating levels of CART comparable to that of insulinoma bearing or control rats. Islet tumor associated anorexia and circulating CART levels are thus not correlated, and in line with this peripheral administration of CART (5-50 mg/kg) produced no effect on feeding behavior. In the rat two alternatively spliced forms of CART mRNA exist and quantitative PCR revealed expression of both forms in the hypothalamus, in the different islet tumors, and in the islets of Langerhans. Immunocytochemistry as well as in situ hybridization localized CART expression to the somatostatin producing islet D cell. A potential endocrine/paracrine role of islet CART remains to be clarified.

Zinc as a paracrine effector in pancreatic islet cell death.

Science.gov (United States)

Kim, B J; Kim, Y H; Kim, S; Kim, J W; Koh, J Y; Oh, S H; Lee, M K; Kim, K W; Lee, M S

2000-03-01

Because of a huge amount of Zn2+ in secretory granules of pancreatic islet beta-cells, Zn2+ released in certain conditions might affect the function or survival of islet cells. We studied potential paracrine effects of endogenous Zn2+ on beta-cell death. Zn2+ induced insulinoma/islet cell death in a dose-dependent manner. Chelation of released endogenous Zn2+ by CaEDTA significantly decreased streptozotocin (STZ)-induced islet cell death in an in vitro culture system simulating in vivo circumstances but not in the conventional culture system. Zn2+ chelation in vivo by continuous CaEDTA infusion significantly decreased the incidence of diabetes after STZ administration. N-(6-methoxy-quinolyl)-para-toluene-sulfonamide staining revealed that Zn2+ was densely deposited in degenerating islet cells 24 h after STZ treatment, which was decreased by CaEDTA infusion. We show here that Zn2+ is not a passive element for insulin storage but an active participant in islet cell death in certain conditions, which in time might contribute to the development of diabetes in aged people.

Glycemia, Hypoglycemia, and Costs of Simultaneous Islet-Kidney or Islet After Kidney Transplantation Versus Intensive Insulin Therapy and Waiting List for Islet Transplantation.

Science.gov (United States)

Gerber, Philipp A; Locher, Rebecca; Zuellig, Richard A; Tschopp, Oliver; Ajdler-Schaeffler, Evelyne; Kron, Philipp; Oberkofler, Christian; Brändle, Michael; Spinas, Giatgen A; Lehmann, Roger

2015-10-01

Long-term data of patients with type 1 diabetes mellitus (T1D) after simultaneous islet-kidney (SIK) or islet-after-kidney transplantation (IAK) are rare and have never been compared to intensified insulin therapy (IIT). Twenty-two patients with T1D and end-stage renal failure undergoing islet transplantation were compared to 70 patients matched for age and diabetes duration treated with IIT and to 13 patients with kidney transplantation alone or simultaneous pancreas-kidney after loss of pancreas function (waiting list for IAK [WLI]). Glycemic control, severe hypoglycemia, insulin requirement, and direct medical costs were analyzed. Glycated hemoglobin decreased significantly from 8.2 ± 1.5 to 6.7 ± 0.9% at the end of follow-up (mean 7.2 ± 2.5 years) in the SIK/IAK and remained constant in IIT (7.8 ± 1.0% and 7.6 ± 1.0) and WLI (7.8 ± 0.8 and 7.9 ± 1.0%). Daily insulin requirement decreased from 0.53 ± 0.15 to 0.29 ± 0.26 U/kg and remained constant in IIT (0.59 ± 0.19 and 0.58 ± 0.23 U/kg) and in WLI (0.76 ± 0.28 and 0.73 ± 0.11 U/kg). Severe hypoglycemia dropped in SIK/IAK from 4.5 ± 9.7 to 0.3 ± 0.7/patient-year and remained constant in IIT (0.1 ± 0.7 and 0.2 ± 0.8/patient-year). Detailed cost analysis revealed US $57,525 of additional cost for islet transplantation 5 years after transplantation. Based on a 5- and 10-year analysis, cost neutrality is assumed to be achieved 15 years after transplantation. This long-term cohort with more than 7 years of follow-up shows that glycemic control in patients with T1D after SIK/IAK transplantation improved, and the rate of severe hypoglycemia decreased significantly as compared to control groups. Cost analysis revealed that islet transplantation is estimated to be cost neutral at 15 years after transplantation.

Mesenchymal stromal cells improve human islet function through released products and extracellular matrix.

Science.gov (United States)

Arzouni, Ahmed A; Vargas-Seymour, Andreia; Rackham, Chloe L; Dhadda, Paramjeet; Huang, Guo-Cai; Choudhary, Pratik; Nardi, Nance; King, Aileen J F; Jones, Peter M

2017-12-01

The aims of the present study were (i) to determine whether the reported beneficial effects of mesenchymal stromal cells (MSCs) on mouse islet function extend to clinically relevant human tissues (islets and MSCs), enabling translation into improved protocols for clinical human islet transplantation; and (ii) to identify possible mechanisms through which human MSCs influence human islet function. Human islets were co-cultured with human adipose tissue-derived MSCs (hASCs) or pre-treated with its products - extracellular matrix (ECM) and annexin A1 (ANXA1). Mouse islets were pre-treated with mouse MSC-derived ECM. Islet insulin secretory function was assessed in vitro by radioimmunoassay. Quantitative RT-PCR was used to screen human adipMSCs for potential ligands of human islet G-protein-coupled receptors. We show that co-culture with hASCs improves human islet secretory function in vitro , as measured by glucose-stimulated insulin secretion, confirming previous reports using rodent tissues. Furthermore, we demonstrate that these beneficial effects on islet function can be partly attributed to the MSC-derived products ECM and ANXA1. Our results suggest that hASCs have the potential to improve the quality of human islets isolated for transplantation therapy of Type 1 diabetes. Furthermore, it may be possible to achieve improvements in human islet quality in a cell-free culture system by using the MSC-derived products ANXA1 and ECM. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Abnormal islet sphingolipid metabolism in type 1 diabetes.

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Holm, Laurits J; Krogvold, Lars; Hasselby, Jane P; Kaur, Simranjeet; Claessens, Laura A; Russell, Mark A; Mathews, Clayton E; Hanssen, Kristian F; Morgan, Noel G; Koeleman, Bobby P C; Roep, Bart O; Gerling, Ivan C; Pociot, Flemming; Dahl-Jørgensen, Knut; Buschard, Karsten

2018-04-18

Sphingolipids play important roles in beta cell physiology, by regulating proinsulin folding and insulin secretion and in controlling apoptosis, as studied in animal models and cell cultures. Here we investigate whether sphingolipid metabolism may contribute to the pathogenesis of human type 1 diabetes and whether increasing the levels of the sphingolipid sulfatide would prevent models of diabetes in NOD mice. We examined the amount and distribution of sulfatide in human pancreatic islets by immunohistochemistry, immunofluorescence and electron microscopy. Transcriptional analysis was used to evaluate expression of sphingolipid-related genes in isolated human islets. Genome-wide association studies (GWAS) and a T cell proliferation assay were used to identify type 1 diabetes related polymorphisms and test how these affect cellular islet autoimmunity. Finally, we treated NOD mice with fenofibrate, a known activator of sulfatide biosynthesis, to evaluate the effect on experimental autoimmune diabetes development. We found reduced amounts of sulfatide, 23% of the levels in control participants, in pancreatic islets of individuals with newly diagnosed type 1 diabetes, which were associated with reduced expression of enzymes involved in sphingolipid metabolism. Next, we discovered eight gene polymorphisms (ORMDL3, SPHK2, B4GALNT1, SLC1A5, GALC, PPARD, PPARG and B4GALT1) involved in sphingolipid metabolism that contribute to the genetic predisposition to type 1 diabetes. These gene polymorphisms correlated with the degree of cellular islet autoimmunity in a cohort of individuals with type 1 diabetes. Finally, using fenofibrate, which activates sulfatide biosynthesis, we completely prevented diabetes in NOD mice and even reversed the disease in half of otherwise diabetic animals. These results indicate that islet sphingolipid metabolism is abnormal in type 1 diabetes and suggest that modulation may represent a novel therapeutic approach. The RNA expression data is

Mesenchymal Stem Cells Enhance Allogeneic Islet Engraftment in Nonhuman Primates

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Berman, Dora M.; Willman, Melissa A.; Han, Dongmei; Kleiner, Gary; Kenyon, Norman M.; Cabrera, Over; Karl, Julie A.; Wiseman, Roger W.; O'Connor, David H.; Bartholomew, Amelia M.; Kenyon, Norma S.

2010-01-01

OBJECTIVE To test the graft-promoting effects of mesenchymal stem cells (MSCs) in a cynomolgus monkey model of islet/bone marrow transplantation. RESEARCH DESIGN AND METHODS Cynomolgus MSCs were obtained from iliac crest aspirate and characterized through passage 11 for phenotype, gene expression, differentiation potential, and karyotype. Allogeneic donor MSCs were cotransplanted intraportally with islets on postoperative day (POD) 0 and intravenously with donor marrow on PODs 5 and 11. Recipients were followed for stabilization of blood glucose levels, reduction of exogenous insulin requirement (EIR), C-peptide levels, changes in peripheral blood T regulatory cells, and chimerism. Destabilization of glycemia and increases in EIR were used as signs of rejection; additional intravenous MSCs were administered to test the effect on reversal of rejection. RESULTS MSC phenotype and a normal karyotype were observed through passage 11. IL-6, IL-10, vascular endothelial growth factor, TGF-β, hepatocyte growth factor, and galectin-1 gene expression levels varied among donors. MSC treatment significantly enhanced islet engraftment and function at 1 month posttransplant (n = 8), as compared with animals that received islets without MSCs (n = 3). Additional infusions of donor or third-party MSCs resulted in reversal of rejection episodes and prolongation of islet function in two animals. Stable islet allograft function was associated with increased numbers of regulatory T-cells in peripheral blood. CONCLUSIONS MSCs may provide an important approach for enhancement of islet engraftment, thereby decreasing the numbers of islets needed to achieve insulin independence. Furthermore, MSCs may serve as a new, safe, and effective antirejection therapy. PMID:20622174

Islet alloautotransplantation: Allogeneic pancreas transplantation followed by transplant pancreatectomy and islet transplantation.

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Nijhoff, M F; Dubbeld, J; van Erkel, A R; van der Boog, P J M; Rabelink, T J; Engelse, M A; de Koning, E J P

2018-04-01

Simultaneous pancreas-kidney (SPK) transplantation is an important treatment option for patients with type 1 diabetes (T1D) and end-stage renal disease (ESRD). Due to complications, in up to 10% of patients, allograft pancreatectomy is necessary shortly after transplantation. Usually the donor pancreas is discarded. Here, we report on a novel procedure to rescue endocrine tissue after allograft pancreatectomy. A 39-year-old woman with T1D and ESRD who had undergone SPK transplantation required emergency allograft pancreatectomy due to bleeding at the vascular anastomosis. Islets were isolated from the removed pancreas allograft, and almost 480 000 islet equivalents were infused into the portal vein. The patient recovered fully. After 3 months, near-normal mixed meal test (fasting glucose 7.0 mmol/L, 2-hour glucose 7.5 mmol/L, maximal stimulated C-peptide 3.25 nmol/L, without insulin use in the preceding 36 hours) was achieved. Glycated hemoglobin while taking a low dose of long-acting insulin was 32.7 mmol/mol hemoglobin (5.3%). When a donor pancreas is lost after transplantation, rescue β cell therapy by islet alloautotransplantation enables optimal use of scarce donor pancreata to optimize glycemic control without additional HLA alloantigen exposure. © 2017 The American Society of Transplantation and the American Society of Transplant Surgeons.

Supravital dithizone staining in the isolation of human and rat pancreatic islets

DEFF Research Database (Denmark)

Hansen, W A; Christie, M R; Kahn, R

1989-01-01

Dithizone, a zinc chelating agent, is known to selectively stain the islets of Langerhans in the pancreas. In the present study, we have used this stain to aid the identification of islets in material obtained by collagenase digestion of human pancreas. Islets were shown to rapidly and reversibly...... techniques for the large scale isolation of functionally intact human islets....

Endothelial dysfunction in metabolic and vascular disorders.

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Polovina, Marija M; Potpara, Tatjana S

2014-03-01

Vascular endothelium has important regulatory functions in the cardiovascular system and a pivotal role in the maintenance of vascular health and metabolic homeostasis. It has long been recognized that endothelial dysfunction participates in the pathogenesis of atherosclerosis from early, preclinical lesions to advanced, thrombotic complications. In addition, endothelial dysfunction has been recently implicated in the development of insulin resistance and type 2 diabetes mellitus (T2DM). Considering that states of insulin resistance (eg, metabolic syndrome, impaired fasting glucose, impaired glucose tolerance, and T2DM) represent the most prevalent metabolic disorders and risk factors for atherosclerosis, it is of considerable scientific and clinical interest that both metabolic and vascular disorders have endothelial dysfunction as a common background. Importantly, endothelial dysfunction has been associated with adverse outcomes in patients with established cardiovascular disease, and a growing body of evidence indicates that endothelial dysfunction also imparts adverse prognosis in states of insulin resistance. In this review, we discuss the association of insulin resistance and T2DM with endothelial dysfunction and vascular disease, with a focus on the underlying mechanisms and prognostic implications of the endothelial dysfunction in metabolic and vascular disorders. We also address current therapeutic strategies for the improvement of endothelial dysfunction.

Ultraviolet light immunomodulation of canine islets for prolongation of allograft survival

International Nuclear Information System (INIS)

Kenyon, N.S.; Strasser, S.; Alejandro, R.

1990-01-01

Ultraviolet (UV) light treatment of donor islets has been shown to be effective for the prolongation of islet allograft survival in rodent models. This study evaluated UV as an immunomodulator of canine islets. The effects of UV irradiation on islet secretory function in vitro revealed a trend of increasing basal insulin release with increasing doses of UV and a corresponding significant decrease in glucose-mediated insulin release (expressed as percentage of basal fractional insulin release) beginning at UV light exposures of 200-300 J/m2 (n = 3, P less than 0.05). Proliferative responses to UV-irradiated allogeneic peripheral blood leukocytes and islets were significantly decreased by 53-112% (P less than 0.05) in 27 of 29 mixed-lymphocyte cultures and by 35-74% (P less than 0.05) in 4 of 5 mixed-lymphocyte islet culture experiments, respectively, beginning at 200-600 J/m2. Autotransplantation of nonirradiated (n = 8) and irradiated islets (600 J/m2, n = 6) resulted in a 1-mo graft survival rate of 75% for the control group and 50% for the irradiated group. Allotransplantation of irradiated islets (600 J/m2) into either nonimmunosuppressed recipients (1 donor to 1 recipient, n = 8) or recipients of subimmunosuppressive doses of cyclosporin (2 donors to 1 recipient, n = 4) resulted in 100% rejection by day 10. In contrast, when islets were cultured for 24 h postirradiation and transplanted into cyclosporin-treated pancreatectomized recipients (2 donors to 1 recipient), 3 of 7 grafts were prolonged beyond day 10 to days 16, 26, and greater than 100

Cost and clinical outcome of islet transplantation in Norway 2010-2015.

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Schive, Simen W; Foss, Aksel; Sahraoui, Afaf; Kloster-Jensen, Kristine; Hafsahl, Geir; Kvalheim, Gunnar; Lundgren, Torbjørn; von Zur-Mühlen, Bengt; Felldin, Marie; Rafael, Ehab; Lempinen, Marko; Korsgren, Olle; Jenssen, Trond G; Mishra, Vinod; Scholz, Hanne

2017-01-01

Islet transplantation is a minimally invasive β-cell replacement strategy. Islet transplantation is a reimbursed treatment in Norway. Here, we summarize the cost and clinical outcome of 31 islet transplantations performed at Oslo University Hospital (OUS) from January 2010 to June 2015. Patients were retrospectively divided into three groups. Thirteen patients received either one or two islet transplantation alone (ITA), while five patients received islet transplantation after previous solid organ transplantation. For the group receiving 2 ITA, Kaplan-Meier estimates show an insulin independence of 20% more than 4 years after their last transplantation. An estimated 70% maintain at least partial graft function, defined as fasting C-peptide >0.1 nmol L -1 , and 47% maintain a HbA1c below 6.5% or 2 percent points lower than before ITA. For all groups combined, we estimate that 44% of the patients have a 50% reduction in insulin requirement 4 years after the initial islet transplantation. The average cost for an islet transplantation procedure was 347 297±60 588 NOK, or 35 424±6182 EUR, of which isolation expenses represent 34%. We hereby add to the common pool of growing experience with islet transplantation and also describe the cost of the treatment at our center. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Optimal formation of genetically modified and functional pancreatic islet spheroids by using hanging-drop strategy.

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Kim, H J; Alam, Z; Hwang, J W; Hwang, Y H; Kim, M J; Yoon, S; Byun, Y; Lee, D Y

2013-03-01

Rejection and hypoxia are important factors causing islet loss at an early stage after pancreatic islet transplantation. Recently, islets have been dissociated into single cells for reaggregation into so-called islet spheroids. Herein, we used a hanging-drop strategy to form islet spheroids to achieve functional equivalence to intact islets. To obtain single islet cells, we dissociated islets with trypsin-EDTA digestion for 10 minutes. To obtain spheroids, we dropped various numbers of single cells (125, 250, or 500 cells/30 μL drop) onto a Petri dish, that was inverted for incubation in humidified air containing 5% CO(2) at 37 °C for 7 days. The aggregated spheroids in the droplets were harvested for further culture. The size of the aggregated islet spheroids depended on the number of single cells (125-500 cells/30 μL droplet). Their morphology was similar to that of intact islets without any cellular damage. When treated with various concentrations of glucose to evaluate responsiveness, their glucose-mediated stimulation index value was similar to that of intact islets, an observation that was attributed to strong cell-to-cell interactions in islet spheroids. However, islet spheroids aggregated in general culture dishes showed abnormal glucose responsiveness owing to weak cell-to-cell interactions. Cell-to-cell interactions in islet spheroids were confirmed with an anti-connexin-36 monoclonal antibody. Finally, nonviral poly(ethylene imine)-mediated interleukin-10 cytokine gene delivered beforehand into dissociated single cells before formation of islet spheroids increased the gene transfection efficacy and interleukin-10 secretion from islet spheroids >4-fold compared with intact islets. These results demonstrated the potential application of genetically modified, functional islet spheroids with of controlled size and morphology using an hanging-drop technique. Copyright © 2013 Elsevier Inc. All rights reserved.

Impact of adverse pancreatic injury at surgical procurement upon islet isolation outcome.

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Andres, Axel; Kin, Tatsuya; O'Gorman, Doug; Bigam, David; Kneteman, Norman; Senior, Peter; Shapiro, Am James

2014-11-01

The consequence of a pancreas injury during the procurement for islet isolation purpose is unknown. The goal of this work was to assess the injuries of the pancreata procured for islet isolation, and to determine their effect on the islet yield. Between January 2007 and October 2013, we prospectively documented every injury of the pancreata processed in our centre for islet isolation. Injuries involving the main duct were classified as major, the others as minor. Donors' characteristics and islet yields were compared between the groups of injuries. A pancreas injury was identified in 42 of 452 pancreata received for islet isolation (9.3%). In 15 cases, the injury was major (3.3% of all pancreata). Although a minor injury did not affect the islet yield, a major injury was significantly associated with unfavourable outcomes (postpurification mean islet equivalent of 364 ± 181, 405 ± 190 and 230 ± 115 × 10(3) for absence of injury, minor injury and major injury, respectively). A major injury was significantly more prevalent in lean and short donors. We recommend assessing the quality of the pancreas in the islet isolation centre before starting the isolation procedure. Each centre should determine its own policy based on its financial resources and on the wait list. © 2014 Steunstichting ESOT.

Lung-Derived Microscaffolds Facilitate Diabetes Reversal after Mouse and Human Intraperitoneal Islet Transplantation.

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Abualhassan, Nasser; Sapozhnikov, Lena; Pawlick, Rena L; Kahana, Meygal; Pepper, Andrew R; Bruni, Antonio; Gala-Lopez, Boris; Kin, Tatsuya; Mitrani, Eduardo; Shapiro, A M James

2016-01-01

There is a need to develop three-dimensional structures that mimic the natural islet tissue microenvironment. Endocrine micro-pancreata (EMPs) made up of acellular organ-derived micro-scaffolds seeded with human islets have been shown to express high levels of key beta-cell specific genes and secrete quantities of insulin per cell similar to freshly isolated human islets in a glucose-regulated manner for more than three months in vitro. The aim of this study was to investigate the capacity of EMPs to restore euglycemia in vivo after transplantation of mouse or human islets in chemically diabetic mice. We proposed that the organ-derived EMPs would restore the extracellular components of the islet microenvironment, generating favorable conditions for islet function and survival. EMPs seeded with 500 mouse islets were implanted intraperitoneally into streptozotocin-induced diabetic mice and reverted diabetes in 67% of mice compared to 13% of controls (p = 0.018, n = 9 per group). Histological analysis of the explanted grafts 60 days post-transplantation stained positive for insulin and exhibited increased vascular density in a collagen-rich background. EMPs were also seeded with human islets and transplanted into the peritoneal cavity of immune-deficient diabetic mice at 250 islet equivalents (IEQ), 500 IEQ and 1000 IEQ. Escalating islet dose increased rates of normoglycemia (50% of the 500 IEQ group and 75% of the 1000 IEQ group, n = 3 per group). Human c-peptide levels were detected 90 days post-transplantation in a dose-response relationship. Herein, we report reversal of diabetes in mice by intraperitoneal transplantation of human islet seeded on EMPs with a human islet dose as low as 500 IEQ.

Design of bioartificial pancreas with functional micro/nano-based encapsulation of islets.

Science.gov (United States)

Kepsutlu, Burcu; Nazli, Caner; Bal, Tugba; Kizilel, Seda

2014-01-01

Type I diabetes mellitus (TIDM), a devastating health issue in all over the world, has been treated by successful transplantation of insulin secreting pancreatic islets. However, serious limitations such as the requirement of immunosuppressive drugs for recipient patients, side effects as a result of long-term use of drugs, and reduced functionality of islets at the transplantation site remain. Bioartificial pancreas that includes islets encapsulated within semi-permeable membrane has been considered as a promising approach to address these requirements. Many studies have focused on micro or nanobased islet immunoisolation systems and tested the efficacy of encapsulated islets using in vitro and in vivo platforms. In this review, we address current progress and obstacles for the development of a bioartificial pancreas using micro/nanobased systems for encapsulation of islets.

Islet-specific T cell clones transfer diabetes to nonobese diabetic (NOD) F1 mice.

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Peterson, J D; Pike, B; McDuffie, M; Haskins, K

1994-09-15

To investigate diabetes resistance to T cell-mediated disease transfer, we administered islet-specific T cell clones to the F1 progeny of nonobese diabetic (NOD) mice that were crossed with various nondiabetes-prone inbred mouse strains. We investigated four diabetogenic CD4+ T cell clones and all induced insulitis and full development of diabetes in (SWR x NOD)F1, (SJL x NOD)F1, and (C57BL/6 x NOD)F1 mice. In contrast, (BALB/c x NOD)F1 and (CBA x NOD)F1 mice were susceptible to disease transfer by some T cell clones but not others, and (C57/L x NOD)F1 mice seemed to be resistant to both insulitis and disease transfer by all of the clones tested. Disease induced by the T cell clones in susceptible F1 strains was age dependent and could only be observed in recipients younger than 13 days old. Full or partial disease resistance did not correlate with the presence or absence of I-E, different levels of Ag expression in islet cells, or differences in APC function. The results from this study suggest that there may be multiple factors contributing to susceptibility of F1 mice to T cell clone-mediated induction of diabetes, including non-MHC-related genetic background, the immunologic maturity of the recipient, and individual characteristics of the T cell clones.

Encapsulation of pancreatic islets for transplantation in diabetes : the untouchable islets

NARCIS (Netherlands)

de Vos, P; Marchetti, P

The aim of encapsulation of pancreatic islets is to transplant in the absence of immunosuppression. It is based on the principle that transplanted tissue is protected from the host immune system by an artificial membrane. Encapsulation allows for application of insulin-secreting cells of animal or

Islet transplantation in diabetic rats normalizes basal and exercise-induced energy metabolism

NARCIS (Netherlands)

Houwing, Harmina; Benthem, L.; Suylichem, P.T.R. van; Leest, J. van der; Strubbe, J.H.; Steffens, A.B.

Transplantation of islets of Langerhans in diabetic rats normalizes resting glucose and insulin levels, but it remains unclear whether islet transplantation restores resting and exercise-induced energy metabolism. Therefore, we compared energy metabolism in islet transplanted rats with energy

mTOR Inhibition and Clinical Transplantation: Pancreas and Islet.

Science.gov (United States)

Berney, Thierry; Andres, Axel; Toso, Christian; Majno, Pietro; Squifflet, Jean-Paul

2018-02-01

This brief overview discusses the beneficial and deleterious effects of mammalian target of rapamycin (mTOR) inhibitors on β cells, and how sirolimus- and everolimus-based immunosuppression have impacted on practices and outcomes of pancreas and islet transplantation. Sirolimus was the cornerstone of immunosuppressive regimens in islet transplantation at the turn of the millenium, but utilization of mTOR inhibitors has progressively decreased from greater than 80% to less than 50% of islet transplant recipients in more recent years. For whole pancreas transplantation, mTOR inhibitors were used in approximately 20% of patients in the early 2000s, but this dropped over the years to less than 10% currently. This decrease is arguably due to less well-tolerated side effects without the advantage of better outcomes. Nonetheless, mTOR inhibitors remain extremely valuable as second-line immunosuppressants in pancreas and islet transplantation.

Streptozotocin Diabetes CORRELATION WITH EXTENT OF DEPRESSION OF PANCREATIC ISLET NICOTINAMIDE ADENINE DINUCLEOTIDE

Science.gov (United States)

Anderson, Tom; Schein, Philip S.; McMenamin, Mary G.; Cooney, David A.

1974-01-01

The diabetogenic activity of streptozotocin has been correlated with a reduction in pyridine nucleotide synthesis in the mouse pancreatic islet. To determine the specificity of this reduction for diabetogenicity, a comparative study of streptozotocin, its cytotoxic moiety, 1-methyl-1-nitrosourea, and alloxan was performed. Streptozotocin administered intraperitoneally (i.p.) producd a dose-related reduction in islet NAD which was proportional to the degree of diabetogenicity. A diabetogenic dose, 200 mg/kg, attained a peak plasma N-nitroso intact streptozotocin concentration of 0.224 μmol/ml and reduced the mean islet NAD from a control of 0.78 to 0.15 pmol. At borderline, 150 mg/kg, and nondiabetogenic, 100 mg/kg, doses, plasma concentrations reached 0.161 and 0.136 μmol/ml, and NAD was 0.36 and 0.86 pmol/islet, respectively. 1-Methyl-1-nitrosourea, 100 mg/kg, attained a maximum N-nitroso intact 1-methyl-1-nitrosourea concentration of 0.162 μmol/ml and reduced the mean NAD to 0.58 pmol/islet, and was nondiabetogenic; 200 mg/kg attained a peak plasma concentration of 0.344 μmol/ml and depressed NAD to 0.38 pmol/islet, and was inconsistently diabetogenic. Islet NAD of 0.4 pmol/islet or greater is required for integrity of the beta cell. A diabetogenic dose of alloxan, 500 mg/kg, did not depress NAD, 0.85 pmol/islet, therefore confirming that its mechanism of diabetogenicity differs from that of streptozotocin. In vivo uptake of [methyl-14C]streptozotocin by islets was 3.8 times that of [methyl-14C]-1-methyl-1-nitrosourea, whereas uptake by the exocrine pancreas favored 1-methyl-1-nitrosourea over streptozotocin 2.4:1. The decreased islet uptake of 1-methyl-1-nitrosourea correlates with the 3.5 times increased molar dosage required to produce islet NAD depression comparable to that of streptozotocin, 150 mg/kg. These studies indicate that the glucose carrier of streptozotocin facilitates uptake of its cytotoxic group, 1-methyl-1-nitrosourea, into islets. PMID

Engineering of microscale three-dimensional pancreatic islet models in vitro and their biomedical applications.

Science.gov (United States)

Gao, Bin; Wang, Lin; Han, Shuang; Pingguan-Murphy, Belinda; Zhang, Xiaohui; Xu, Feng

2016-08-01

Diabetes now is the most common chronic disease in the world inducing heavy burden for the people's health. Based on this, diabetes research such as islet function has become a hot topic in medical institutes of the world. Today, in medical institutes, the conventional experiment platform in vitro is monolayer cell culture. However, with the development of micro- and nano-technologies, several microengineering methods have been developed to fabricate three-dimensional (3D) islet models in vitro which can better mimic the islet of pancreases in vivo. These in vitro islet models have shown better cell function than monolayer cells, indicating their great potential as better experimental platforms to elucidate islet behaviors under both physiological and pathological conditions, such as the molecular mechanisms of diabetes and clinical islet transplantation. In this review, we present the state-of-the-art advances in the microengineering methods for fabricating microscale islet models in vitro. We hope this will help researchers to better understand the progress in the engineering 3D islet models and their biomedical applications such as drug screening and islet transplantation.

Serum Cytokines as Biomarkers in Islet Cell Transplantation for Type 1 Diabetes

NARCIS (Netherlands)

van der Torren, Cornelis R; Verrijn Stuart, Annemarie A; Lee, DaHae; Meerding, Jenny; van de Velde, Ursule; Pipeleers, Daniel; Gillard, Pieter; Keymeulen, Bart; de Jager, Wilco; Roep, Bart O

2016-01-01

BACKGROUND: Islet cell transplantation holds a potential cure for type 1 diabetes, but many islet recipients do not reach long-lasting insulin independence. In this exploratory study, we investigated whether serum cytokines, chemokines and adipokines are associated with the clinical outcome of islet

Vitality of pancreatic islets labeled for magnetic resonance imaging with iron particles.

Science.gov (United States)

Berkova, Z; Kriz, J; Girman, P; Zacharovova, K; Koblas, T; Dovolilova, E; Saudek, F

2005-10-01

We previously described an in vivo method for pancreatic islet visualization using magnetic resonance imaging with the aid of superparamagnetic nanoparticles of iron oxide (Resovist) or by magnetic beads precoated with antibodies (Dynabeads). The aim of this study was to investigate the in vitro effect of islet labeling on their quality. Isolated rat islets were cultivated for 48 hours with a contrast agent or, in the case of magnetic antibody-coated beads, for only 2 hours. The ability to secrete insulin was tested by a static insulin release assay and the results were expressed as a stimulation index. Staining with propidium iodide and acridine orange was performed to determine the ratio of live to dead cells. Stimulation indices in the Resovist islets (n = 23) vs controls (n = 14) were 15.3 and 15.0, respectively, and in the Dynabeads islets (n = 15) vs controls (n = 12) 21.3 and 19.9, respectively. The vitality of the Resovist islets vs controls determined by live/dead cells ratio was 90.8% and 91.1%, respectively (n = 20), and in the Dynabeads islets vs controls was 89.4% and 91.8%, respectively (n = 11). Islet labeling with the contrast agent as well as with specific antibodies with iron beads did not change the vitality and insulin-secreting capacity assessed in vitro (P > .05). Magnetic resonance using iron nanoparticles represents the only method for in-vivo visualization of transplanted islets so far. Our data represent an important contribution for its clinical use.

Pancreatic Islet Protein Complexes and Their Dysregulation in Type 2 Diabetes

DEFF Research Database (Denmark)

Pedersen, Helle Krogh; Gudmundsdottir, Valborg; Brunak, Søren

2017-01-01

Type 2 diabetes (T2D) is a complex disease that involves multiple genes. Numerous risk loci have already been associated with T2D, although many susceptibility genes remain to be identified given heritability estimates. Systems biology approaches hold potential for discovering novel T2D genes...... by considering their biological context, such as tissue-specific protein interaction partners. Pancreatic islets are a key T2D tissue and many of the known genetic risk variants lead to impaired islet function, hence a better understanding of the islet-specific dysregulation in the disease-state is essential...... to unveil the full potential of person-specific profiles. Here we identify 3,692 overlapping pancreatic islet protein complexes (containing 10,805 genes) by integrating islet gene and protein expression data with protein interactions. We found 24 of these complexes to be significantly enriched for genes...

Imatinib prevents beta cell death in vitro but does not improve islet transplantation outcome.

Science.gov (United States)

King, Aileen J F; Griffiths, Lisa A; Persaud, Shanta J; Jones, Peter M; Howell, Simon L; Welsh, Nils

2016-05-01

Introduction Improving islet transplantation outcome could not only bring benefits to individual patients but also widen the patient pool to which this life-changing treatment is available. Imatinib has previously been shown to protect beta cells from apoptosis in a variety of in vitro and in vivo models. The aim of this study was to investigate whether imatinib could be used to improve islet transplantation outcome. Methods Islets were isolated from C57Bl/6 mice and pre-cultured with imatinib prior to exposure to streptozotocin and cytokines in vitro. Cell viability and glucose-induced insulin secretion were measured. For transplantation experiments, islets were pre-cultured with imatinib for either 72 h or 24 h prior to transplantation into streptozotocin-diabetic C57Bl/6 mice. In one experimental series mice were also administered imatinib after islet transplantation. Results Imatinib partially protected islets from beta cell death in vitro. However, pre-culturing islets in imatinib or administering the drug to the mice in the days following islet transplantation did not improve blood glucose concentrations more than control-cultured islets. Conclusion Although imatinib protected against beta cell death from cytokines and streptozotocin in vitro, it did not significantly improve syngeneic islet transplantation outcome.

The Spleen Is an Ideal Site for Inducing Transplanted Islet Graft Expansion in Mice.

Directory of Open Access Journals (Sweden)

Takeshi Itoh

Full Text Available Alternative islet transplantation sites have the potential to reduce the marginal number of islets required to ameliorate hyperglycemia in recipients with diabetes. Previously, we reported that T cell leukemia homeobox 1 (Tlx1+ stem cells in the spleen effectively regenerated into insulin-producing cells in the pancreas of non-obese diabetic mice with end-stage disease. Thus, we investigated the spleen as a potential alternative islet transplantation site. Streptozotocin-induced diabetic C57BL/6 mice received syngeneic islets into the portal vein (PV, beneath the kidney capsule (KC, or into the spleen (SP. The marginal number of islets by PV, KC, or SP was 200, 100, and 50, respectively. Some plasma inflammatory cytokine levels in the SP group were significantly lower than those of the PV group after receiving a marginal number of islets, indicating reduced inflammation in the SP group. Insulin contents were increased 280 days after islet transplantation compared with those immediately following transplantation (p<0.05. Additionally, Tlx1-related genes, including Rrm2b and Pla2g2d, were up-regulated, which indicates that islet grafts expanded in the spleen. The spleen is an ideal candidate for an alternative islet transplantation site because of the resulting reduced inflammation and expansion of the islet graft.

Photochemical (PUVA) treatment of isolated rat islets

International Nuclear Information System (INIS)

Schmidt, S.; Wilke, B.; Kloeting, I.

1984-01-01

Isolated rat islets were irradiated with long-wave ultraviolet light alone or in combination with the photosensitizer 8-methoxypsoralen. The influence on specific beta cell functions was determined with the aim to find out experimental conditions which allow the use of such islets for transplantation. Short-term effects: Ultraviolet light affected [ 3 H]leucine incorporation into (pro)insulin (5 J/cm 2 : 53.8 %, 10 J/cm 2 : 41.0 % of the controls) and insulin release was slightly reduced. 8-methoxypsoralen enhanced the irradiation effect. Long-term effects: A restoration of irradiation-affected beta cell function was detected after 5 days of culture unless the dose exceeded 2 J/cm 2 (0.1 μM 8-methoxypsoralen) or 1 J/cm 2 (1 μM 8-methoxypsoralen). After functional restoration islets were used for transplantation experiments. (author)

Photochemical (PUVA) treatment of isolated rat islets

Energy Technology Data Exchange (ETDEWEB)

Schmidt, S; Wilke, B; Kloeting, I [Zentralinstitut fuer Diabetes, Karlsburg (German Democratic Republic)

1984-05-01

Isolated rat islets were irradiated with long-wave ultraviolet light alone or in combination with the photosensitizer 8-methoxypsoralen. The influence on specific beta cell functions was determined with the aim to find out experimental conditions which allow the use of such islets for transplantation. Short-term effects: Ultraviolet light affected (/sup 3/H)leucine incorporation into (pro)insulin (5 J/cm/sup 2/ : 53.8 %, 10 J/cm/sup 2/ : 41.0 % of the controls) and insulin release was slightly reduced. 8-methoxypsoralen enhanced the irradiation effect. Long-term effects: A restoration of irradiation-affected beta cell function was detected after 5 days of culture unless the dose exceeded 2 J/cm/sup 2/ (0.1 ..mu..M 8-methoxypsoralen) or 1 J/cm/sup 2/ (1 ..mu..M 8-methoxypsoralen). After functional restoration islets were used for transplantation experiments.

Preservation of beta cell function in adult human pancreatic islets for several months in vitro

DEFF Research Database (Denmark)

Brunstedt, J; Andersson, A; Frimodt-Møller, C

1979-01-01

Islets of Langerhans were isolated from four human kidney donors, aged 16 to 21 years by the collagenase method described for isolation of rodent islets. So far the human islets have been kept in tissue culture, without attachment, in medium RPMI 1640 supplemented with 10% calf serum for more tha...... technique presents a valuable tool for studying chronic effects of metabolites and hormones on islet function, as well as for islet storage prior to transplantation into humans.......Islets of Langerhans were isolated from four human kidney donors, aged 16 to 21 years by the collagenase method described for isolation of rodent islets. So far the human islets have been kept in tissue culture, without attachment, in medium RPMI 1640 supplemented with 10% calf serum for more than...

Adipose stem cells from chronic pancreatitis patients improve mouse and human islet survival and function.

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Song, Lili; Sun, Zhen; Kim, Do-Sung; Gou, Wenyu; Strange, Charlie; Dong, Huansheng; Cui, Wanxing; Gilkeson, Gary; Morgan, Katherine A; Adams, David B; Wang, Hongjun

2017-08-30

Chronic pancreatitis has surgical options including total pancreatectomy to control pain. To avoid surgical diabetes, the explanted pancreas can have islets harvested and transplanted. Immediately following total pancreatectomy with islet autotransplantation (TP-IAT), many islet cells die due to isolation and transplantation stresses. The percentage of patients remaining insulin free after TP-IAT is therefore low. We determined whether cotransplantation of adipose-derived mesenchymal stem cells (ASCs) from chronic pancreatitis patients (CP-ASCs) would protect islets after transplantation. In a marginal mass islet transplantation model, islets from C57BL/6 mice were cotransplanted with CP-ASCs into syngeneic streptozotocin-treated diabetic mice. Treatment response was defined by the percentage of recipients reaching normoglycemia, and by the area under the curve for glucose and c-peptide in a glucose tolerance test. Macrophage infiltration, β-cell apoptosis, and islet graft vasculature were measured in transplanted islet grafts by immunohistochemistry. mRNA expression profiling of 84 apoptosis-related genes in islet grafts transplanted alone or with CP-ASCs was measured by the RT 2 Profiler™ Apoptosis PCR Array. The impact of insulin-like growth factor-1 (IGF-1) on islet apoptosis was determined in islets stimulated with cytokines (IL-1β and IFN-γ) in the presence and absence of CP-ASC conditioned medium. CP-ASC-treated mice were more often normoglycemic compared to mice receiving islets alone. ASC cotransplantation reduced macrophage infiltration, β-cell death, suppressed expression of TNF-α and Bcl-2 modifying factor (BMF), and upregulated expressions of IGF-1 and TNF Receptor Superfamily Member 11b (TNFRSF11B) in islet grafts. Islets cultured in conditioned medium from CP-ASCs showed reduced cell death. This protective effect was diminished when IGF-1 was blocked in the conditioned medium by the anti-IGF-1 antibody. Cotransplantation of islets with ASCs

Foodborne cereulide causes beta-cell dysfunction and apoptosis.

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Roman Vangoitsenhoven

Full Text Available To study the effects of cereulide, a food toxin often found at low concentrations in take-away meals, on beta-cell survival and function.Cell death was quantified by Hoechst/Propidium Iodide in mouse (MIN6 and rat (INS-1E beta-cell lines, whole mouse islets and control cell lines (HepG2 and COS-1. Beta-cell function was studied by glucose-stimulated insulin secretion (GSIS. Mechanisms of toxicity were evaluated in MIN6 cells by mRNA profiling, electron microscopy and mitochondrial function tests.24 h exposure to 5 ng/ml cereulide rendered almost all MIN6, INS-1E and pancreatic islets apoptotic, whereas cell death did not increase in the control cell lines. In MIN6 cells and murine islets, GSIS capacity was lost following 24 h exposure to 0.5 ng/ml cereulide (P<0.05. Cereulide exposure induced markers of mitochondrial stress including Puma (p53 up-regulated modulator of apoptosis, P<0.05 and general pro-apoptotic signals as Chop (CCAAT/-enhancer-binding protein homologous protein. Mitochondria appeared swollen upon transmission electron microscopy, basal respiration rate was reduced by 52% (P<0.05 and reactive oxygen species increased by more than twofold (P<0.05 following 24 h exposure to 0.25 and 0.50 ng/ml cereulide, respectively.Cereulide causes apoptotic beta-cell death at low concentrations and impairs beta-cell function at even lower concentrations, with mitochondrial dysfunction underlying these defects. Thus, exposure to cereulide even at concentrations too low to cause systemic effects appears deleterious to the beta-cell.

Differences in glucose-stimulated insulin secretion in vitro of islets from human, nonhuman primate, and porcine origin.

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Mueller, Kate R; Balamurugan, A N; Cline, Gary W; Pongratz, Rebecca L; Hooper, Rebecca L; Weegman, Bradley P; Kitzmann, Jennifer P; Taylor, Michael J; Graham, Melanie L; Schuurman, Henk-Jan; Papas, Klearchos K

2013-01-01

Porcine islet xenotransplantation is considered a potential cell-based therapy for type 1 diabetes. It is currently being evaluated in diabetic nonhuman primates (NHP) to assess safety and efficacy of the islet product. However, due to a variety of distinct differences between the respective species, including the insulin secretory characteristics of islets, the suitability and predictive value of the preclinical model in the extrapolation to the clinical setting remain a critical issue. Islets isolated from human (n = 3), NHP (n = 2), adult pig (AP, n = 3), and juvenile pig (JP, n = 4) pancreata were perifused with medium at basal glucose (2.5 mm) followed by high glucose (16.7 mm) concentrations. The total glucose-stimulated insulin secretion (GSIS) was calculated from generated insulin secretion profiles. Nonhuman primate islets exhibited GSIS 3-fold higher than AP islets, while AP and JP islets exhibited GSIS 1/3 and 1/30 of human islets, respectively. The insulin content of NHP and AP islets was similar to that of human islets, whereas that of JP islets was 1/5 of human islets. Despite the fact that human, NHP, and AP islets contain similar amounts of insulin, the much higher GSIS for NHP islets than for AP and JP islets suggests the need for increased dosing of islets from JP and AP in pig-to-NHP transplantation. Porcine islet xenotransplantation to humans may require significantly higher dosing given the lower GSIS of AP islets compared to human islets. © 2013 John Wiley & Sons A/S.

Maternal Diet Supplementation with n-6/n-3 Essential Fatty Acids in a 1.2 : 1.0 Ratio Attenuates Metabolic Dysfunction in MSG-Induced Obese Mice

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Josiane Morais Martin

2016-01-01

Full Text Available Essential polyunsaturated fatty acids (PUFAs prevent cardiometabolic diseases. We aimed to study whether a diet supplemented with a mixture of n-6/n-3 PUFAs, during perinatal life, attenuates outcomes of long-term metabolic dysfunction in prediabetic and obese mice. Seventy-day-old virgin female mice were mated. From the conception day, dams were fed a diet supplemented with sunflower oil and flaxseed powder (containing an n-6/n-3 PUFAs ratio of 1.2 : 1.0 throughout pregnancy and lactation, while control dams received a commercial diet. Newborn mice were treated with monosodium L-glutamate (MSG, 4 mg g−1 body weight per day for the first 5 days of age. A batch of weaned pups was sacrificed to quantify the brain and pancreas total lipids; another batch were fed a commercial diet until 90 days of age, where glucose homeostasis and glucose-induced insulin secretion (GIIS as well as retroperitoneal fat and Lee index were assessed. MSG-treated mice developed obesity, glucose intolerance, insulin resistance, pancreatic islet dysfunction, and higher fat stores. Maternal flaxseed diet-supplementation decreased n-6/n-3 PUFAs ratio in the brain and pancreas and blocked glucose intolerance, insulin resistance, GIIS impairment, and obesity development. The n-6/n-3 essential PUFAs in a ratio of 1.2 : 1.0 supplemented in maternal diet during pregnancy and lactation prevent metabolic dysfunction in MSG-obesity model.

Endothelial microparticles released by activated protein C protect beta cells through EPCR/PAR1 and annexin A1/FPR2 pathways in islets.

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Kreutter, Guillaume; Kassem, Mohamad; El Habhab, Ali; Baltzinger, Philippe; Abbas, Malak; Boisrame-Helms, Julie; Amoura, Lamia; Peluso, Jean; Yver, Blandine; Fatiha, Zobairi; Ubeaud-Sequier, Geneviève; Kessler, Laurence; Toti, Florence

2017-11-01

Islet transplantation is associated with early ischaemia/reperfusion, localized coagulation and redox-sensitive endothelial dysfunction. In animal models, islet cytoprotection by activated protein C (aPC) restores islet vascularization and protects graft function, suggesting that aPC triggers various lineages. aPC also prompts the release of endothelial MP that bear EPCR, its specific receptor. Microparticles (MP) are plasma membrane procoagulant vesicles, surrogate markers of stress and cellular effectors. We measured the cytoprotective effects of aPC on endothelial and insulin-secreting Rin-m5f β-cells and its role in autocrine and paracrine MP-mediated cell crosstalk under conditions of oxidative stress. MP from aPC-treated primary endothelial (EC) or β-cells were applied to H 2 O 2 -treated Rin-m5f. aPC activity was measured by enzymatic assay and ROS species by dihydroethidium. The capture of PKH26-stained MP and the expression of EPCR were probed by fluorescence microscopy and apoptosis by flow cytometry. aPC treatment enhanced both annexin A1 (ANXA1) and PAR-1 expression in EC and to a lesser extent in β-cells. MP from aPC-treated EC (eM aPC ) exhibited high EPCR and annexin A1 content, protected β-cells, restored insulin secretion and were captured by 80% of β cells in a phosphatidylserine and ANXA1-dependent mechanism. eMP activated EPCR/PAR-1 and ANXA1/FPR2-dependent pathways and up-regulated the expression of EPCR, and of FPR2/ALX, the ANXA1 receptor. Cytoprotection was confirmed in H 2 O 2 -treated rat islets with increased viability (62% versus 48% H 2 O 2 ), reduced apoptosis and preserved insulin secretion in response to glucose elevation (16 versus 5 ng/ml insulin per 10 islets). MP may prove a promising therapeutic tool in the protection of transplanted islets. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Ancestral genomic duplication of the insulin gene in tilapia: An analysis of possible implications for clinical islet xenotransplantation using donor islets from transgenic tilapia expressing a humanized insulin gene.

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Hrytsenko, Olga; Pohajdak, Bill; Wright, James R

2016-07-03

Tilapia, a teleost fish, have multiple large anatomically discrete islets which are easy to harvest, and when transplanted into diabetic murine recipients, provide normoglycemia and mammalian-like glucose tolerance profiles. Tilapia insulin differs structurally from human insulin which could preclude their use as islet donors for xenotransplantation. Therefore, we produced transgenic tilapia with islets expressing a humanized insulin gene. It is now known that fish genomes may possess an ancestral duplication and so tilapia may have a second insulin gene. Therefore, we cloned, sequenced, and characterized the tilapia insulin 2 transcript and found that its expression is negligible in islets, is not islet-specific, and would not likely need to be silenced in our transgenic fish.

High glucose suppresses human islet insulin biosynthesis by inducing miR-133a leading to decreased polypyrimidine tract binding protein-expression

DEFF Research Database (Denmark)

Fred, Rikard G; Bang-Berthelsen, Claus H; Mandrup-Poulsen, Thomas

2010-01-01

BACKGROUND: Prolonged periods of high glucose exposure results in human islet dysfunction in vitro. The underlying mechanisms behind this effect of high glucose are, however, unknown. The polypyrimidine tract binding protein (PTB) is required for stabilization of insulin mRNA and the PTB mRNA 3......'-UTR contains binding sites for the microRNA molecules miR-133a, miR-124a and miR-146. The aim of this study was therefore to investigate whether high glucose increased the levels of these three miRNAs in association with lower PTB levels and lower insulin biosynthesis rates. METHODOLOGY...... for real-time RT-PCR analysis of miR-133a, miR-124a, miR-146, insulin mRNA and PTB mRNA contents. Insulin biosynthesis rates were determined by radioactive labeling and immunoprecipitation. Synthetic miR-133a precursor and inhibitor were delivered to dispersed islet cells by lipofection, and PTB...

Retention of gene expression in porcine islets after agarose encapsulation and long-term culture

International Nuclear Information System (INIS)

Dumpala, Pradeep R.; Holdcraft, Robert W.; Martis, Prithy C.; Laramore, Melissa A.; Parker, Thomas S.; Levine, Daniel M.; Smith, Barry H.; Gazda, Lawrence S.

2016-01-01

Agarose encapsulation of porcine islets allows extended in vitro culture, providing ample time to determine the functional capacity of the islets and conduct comprehensive microbiological safety testing prior to implantation as a treatment for type 1 diabetes mellitus. However, the effect that agarose encapsulation and long-term culture may have on porcine islet gene expression is unknown. The aim of the present study was to compare the transcriptome of encapsulated porcine islets following long-term in vitro culture against free islets cultured overnight. Global gene expression analysis revealed no significant change in the expression of 98.47% of genes. This indicates that the gene expression profile of free islets is highly conserved following encapsulation and long-term culture. Importantly, the expression levels of genes that code for critical hormones secreted by islets (insulin, glucagon, and somatostatin) as well as transcripts encoding proteins involved in their packaging and secretion are unchanged. While a small number of genes known to play roles in the insulin secretion and insulin signaling pathways are differentially expressed, our results show that overall gene expression is retained following islet isolation, agarose encapsulation, and long-term culture. - Highlights: • Effect of agarose encapsulation and 8 week culture on porcine islets was analyzed. • Transcriptome analysis revealed no significant change in a majority (98%) of genes. • Agarose encapsulation allows for long-term culture of porcine islets. • Islet culture allows for functional and microbial testing prior to clinical use.

Retention of gene expression in porcine islets after agarose encapsulation and long-term culture

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Dumpala, Pradeep R., E-mail: pdumpala@rixd.org [The Rogosin Institute – Xenia Division, 740 Birch Road, Xenia, OH 45385 (United States); Holdcraft, Robert W.; Martis, Prithy C.; Laramore, Melissa A. [The Rogosin Institute – Xenia Division, 740 Birch Road, Xenia, OH 45385 (United States); Parker, Thomas S.; Levine, Daniel M. [The Rogosin Institute, 505 East 70th Street, New York, NY 10021 (United States); Smith, Barry H. [The Rogosin Institute, 505 East 70th Street, New York, NY 10021 (United States); NewYork-Presbyterian Hospital, Weill Medical College of Cornell University, 1300 York Avenue, New York, NY 10021 (United States); Gazda, Lawrence S. [The Rogosin Institute – Xenia Division, 740 Birch Road, Xenia, OH 45385 (United States)

2016-08-05

Agarose encapsulation of porcine islets allows extended in vitro culture, providing ample time to determine the functional capacity of the islets and conduct comprehensive microbiological safety testing prior to implantation as a treatment for type 1 diabetes mellitus. However, the effect that agarose encapsulation and long-term culture may have on porcine islet gene expression is unknown. The aim of the present study was to compare the transcriptome of encapsulated porcine islets following long-term in vitro culture against free islets cultured overnight. Global gene expression analysis revealed no significant change in the expression of 98.47% of genes. This indicates that the gene expression profile of free islets is highly conserved following encapsulation and long-term culture. Importantly, the expression levels of genes that code for critical hormones secreted by islets (insulin, glucagon, and somatostatin) as well as transcripts encoding proteins involved in their packaging and secretion are unchanged. While a small number of genes known to play roles in the insulin secretion and insulin signaling pathways are differentially expressed, our results show that overall gene expression is retained following islet isolation, agarose encapsulation, and long-term culture. - Highlights: • Effect of agarose encapsulation and 8 week culture on porcine islets was analyzed. • Transcriptome analysis revealed no significant change in a majority (98%) of genes. • Agarose encapsulation allows for long-term culture of porcine islets. • Islet culture allows for functional and microbial testing prior to clinical use.

Integrative analysis of a cross-loci regulation network identifies App as a gene regulating insulin secretion from pancreatic islets.

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Zhidong Tu

Full Text Available Complex diseases result from molecular changes induced by multiple genetic factors and the environment. To derive a systems view of how genetic loci interact in the context of tissue-specific molecular networks, we constructed an F2 intercross comprised of >500 mice from diabetes-resistant (B6 and diabetes-susceptible (BTBR mouse strains made genetically obese by the Leptin(ob/ob mutation (Lep(ob. High-density genotypes, diabetes-related clinical traits, and whole-transcriptome expression profiling in five tissues (white adipose, liver, pancreatic islets, hypothalamus, and gastrocnemius muscle were determined for all mice. We performed an integrative analysis to investigate the inter-relationship among genetic factors, expression traits, and plasma insulin, a hallmark diabetes trait. Among five tissues under study, there are extensive protein-protein interactions between genes responding to different loci in adipose and pancreatic islets that potentially jointly participated in the regulation of plasma insulin. We developed a novel ranking scheme based on cross-loci protein-protein network topology and gene expression to assess each gene's potential to regulate plasma insulin. Unique candidate genes were identified in adipose tissue and islets. In islets, the Alzheimer's gene App was identified as a top candidate regulator. Islets from 17-week-old, but not 10-week-old, App knockout mice showed increased insulin secretion in response to glucose or a membrane-permeant cAMP analog, in agreement with the predictions of the network model. Our result provides a novel hypothesis on the mechanism for the connection between two aging-related diseases: Alzheimer's disease and type 2 diabetes.

Assessment of DNA synthesis in Islet-1+ cells in the adult murine heart

International Nuclear Information System (INIS)

Weinberger, Florian; Mehrkens, Dennis; Starbatty, Jutta; Nicol, Philipp; Eschenhagen, Thomas

2015-01-01

Highlights: • Islet-1 was expressed in the adult heart. • Islet-1-positive cells did not proliferate in the adult heart. • Sinoatrial node cells did not proliferate in the adult heart. - Abstract: Rationale: Islet-1 positive (Islet-1 + ) cardiac progenitor cells give rise to the right ventricle, atria and outflow tract during murine cardiac development. In the adult heart Islet-1 expression is limited to parasympathetic neurons, few cardiomyocytes, smooth muscle cells, within the proximal aorta and pulmonary artery and sinoatrial node cells. Its role in these cells is unknown. Here we tested the hypothesis that Islet-1 + cells retain proliferative activity and may therefore play a role in regenerating specialized regions in the heart. Methods and results: DNA synthesis was analyzed by the incorporation of tritiated thymidine ( 3 H-thymidine) in Isl-1-nLacZ mice, a transgenic model with an insertion of a nuclear beta-galactosidase in the Islet-1 locus. Mice received daily injections of 3 H-thymidine for 5 days. DNA synthesis was visualized throughout the heart by dipping autoradiography of cryosections. Colocalization of an nLacZ-signal and silver grains would indicate DNA synthesis in Islet-1 + cells. Whereas Islet − non-myocyte nuclei were regularly marked by accumulation of silver grains, colocalization with nLacZ-signals was not detected in >25,000 cells analyzed. Conclusions: Islet-1 + cells are quiescent in the adult heart, suggesting that, under normal conditions, even pacemaking cells do not proliferate at higher rates than normal cardiac myocytes

Transplanted human pancreatic islets after long-term insulin independence

DEFF Research Database (Denmark)

Muller, Y D; Gupta, Shashank; Morel, P

2013-01-01

Long-term insulin independence after islets of Langerhans transplantation is rarely achieved. The aims of this study were to identify the histological and immunological features of islets transplanted in a type 1 diabetic patient who died of a cerebral hemorrhage after >13 years insulin independe...

Potential differentiation of islet-like cells from pregnant cow-derived placental stem cells.

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Peng, Shao-Yu; Chou, Chien-Wen; Kuo, Yu-Hsuan; Shen, Perng-Chih; Shaw, S W Steven

2017-06-01

Type 1 diabetes is an autoimmune disease that destroys islet cells and results in insufficient insulin secretion by pancreatic β-cells. Islet transplantation from donors is an approach used for treating patients with diabetes; however, this therapy is difficult to implement because of the lack of donors. Nevertheless, several stem cells have the potential to differentiate from islet-like cells and enable insulin secretion for treating diabetes in animal models. For example, placenta is considered a waste material and can be harvested noninvasively during delivery without ethical or moral concerns. To date, the differentiation of islet-like cells from cow-derived placental stem cells (CPSCs) has yet to be demonstrated. The investigation of potential differentiation of islet-like cells from CPSCs was conducted by supplementation with nicotinamide, exendin-4, glucose, and poly-d-lysine and was detected through reverse transcription polymerase chain reaction, dithizone staining, and immunocytochemical methods. Our results indicated that CPSCs are established and express mesenchymal stem cell surface antigen markers, such as CD73, CD166, β-integrin, and Oct-4, but not hematopoietic stem cell surface antigen markers, such as CD45. After induction, the CPSCs successfully differentiated into islet-like cells. The CPSC-derived islet-like cells expressed islet cell development-related genes, such as insulin, glucagon, pax-4, Nkx6.1, pax-6, and Fox. Moreover, CPSC-derived islet-like cells can be stained with zinc ions, which are widely distributed in the islet cells and enable insulin secretion. Altogether, islet-like cells have the potential to be differentiated from CPSCs without gene manipulation, and can be used in diabetic animal models in the future for preclinical and drug testing trial investigations. Copyright © 2017. Published by Elsevier B.V.

Mechanisms of endothelial dysfunction in resistance arteries from patients with end-stage renal disease.

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Leanid Luksha

Full Text Available The study focuses on the mechanisms of endothelial dysfunction in the uremic milieu. Subcutaneous resistance arteries from 35 end-stage renal disease (ESRD patients and 28 matched controls were studied ex-vivo. Basal and receptor-dependent effects of endothelium-derived factors, expression of endothelial NO synthase (eNOS, prerequisites for myoendothelial gap junctions (MEGJ, and associations between endothelium-dependent responses and plasma levels of endothelial dysfunction markers were assessed. The contribution of endothelium-derived hyperpolarizing factor (EDHF to endothelium-dependent relaxation was impaired in uremic arteries after stimulation with bradykinin, but not acetylcholine, reflecting the agonist-specific differences. Diminished vasodilator influences of the endothelium on basal tone and enhanced plasma levels of asymmetrical dimethyl L-arginine (ADMA suggest impairment in NO-mediated regulation of uremic arteries. eNOS expression and contribution of MEGJs to EDHF type responses were unaltered. Plasma levels of ADMA were negatively associated with endothelium-dependent responses in uremic arteries. Preserved responses of smooth muscle to pinacidil and NO-donor indicate alterations within the endothelium and tolerance of vasodilator mechanisms to the uremic retention products at the level of smooth muscle. We conclude that both EDHF and NO pathways that control resistance artery tone are impaired in the uremic milieu. For the first time, we validate the alterations in EDHF type responses linked to kinin receptors in ESRD patients. The association between plasma ADMA concentrations and endothelial function in uremic resistance vasculature may have diagnostic and future therapeutic implications.

Increased expression of SNARE proteins and synaptotagmin IV in islets from pregnant rats and in vitro prolactin-treated neonatal islets

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DANIEL A CUNHA

2006-01-01

Full Text Available During pregnancy and the perinatal period of life, prolactin (PRL and other lactogenic substances induce adaptation and maturation of the stimulus-secretion coupling system in pancreatic β-cells. Since the SNARE molecules, SNAP-25, syntaxin 1, VAMP-2, and synaptotagmins participate in insulin secretion, we investigated whether the improved secretory response to glucose during these periods involves alteration in the expression of these proteins. mRNA was extracted from neonatal rat islets cultured for 5 days in the presence of PRL and from pregnant rats (17th-18th days of pregnancy and reverse transcribed. The expression of genes was analyzed by semi-quantitative RT-PCR assay. The expression of proteins was analyzed by Western blotting and confocal microscopy. Transcription and expression of all SNARE genes and proteins were increased in islets from pregnant and PRL-treated neonatal rats when compared with controls. The only exception was VAMP-2 production in islets from pregnant rats. Increased mRNA and protein expression of synaptotagmin IV, but not the isoform I, also was observed in islets from pregnant and PRL-treated rats. This effect was not inhibited by wortmannin or PD098059, inhibitors of the PI3-kinase and MAPK pathways, respectively. As revealed by confocal laser microscopy, both syntaxin 1A and synaptotagmin IV were immunolocated in islet cells, including the insulin-containing cells. These results indicate that PRL modulates the final steps of insulin secretion by increasing the expression of proteins involved in membrane fusion.

Successful pregnancy and delivery after simultaneous islet-kidney transplantation.

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Assalino, Michela; Podetta, Michele; Demuylder-Mischler, Sandrine; Francini, Katyuska; Pernin, Nadine; Randin, Jean-Pierre; Bosco, Domenico; Andres, Axel; Berney, Thierry

2018-04-19

Allogeneic islet of Langerhans transplantation is a recognized beta-cell replacement therapy for patients affected by type 1 diabetes mellitus. Type 1 diabetes mellitus is a condition associated with an increased risk of adverse outcomes for pregnant women and fetuses. We report the case of a 29-year-old woman with type 1 diabetes mellitus, who underwent successful allogeneic islet transplantation with simultaneous kidney transplantation. She achieved durable insulin independence after 2 islet infusions. Pregnancy was desired and planned 2 years after the last islet infusion. Multidisciplinary monitoring of pregnancy was carried out and the immunosuppressive regimen was adapted. Euglycemia was maintained throughout pregnancy without the need for exogenous insulin. After an uneventful pregnancy, she delivered on term an otherwise healthy male child with imperforate anus that was immediately surgically corrected. In conclusion, allogeneic islet transplantation is a suitable treatment for women of childbearing age with complicated type 1 diabetes mellitus, allowing physiologic glycemic control during pregnancy with a low risk of graft loss. This target can be achieved only by a tight multidisciplinary follow-up, including immunosuppressive therapy adaptation and adequate diabetes and obstetrical monitoring. © 2018 The American Society of Transplantation and the American Society of Transplant Surgeons.

First update of the International Xenotransplantation Association consensus statement on conditions for undertaking clinical trials of porcine islet products in type 1 diabetes--Chapter 3: Porcine islet product manufacturing and release testing criteria.

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Rayat, Gina R; Gazda, Lawrence S; Hawthorne, Wayne J; Hering, Bernhard J; Hosking, Peter; Matsumoto, Shinichi; Rajotte, Ray V

2016-01-01

In the 2009 IXA consensus, the requirements for the quality and control of manufacturing of porcine islet products were based on the U.S. regulatory framework where the porcine islet products fall within the definition of somatic cell therapy under the statutory authority of the U.S. Food and Drug Administration (FDA). In addition, porcine islet products require pre-market approval as a biologic product under the Public Health Services Act and they meet the definition of a drug under the Federal Food, Drug, and Cosmetic Act (FD&C Act). Thus, they are subject to applicable provisions of the law and as such, control of manufacturing as well as reproducibility and consistency of porcine islet products, safety of porcine islet products, and characterization of porcine islet products must be met before proceeding to clinical trials. In terms of control of manufacturing as well as reproducibility and consistency of porcine islet products, the manufacturing facility must be in compliance with current Good Manufacturing Practices (cGMP) guidelines appropriate for the initiation of Phase 1/2 clinical trials. Sponsors intending to conduct a Phase 1/2 trial of islet xenotransplantation products must be able to demonstrate the safety of the product through the establishment of particular quality assurance and quality control procedures. All materials (including animal source and pancreas) used in the manufacturing process of the porcine islet products must be free of adventitious agents. The final porcine islet product must undergo tests for the presence of these adventitious agents including sterility, mycoplasma (if they are cultured), and endotoxin. Assessments of the final product must include the safety specifications mentioned above even if the results are not available until after release as these data would be useful for patient diagnosis and treatment if necessary. In addition, a plan of action must be in place for patient notification and treatment in case the

Efficacy of DHMEQ, a NF-κB inhibitor, in islet transplantation: II. Induction DHMEQ treatment ameliorates subsequent alloimmune responses and permits long-term islet allograft acceptance.

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Watanabe, Masaaki; Yamashita, Kenichiro; Kamachi, Hirofumi; Kuraya, Daisuke; Koshizuka, Yasuyuki; Shibasaki, Susumu; Asahi, Yoh; Ono, Hitoshi; Emoto, Shin; Ogura, Masaomi; Yoshida, Tadashi; Ozaki, Michitaka; Umezawa, Kazuo; Matsushita, Michiaki; Todo, Satoru

2013-09-15

Long-term graft deterioration remains a major obstacle in the success of pancreatic islet transplantation (PITx). Antigen-independent inflammatory and innate immune responses strengthen subsequent antigen-dependent immunity; further, activation of nuclear factor (NF)-κB plays a key role during these responses. In this study, we tested our hypothesis that, by the inhibition of NF-κB activation, the suppression of these early responses after PITx could facilitate graft acceptance. Full major histocompatibility complex (MHC)-mismatched BALB/c (H-2) mice islets were transplanted into streptozotocin-induced diabetic C57BL/6 (B6: H-2) mice. The NF-κB inhibitor dehydroxymethylepoxyquinomicin (DHMEQ) was administered for either 3 or 14 days after PITx. To some PITx recipients, tacrolimus was also administered. Islet allograft survival, alloimmune responses, and in vitro effects of DHMEQ on dendritic cells (DCs) were assessed. With a vehicle treatment, 600 islet allografts were promptly rejected after PITx. In contrast, 3-day treatment with DHMEQ, followed by 2-week treatment with tacrolimus, allowed permanent acceptance of islet allografts. The endogenous danger-signaling molecule high mobility group complex 1 (HMGB1) was elevated in sera shortly after PITx, whereas DHMEQ administration abolished this elevation. DHMEQ suppressed HMGB1-driven cellular activation and proinflammatory cytokine secretion in mouse bone marrow-derived DCs and significantly reduced the capacity of DCs to prime allogeneic T-cell proliferation in vitro. Finally, the DHMEQ plus tacrolimus regimen reverted the diabetic state with only 300 islet allografts. Inhibition of NF-κB activation by DHMEQ shortly after PITx suppresses HMGB1, which activates DCs and strengthens the magnitude of alloimmune responses; this permits long-term islet allograft acceptance, even in case of fewer islet allografts.

Alginate Microencapsulation of Human Islets Does Not Increase Susceptibility to Acute Hypoxia

Science.gov (United States)

Hals, I. K.; Rokstad, A. M.; Strand, B. L.; Oberholzer, J.; Grill, V.

2013-01-01

Islet transplantation in diabetes is hampered by the need of life-long immunosuppression. Encapsulation provides partial immunoprotection but could possibly limit oxygen supply, a factor that may enhance hypoxia-induced beta cell death in the early posttransplantation period. Here we tested susceptibility of alginate microencapsulated human islets to experimental hypoxia (0.1–0.3% O2 for 8 h, followed by reoxygenation) on viability and functional parameters. Hypoxia reduced viability as measured by MTT by 33.8 ± 3.5% in encapsulated and 42.9 ± 5.2% in nonencapsulated islets (P microencapsulation of human islets does not increase susceptibility to acute hypoxia. This is a positive finding in relation to potential use of encapsulation for islet transplantation. PMID:24364039

Transcriptional Regulation of Chemokine Genes: A Link to Pancreatic Islet Inflammation?

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Susan J. Burke

2015-05-01

Full Text Available Enhanced expression of chemotactic cytokines (aka chemokines within pancreatic islets likely contributes to islet inflammation by regulating the recruitment and activation of various leukocyte populations, including macrophages, neutrophils, and T-lymphocytes. Because of the powerful actions of these chemokines, precise transcriptional control is required. In this review, we highlight what is known about the signals and mechanisms that govern the transcription of genes encoding specific chemokine proteins in pancreatic islet β-cells, which include contributions from the NF-κB and STAT1 pathways. We further discuss increased chemokine expression in pancreatic islets during autoimmune-mediated and obesity-related development of diabetes.

Approaches for imaging islets: recent advances and future prospects.

NARCIS (Netherlands)

Ahlgren, U.; Gotthardt, M.

2010-01-01

The establishment of improved technologies for imaging of the pancreas is a key element in addressing several aspects of diabetes pathogenesis. In this respect, the development of a protocol that allows for non-invasive scoring of human islets, or islet beta-cells, is of particular importance. The

Kidney Versus Islet Allograft Survival After Induction of Mixed Chimerism With Combined Donor Bone Marrow Transplantation.

Science.gov (United States)

Oura, Tetsu; Ko, Dicken S C; Boskovic, Svjetlan; O'Neil, John J; Chipashvili, Vaja; Koulmanda, Maria; Hotta, Kiyohiko; Kawai, Kento; Nadazdin, Ognjenka; Smith, R Neal; Cosimi, A B; Kawai, Tatsuo

2016-01-01

We have previously reported successful induction of transient mixed chimerism and long-term acceptance of renal allografts in MHC mismatched nonhuman primates. In this study, we attempted to extend this tolerance induction approach to islet allografts. A total of eight recipients underwent MHC mismatched combined islet and bone marrow (BM) transplantation after induction of diabetes by streptozotocin. Three recipients were treated after a nonmyeloablative conditioning regimen that included low-dose total body and thymic irradiation, horse Atgam (ATG), six doses of anti-CD154 monoclonal antibody (mAb), and a 1-month course of cyclosporine (CyA) (Islet A). In Islet B, anti-CD8 mAb was administered in place of CyA. In Islet C, two recipients were treated with Islet B, but without ATG. The results were compared with previously reported results of eight cynomolgus monkeys that received combined kidney and BM transplantation (Kidney A) following the same conditioning regimen used in Islet A. The majority of kidney/BM recipients achieved long-term renal allograft survival after induction of transient chimerism. However, prolonged islet survival was not achieved in similarly conditioned islet/BM recipients (Islet A), despite induction of comparable levels of chimerism. In order to rule out islet allograft loss due to CyA toxicity, three recipients were treated with anti-CD8 mAb in place of CyA. Although these recipients developed significantly superior mixed chimerism and more prolonged islet allograft survival (61, 103, and 113 days), islet function was lost soon after the disappearance of chimerism. In Islet C recipients, neither prolonged chimerism nor islet survival was observed (30 and 40 days). Significant improvement of mixed chimerism induction and islet allograft survival were achieved with a CyA-free regimen that included anti-CD8 mAb. However, unlike the kidney allograft, islet allograft tolerance was not induced with transient chimerism. Induction of more

Treatment of diabetic rats with encapsulated islets

OpenAIRE

Sweet, Ian R; Yanay, Ofer; Waldron, Lanaya; Gilbert, Merle; Fuller, Jessica M; Tupling, Terry; Lernmark, Ake; Osborne, William R A

2008-01-01

Immunoprotection of islets using bioisolator systems permits introduction of allogeneic cells to diabetic patients without the need for immunosuppression. Using TheraCyte? immunoisolation devices, we investigated two rat models of type 1 diabetes mellitus (T1DM), BB rats and rats made diabetic by streptozotocin (STZ) treatment. We chose to implant islets after the onset of diabetes to mimic the probable treatment of children with T1DM as they are usually diagnosed after disease onset. We enca...

A bilaminated decellularized scaffold for islet transplantation: Structure, properties and functions in diabetic mice.

Science.gov (United States)

Wang, Xi; Wang, Kai; Zhang, Wei; Qiang, Ming; Luo, Ying

2017-09-01

Ectopic transplantation of islets provides a beta cell-replacement approach that may allow the recovery of physiological regulation of the blood sugar level in patients with Type I diabetes (T1D). In development of new extrahepatic islet transplantation protocols in support of the islet engraftment, it is pivotal to develop scaffold materials with multifaceted functions to provide beneficial microenvironment, mediate host response in favor of vascularization/islet integration and maintain long-term islet function at the transplantation site. In this study, a new composite bilaminar decellularized scaffold (CDS) was fabricated with differential structural, degradation and mechanical properties by the combination of a fast-degrading porous collagen matrix and a mechanically supportive porcine pericardium. When investigated in the epididymal fat pad in syngeneic mouse models, it was shown that CDS could serve as superior scaffolds to promote islet adhesion and viability, and islet-CDS constructs also allowed rapid reversal of the hyperglycemic condition in the host. The engraftment and effects of islets were achieved at low islet numbers, accompanied by minimal adverse tissue reactions and optimal islet integration with the surrounding fat tissue. The bioactive surface, mechanical/chemical durability and biocompatibility of the CDS may all have played important roles in facilitating the engraftment of islets. Our study provided new insights into scaffold's function in the interplay of cells, materials and host tissue and the extracellular matrix-based scaffolds have potential for clinical translation in the beta cell-replacement therapy to treat T1D. Copyright © 2017 Elsevier Ltd. All rights reserved.

Methylated trivalent arsenicals are potent inhibitors of glucose stimulated insulin secretion by murine pancreatic islets

International Nuclear Information System (INIS)

Douillet, Christelle; Currier, Jenna; Saunders, Jesse; Bodnar, Wanda M.; Matoušek, Tomáš; Stýblo, Miroslav

2013-01-01

Epidemiologic evidence has linked chronic exposure to inorganic arsenic (iAs) with an increased prevalence of diabetes mellitus. Laboratory studies have identified several mechanisms by which iAs can impair glucose homeostasis. We have previously shown that micromolar concentrations of arsenite (iAs III ) or its methylated trivalent metabolites, methylarsonite (MAs III ) and dimethylarsinite (DMAs III ), inhibit the insulin-activated signal transduction pathway, resulting in insulin resistance in adipocytes. Our present study examined effects of the trivalent arsenicals on insulin secretion by intact pancreatic islets isolated from C57BL/6 mice. We found that 48-hour exposures to low subtoxic concentrations of iAs III , MAs III or DMAs III inhibited glucose-stimulated insulin secretion (GSIS), but not basal insulin secretion. MAs III and DMAs III were more potent than iAs III as GSIS inhibitors with estimated IC 50 ≤ 0.1 μM. The exposures had little or no effects on insulin content of the islets or on insulin expression, suggesting that trivalent arsenicals interfere with mechanisms regulating packaging of the insulin transport vesicles or with translocation of these vesicles to the plasma membrane. Notably, the inhibition of GSIS by iAs III , MAs III or DMAs III could be reversed by a 24-hour incubation of the islets in arsenic-free medium. These results suggest that the insulin producing pancreatic β-cells are among the targets for iAs exposure and that the inhibition of GSIS by low concentrations of the methylated metabolites of iAs may be the key mechanism of iAs-induced diabetes. - Highlights: ► Trivalent arsenicals inhibit glucose stimulated insulin secretion by pancreatic islets. ► MAs III and DMAs III are more potent inhibitors than arsenite with IC 50 ∼ 0.1 μM. ► The arsenicals have little or no effects on insulin expression in pancreatic islets. ► The inhibition of insulin secretion by arsenite, MAs III or DMAs III is reversible. ► Thus

Inhibition of carbachol-induced formation of inositolphosphates in isolated pancreatic islets

DEFF Research Database (Denmark)

Kardasz, A.M.J.; Capito, Kirsten; Hansen, Svend Erik

1991-01-01

Medicinsk biokemi, feed-back inhibition, phospholipase C, pancreatic islets, Calcium, proteinkinase C......Medicinsk biokemi, feed-back inhibition, phospholipase C, pancreatic islets, Calcium, proteinkinase C...

Improving pancreatic islet in vitro functionality and transplantation efficiency by using heparin mimetic peptide nanofiber gels.

Science.gov (United States)

Uzunalli, Gozde; Tumtas, Yasin; Delibasi, Tuncay; Yasa, Oncay; Mercan, Sercan; Guler, Mustafa O; Tekinay, Ayse B

2015-08-01

Pancreatic islet transplantation is a promising treatment for type 1 diabetes. However, viability and functionality of the islets after transplantation are limited due to loss of integrity and destruction of blood vessel networks. Thus, it is important to provide a proper mechanically and biologically supportive environment for enhancing both in vitro islet culture and transplantation efficiency. Here, we demonstrate that heparin mimetic peptide amphiphile (HM-PA) nanofibrous network is a promising platform for these purposes. The islets cultured with peptide nanofiber gel containing growth factors exhibited a similar glucose stimulation index as that of the freshly isolated islets even after 7 days. After transplantation of islets to STZ-induced diabetic rats, 28 day-long monitoring displayed that islets that were transplanted in HM-PA nanofiber gels maintained better blood glucose levels at normal levels compared to the only islet transplantation group. In addition, intraperitoneal glucose tolerance test revealed that animals that were transplanted with islets within peptide gels showed a similar pattern with the healthy control group. Histological assessment showed that islets transplanted within peptide nanofiber gels demonstrated better islet integrity due to increased blood vessel density. This work demonstrates that using the HM-PA nanofiber gel platform enhances the islets function and islet transplantation efficiency both in vitro and in vivo. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Application of Rotating Wall Vessel (RWV) Cell Culture for Pancreas Islet Cell Transplantation

Science.gov (United States)

Rutzky, Lynne P.

1998-01-01

Type I insulin-dependent diabetes mellitus (IDDM) remains a major cause of morbidity and mortality in both pediatric and adult populations, despite significant advances in medical management. While insulin therapy treats symptoms of acute diabetes, it fails to prevent chronic complications such as microvascular disease, blindness, neuropathy, and chronic renal failure. Strict control of blood glucose concentrations delays but does not prevent the onset and progression of secondary complications. Although, whole pancreas transplantation restores physiological blood glucose levels, a continuous process of allograft rejection causes vascular and exocrine-related complications. Recent advances in methods for isolation and purification of pancreatic islets make transplantation of islet allografts an attractive alternative to whole pancreas transplantation. However, immunosuppressive drugs are necessary to prevent rejection of islet allografts and many of these drugs are known to be toxic to the islets. Since auto-transplants of isolated islets following total pancreatectomy survive and function in vivo, it is apparent that a major obstacle to successful clinical islet transplantation is the immunogenicity of the islet allografts.

Animal Models of Diabetes Mellitus for Islet Transplantation

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Naoaki Sakata

2012-01-01

Full Text Available Due to current improvements in techniques for islet isolation and transplantation and protocols for immunosuppressants, islet transplantation has become an effective treatment for severe diabetes patients. Many diabetic animal models have contributed to such improvements. In this paper, we focus on 3 types of models with different mechanisms for inducing diabetes mellitus (DM: models induced by drugs including streptozotocin (STZ, pancreatomized models, and spontaneous models due to autoimmunity. STZ-induced diabetes is one of the most commonly used experimental diabetic models and is employed using many specimens including rodents, pigs or monkeys. The management of STZ models is well established for islet studies. Pancreatomized models reveal different aspects compared to STZ-induced models in terms of loss of function in the increase and decrease of blood glucose and therefore are useful for evaluating the condition in total pancreatomized patients. Spontaneous models are useful for preclinical studies including the assessment of immunosuppressants because such models involve the same mechanisms as type 1 DM in the clinical setting. In conclusion, islet researchers should select suitable diabetic animal models according to the aim of the study.

Role of Natural Killer Cells in the Innate Immune System After Intraportal Islet Transplantation in Mice.

Science.gov (United States)

Saeki, Y; Ishiyama, K; Ishida, N; Tanaka, Y; Ohdan, H

Both liver natural killer (NK) and NK T cells of the innate immune system play a crucial role in islet graft loss after intraportal islet transplantation, although a relationship between NK and NK T cells in islet loss has not been proven. In this study, we investigated the role of NK cells in the innate immune system in islet graft loss after intraportal islet transplantation. To investigate the involvement of liver NK cells in islet destruction, we assessed the differences in graft survival after intraportal islet transplantation between CD1d -/- diabetic mice and NK cell-depleted CD1d -/- diabetic mice. The transplantation of 400 islets into the liver was sufficient to reverse hyperglycemia in wild-type diabetic mice (100%, 4/4). However, normoglycemia could not be achieved when 200 islets were transplanted (0%, 0/4). In contrast, intraportal transplantation of 200 islets in NK cell-depleted CD1d -/- diabetic mice ameliorated hyperglycemia in 71% of cases (5/7), whereas transplantation of the same number of islets in CD1d -/- diabetic mice did not (0%, 0/4). Histologic findings also confirmed that intact islets were observed in NK cell-depleted CD1d -/- diabetic mice, but were difficult to observe in CD1d -/- diabetic mice. The involvement of liver NK cells in the innate immune system related to islet graft loss after intraportal islet transplantation is revealed by improved graft survival and function in NK cell-depleted CD1d -/- diabetic mice. Our data reveal that regulation of NK cell activity is particularly important when insufficient islet numbers are used for transplantation. Copyright © 2016 Elsevier Inc. All rights reserved.

Encapsulated Islet Transplantation: Where Do We Stand?

Science.gov (United States)

Vaithilingam, Vijayaganapathy; Bal, Sumeet; Tuch, Bernard E

2017-01-01

Transplantation of pancreatic islets encapsulated within immuno-protective microcapsules is a strategy that has the potential to overcome graft rejection without the need for toxic immunosuppressive medication. However, despite promising preclinical studies, clinical trials using encapsulated islets have lacked long-term efficacy, and although generally considered clinically safe, have not been encouraging overall. One of the major factors limiting the long-term function of encapsulated islets is the host's immunological reaction to the transplanted graft which is often manifested as pericapsular fibrotic overgrowth (PFO). PFO forms a barrier on the capsule surface that prevents the ingress of oxygen and nutrients leading to islet cell starvation, hypoxia and death. The mechanism of PFO formation is still not elucidated fully and studies using a pig model have tried to understand the host immune response to empty alginate microcapsules. In this review, the varied strategies to overcome or reduce PFO are discussed, including alginate purification, altering microcapsule geometry, modifying alginate chemical composition, co-encapsulation with immunomodulatory cells, administration of pharmacological agents, and alternative transplantation sites. Nanoencapsulation technologies, such as conformal and layer-by-layer coating technologies, as well as nanofiber, thin-film nanoporous devices, and silicone based NanoGland devices are also addressed. Finally, this review outlines recent progress in imaging technologies to track encapsulated cells, as well as promising perspectives concerning the production of insulin-producing cells from stem cells for encapsulation.

Stem Cells as a Tool to Improve Outcomes of Islet Transplantation

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Emily Sims

2012-01-01

Full Text Available The publication of the promising results of the Edmonton protocol in 2000 generated optimism for islet transplantation as a potential cure for Type 1 Diabetes Mellitus. Unfortunately, follow-up data revealed that less than 10% of patients achieved long-term insulin independence. More recent data from other large trials like the Collaborative Islet Transplant Registry show incremental improvement with 44% of islet transplant recipients maintaining insulin independence at three years of follow-up. Multiple underlying issues have been identified that contribute to islet graft failure, and newer research has attempted to address these problems. Stem cells have been utilized not only as a functional replacement for β cells, but also as companion or supportive cells to address a variety of different obstacles that prevent ideal graft viability and function. In this paper, we outline the manners in which stem cells have been applied to address barriers to the achievement of long-term insulin independence following islet transplantation.

On the use of [18F]DOPA as an imaging biomarker for transplanted islet mass

International Nuclear Information System (INIS)

Eriksson, Olof; Mintz, Akiva; Liu, Chengyang; Yu, Ming; Naji, Ali; Alavi, Abass

2014-01-01

Islet transplantation is being developed as a potential cure for patients with type 1 diabetes. There is a need for non-invasive imaging techniques for the quantification of transplanted islets, as current transplantation sites are associated with a substantial loss of islet viability. The dopaminergic metabolic pathway is present in the islets; therefore, we propose Fluorine-18 labeled L-3,4-dihydroxyphenylalanine ([ 18 F]DOPA) as a biomarker for transplanted islet mass. The expression of enzymes involved in the dopaminergic metabolic pathway was investigated in both native and transplanted human islets. The specific uptake of [ 18 F]DOPA in islets and immortalized beta cells was studied in vitro by selective blocking of dopa decarboxylase (DDC). Initial in vivo positron emission tomography (PET) imaging of viable subcutaneous human islets was performed using [ 18 F]DOPA. DDC and vesicular monoamine transporter 2 are co-localized with insulin in the native human pancreas, and the expression is retained after transplantation. Islet uptake of the [ 18 F]DOPA could be modulated by inhibiting DDC, indicating that the uptake followed the normal dopaminergic metabolic pathway. In vivo imaging revealed [ 18 F]DOPA uptake at the site of the functional islet graft. Based on the in vitro and in vivo results presented in this study, we propose to further validate [ 18 F]DOPA-PET as a sensitive imaging modality for imaging extrahepatically transplanted islets. (author)

Protective efficacy of folic acid and vitamin B12 against nicotine-induced toxicity in pancreatic islets of the rat

Directory of Open Access Journals (Sweden)

Bhattacharjee Ankita

2015-06-01

Full Text Available Although cigarette smoking is associated with insulin resistance and an increased risk for type 2 diabetes, few studies have examined the effect of nicotine on the adult endocrine pancreas. In this study, male Wister rats were treated with nicotine (3 mg/kg body weight/day with or without supplementation of folic acid (36 μg/kg body weight/day or vitamin B12 (0.63 μg/kg body weight/day alone or in combination. Fasting blood glucose, insulin and HBA1C level and different oxidative and anti-oxidative stress parameters were measured and pancreatic tissue sections were stained with eosin-haematoxylene. Data were analysed by nonparametric statistics. The results revealed that nicotine induced prediabetes condition with subsequent damage to pancreatic islets in rats. Nicotine also caused oxidative stress in pancreatic tissue as evidenced by increased nitric oxide and malondialdehyde level and decreased superoxide dismutase, catalase and reduced glutathione level. Compared to vitamin B12 supplementation, folic acid blunted the nicotine-induced toxicity in pancreatic islets with higher efficacy. Further, folic acid and vitamin B12 in combination were able to confer significant protection on pancreatic islets against nicotine induced toxicity. These results suggest that supplementation of folic acid and vitamin B12 in combination may be a possible strategy of detoxification against nicotine-induced toxicity in pancreatic islets of the rat.

Glucose cycling is markedly enhanced in pancreatic islets of obese hyperglycemic mice

International Nuclear Information System (INIS)

Khan, A.; Chandramouli, V.; Ostenson, C.G.; Berggren, P.O.; Loew, H.L.; Landau, B.R.; Efendic, S.

1990-01-01

Pancreatic islets from fed 7-month old lean and obese hyperglycemic mice (ob/ob) were incubated with 3H2O and 5.5 mM or 16.7 mM glucose. Incorporation of 3H into the medium glucose was taken as the measure of glucose-6-P hydrolysis to glucose. Glucose utilization was measured from the yield of 3H2O from [5-3H]glucose. Only 3-4% of the glucose phosphorylated was dephosphorylated by the lean mouse islets irrespective of the glucose concentration. In contrast, the ob/ob mouse islets at 5.5 mM glucose dephosphorylated 18% of the glucose phosphorylated and 30% at 16.7 mM. Thus, the islets of hyperglycemic mice demonstrate increased glucose cycling as compared to the islets of normoglycemic lean mice

Immunogenicity of Anti-HLA Antibodies in Pancreas and Islet Transplantation.

Science.gov (United States)

Chaigne, Benjamin; Geneugelijk, Kirsten; Bédat, Benoît; Ahmed, Mohamed Alibashe; Hönger, Gideon; De Seigneux, Sophie; Demuylder-Mischler, Sandrine; Berney, Thierry; Spierings, Eric; Ferrari-Lacraz, Sylvie; Villard, Jean

2016-11-01

The aim of the current study was to characterize the anti-HLA antibodies before and after pancreatic islet or pancreas transplantation. We assessed the risk of anti-donor-specific antibody (DSA) sensitization in a single-center, retrospective clinical study at Geneva University Hospital. Data regarding clinical characteristics, graft outcome, HLA mismatch, donor HLA immunogenicity, and anti-HLA antibody characteristics were collected. Between January 2008 and July 2014, 18 patients received islet transplants, and 26 patients received a pancreas transplant. Eleven out of 18 patients (61.1%) in the islet group and 12 out of 26 patients (46.2%) in the pancreas group had anti-HLA antibodies. Six patients (33.3%) developed DSAs against HLA of the islets, and 10 patients (38.4%) developed DSAs against HLA of the pancreas. Most of the DSAs were at a low level. Several parameters such as gender, number of times cells were transplanted, HLA mismatch, eplet mismatch and PIRCHE-II numbers, rejection, and infection were analyzed. Only the number of PIRCHE-II was associated with the development of anti-HLA class II de novo DSAs. Overall, the development of de novo DSAs did not influence graft survival as estimated by insulin independence. Our results indicated that pretransplant DSAs at low levels do not restrict islet or pancreas transplantation [especially islet transplantation (27.8% vs. 15.4.%)]. De novo DSAs do occur at a similar rate in both pancreas and islet transplant recipients (mainly of class II), and the immunogenicity of donor HLA is a parameter that should be taken into consideration. When combined with an immunosuppressive regimen and close follow-up, development of low levels of DSAs was not found to result in reduced graft survival or graft function in the current study.

Liver-derived systemic factors drive β-cell hyperplasia in insulin resistant states

Energy Technology Data Exchange (ETDEWEB)

El Ouaamari, Abdelfattah; Kawamori, Dan; Dirice, Ercument; Liew, Chong Wee; Shadrach, Jennifer L.; Hu, Jiang; Katsuta, Hitoshi; Hollister-Lock, Jennifer; Qian, Weijun; Wagers, Amy J.; Kulkarni, Rohit N.

2013-02-21

Integrative organ cross-talk regulates key aspects of energy homeostasis and its dysregulation may underlie metabolic disorders such as obesity and diabetes. To test the hypothesis that cross-talk between the liver and pancreatic islets modulates β-cell growth in response to insulin resistance, we used the Liver-specific Insulin Receptor Knockout (LIRKO) mouse, a unique model that exhibits dramatic islet hyperplasia. Using complementary in vivo parabiosis and transplantation assays, and in vitro islet culture approaches, we demonstrate that humoral, non-neural, non-cell autonomous factor(s) induce β-cell proliferation in LIRKO mice. Furthermore, we report that a hepatocyte-derived factor(s) stimulates mouse and human β-cell proliferation in ex vivo assays, independent of ambient glucose and insulin levels. These data implicate the liver as a critical source of β-cell growth factors in insulin resistant states.

A simple assay for the detection of antibodies to endocrine islet cell surface antigens

International Nuclear Information System (INIS)

Contreas, G.; Madsen, O.D.; Vissing, H.; Lernmark, Aa.

1986-01-01

A simple and sensitive immunoradiometric assay for the detection of islet cell surface antibodies (CIRMA) has been developed. Live, transformed islet cells derived from a liver metastasis of a transplantable islet cell tumor were grown in removable microtiter wells and incubated with antibody. Cell-bound antibodies were quantitated using 125 I-labelled second antibodies. The assay was used to detect islet cell antibodies present in sera from non-diabetic and diabetic BB rats and proved to be particularly effective for screening hybridoma supernatants in order to identify monoclonal antibodies recognizing islet cell surface antigens. (Auth.)

Occurance of apoptosis during ischemia in porcine pancreas islet cells.

Science.gov (United States)

Stadlbauer, V; Schaffellner, S; Iberer, F; Lackner, C; Liegl, B; Zink, B; Kniepeiss, D; Tscheliessnigg, K H

2003-03-01

Pancreas islet transplantation is a potential treatment of diabetes mellitus and porcine organs provide an easily available source of cells. Unfortunately quality and quantity of isolated islets are still not satisfactory. Apoptosis occurs in freshly isolated islets and plays a significant role in early graft loss. We evaluated the influence of four storage solutions on porcine pancreas islets. After warm ischemia of 15-20 minutes 12 organs were stored in 4 cold preservation solutions: Histidine-Tryptophan-Ketoglutarate solution (HTK), Hank's buffered saline solution (HBSS), University of Wisconsin (UW) solution and Ringer-Lactate (R). After cold ischemia for 100 minutes, organs were fixed in 3% formalin. Apoptotic cells were counted on hematocylin-eosin stainings. Most apoptotic cells were found in organs stored in R. Low numbers were found in the other groups. The difference between organs stored in R and organs stored in UW, HTK, or HBSS was highly significant. No significant difference could be found between UW, HTK and HBSS. Cold and warm ischemia of the pancreas seems to induce apoptosis in islet cells. Preservation solutions cause less apoptosis than electrolyte solution. No significant differences could be found among the preservation solutions.

Origin of induced pancreatic islet tumors: a radioautographic study

International Nuclear Information System (INIS)

Michels, J.E.; Bauer, G.E.; Dixit, P.K.

1987-01-01

Endocrine tumors of the pancreas are induced in a high percentage of young rats by injections of streptozotocin and nicotinamide (SZ/NA). Benign tumors first appear 20 to 36 weeks after drug injections. To determine the possible site of their origin, the incorporation of [ 3 H]thymidine into islets, ducts, acini, microtumors, and gross tumors was examined by radioautography of histologic sections at 1 to 36 weeks after drug injection. Drug treatment led to early (1- to 6-week) increases in nuclear 3 H labeling of exocrine pancreatic structures (ductal and acinar cells), which may involve DNA repair processes. A secondary increase in labeling of duct cells during the period of tumor emergence supports the assumption that SZ/NA-induced tumors are of ductal origin. Microtumors and gross tumors also exhibited markedly elevated rates of [ 3 H]thymidine incorporation compared to control islets. Nontumorous islet tissue, which exhibited a gradual decrease in volume due to B-cell destruction by the drug injection, showed about 10-fold higher 3 H labeling than islets of controls at all time points. The results suggest that in addition to ductal precursors, islets that survive SZ/NA-induced injury may also provide sites of focal endocrine cell differentiation to tumor tissue. Once established, both microtumors and gross tumors continue to grow by accelerated cell division

MULTIHORMONAL ISLET CELL CARCINOMAS IN THREE KOMODO DRAGONS (VARANUS KOMODOENSIS).

Science.gov (United States)

Eustace, Ronan; Garner, Michael M; Cook, Kimberly; Miller, Christine; Kiupel, Matti

2017-03-01

  Multihormonal pancreatic islet cell carcinomas were found in one female and two male captive geriatric Komodo dragons (Varanus komodoensis). Gross changes in the pancreas were visible in two of the cases. Clinical signs noted in the Komodo dragons were lethargy, weakness, and anorexia. Histologically, the tumors were comprised of nests and cords of well-differentiated neoplastic islet cells with scant amounts of eosinophilic cytoplasm and round, euchromatic nuclei, with rare mitoses. Infiltration by the islet cell tumor into the surrounding acinar tissue was observed in all cases, but no metastatic foci were seen. Multihormone expression was observed in all tumors, which labeled strongly positive for glucagon and somatostatin and focally positive for polypeptide. Pancreatic islet cell neoplasms should be considered in the differential diagnosis for geriatric Komodo dragons presenting with weakness, lethargy, and poor appetite.

Current Status of Immunomodulatory and Cellular Therapies in Preclinical and Clinical Islet Transplantation

Directory of Open Access Journals (Sweden)

Preeti Chhabra

2011-01-01

Full Text Available Clinical islet transplantation is a -cell replacement strategy that represents a possible definitive intervention for patients with type 1 diabetes, offering substantial benefits in terms of lowering daily insulin requirements and reducing incidences of debilitating hypoglycemic episodes and unawareness. Despite impressive advances in this field, a limiting supply of islets, inadequate means for preventing islet rejection, and the deleterious diabetogenic and nephrotoxic side effects associated with chronic immunosuppressive therapy preclude its wide-spread applicability. Islet transplantation however allows a window of opportunity for attempting various therapeutic manipulations of islets prior to transplantation aimed at achieving superior transplant outcomes. In this paper, we will focus on the current status of various immunosuppressive and cellular therapies that promote graft function and survival in preclinical and clinical islet transplantation with special emphasis on the tolerance-inducing capacity of regulatory T cells as well as the -cells regenerative capacity of stem cells.

Sympathetic Innervation during Development Is Necessary for Pancreatic Islet Architecture and Functional Maturation

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Philip Borden

2013-07-01

Full Text Available Sympathetic neurons depend on target-derived neurotrophic cues to control their survival and growth. However, whether sympathetic innervation contributes reciprocally to the development of target tissues is less clear. Here, we report that sympathetic innervation is necessary for the formation of the pancreatic islets of Langerhans and for their functional maturation. Genetic or pharmacological ablation of sympathetic innervation during development resulted in altered islet architecture, reduced insulin secretion, and impaired glucose tolerance in mice. Similar defects were observed with pharmacological blockade of β-adrenergic signaling. Conversely, the administration of a β-adrenergic agonist restored islet morphology and glucose tolerance in deinnervated animals. Furthermore, in neuron-islet cocultures, sympathetic neurons promoted islet cell migration in a β-adrenergic-dependent manner. This study reveals that islet architecture requires extrinsic inductive cues from neighboring tissues such as sympathetic nerves and suggests that early perturbations in sympathetic innervation might underlie metabolic disorders.

65Zinc and endogenous zinc content and distribution in islets in relationship to insulin content

International Nuclear Information System (INIS)

Figlewicz, D.P.; Forhan, S.E.; Hodgson, A.T.; Grodsky, G.M.

1984-01-01

Uptake of 65 Zn and distribution of 65 Zn, total zinc, and insulin were measured in rat islets and islet granules under different conditions of islet culture. Specific activity of islet zinc ( 65 Zn/zinc) was less than 15% that of extracellular zinc even after 48 h. In contrast, once in the islet, 65 Zn approached 70% of equilibrium with granular zinc in 24 h and apparent equilibrium by 48 h. During a 24-h culture, at either high or low glucose, reduction of both islet zinc and insulin occurred. However, zinc depletion was greater than that predicted if zinc loss was proportional to insulin depletion and occurred only from the granular compartment, which represents only one third of the total islet zinc. Extension of culture to 48 h caused additional insulin depletion, but islet zinc was unchanged. Omission of calcium during the 48-h culture caused a predicted increase in insulin retention, presumably by inhibiting secretion; however, zinc retention was not increased proportionately. Pretreatment of rats with tolbutamide caused a massive depletion of insulin stored in isolated islets, with little change in total islet zinc; subsequent culture of these islets resulted in a greater loss of granular zinc than predicted from the small loss of granular insulin. None of the conditions tested affected the percentage of either 65 Zn or total zinc that was distributed in the islet granules. Results show that zinc exists in a metabolically labile islet compartment(s) as well as in secretory granules; and extra-granular zinc, although not directly associated with insulin storage, may act as a reservoir for granular zinc and may regulate insulin synthesis, storage, and secretion in ways as yet unknown

Anti-Inflammatory Strategies in Intrahepatic Islet Transplantation: A Comparative Study in Preclinical Models.

Science.gov (United States)

Citro, Antonio; Cantarelli, Elisa; Pellegrini, Silvia; Dugnani, Erica; Piemonti, Lorenzo

2018-02-01

The identification of pathway(s) playing a pivotal role in peritransplant detrimental inflammatory events represents the crucial step toward a better management and outcome of pancreatic islet transplanted patients. Recently, we selected the CXCR1/2 inhibition as a relevant strategy in enhancing pancreatic islet survival after transplantation. Here, the most clinically used anti-inflammatory compounds (IL1-receptor antagonist, steroids, and TNF-α inhibitor) alone or in combination with a CXCR1/2 inhibitor were evaluated in their ability to improve engraftment or delay graft rejection. To rule out bias related to transplantation site, we used well-established preclinical syngeneic (250 C57BL/6 equivalent islets in C57BL/6) and allogeneic (400 Balb/c equivalent islets in C57BL6) intrahepatic islet transplantation platforms. In mice, we confirmed that targeting the CXCR1/2 pathway is crucial in preserving islet function and improving engraftment. In the allogeneic setting, CXCR1/2 inhibitor alone could reduce the overall recruitment of transplant-induced leukocytes and significantly prolong the time to graft rejection both as a single agent and in combination with immunosuppression. No other anti-inflammatory compounds tested (IL1-receptor antagonist, steroids, and TNF-α inhibitor) alone or in combination with CXCR1/2 inhibitor improve islet engraftment and significantly delay graft rejection in the presence of MMF + FK-506 immunosuppressive treatment. These findings indicate that only the CXCR1/2-mediated axis plays a crucial role in controlling the islet damage and should be a target for intervention to improve the efficiency of islet transplantation.

Pig Pancreas Anatomy: Implications for Pancreas Procurement, Preservation, and Islet Isolation

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Ferrer, Joana; Scott, William E; Weegman, Bradley P; Suszynski, Thomas M; Sutherland, David E R; Hering, Bernhard J; Papas, Klearchos K

2009-01-01

Background Islet transplantation is emerging as a treatment option for selected patients with type 1 diabetes. The limited human islet supply from cadavers and poor islet yield and quality remain substantial impediments to progress in the field. Use of porcine islets holds great promise for large-scale application of islet transplantation. Consistent isolation of porcine islets is dependent on advances in pancreas procurement and preservation, and islet isolation requiring detailed knowledge of the porcine pancreatic anatomy. The primary aim of this study was to describe the vascular and ductal anatomy of the porcine pancreas in order to guide and improve organ preservation and enzyme perfusion. Methods Pancreata were removed by en bloc viscerectomy from 65 female Landrace pigs. Results 15% of organs exhibited inconsistent vascular branching from the celiac trunk. All organs had uniform patterns of branching at the superior mesenteric artery. The superior and inferior mesenteric veins (IMV) merged to become the portal vein in all but one case in which the IMV drained into the splenic vein. 97% of pancreata had three lobes: duodenal (DL), connecting (CL), and splenic (SL); 39% demonstrated ductal communication between the CL and the other two lobes; 50% had ductal communication only between the CL and DL; and 11% presented other types of ductal delineation. Conclusions Accounting for the variations in vascular and ductal anatomy, as detailed in this study, will facilitate development of protocols for preservation, optimal enzyme administration, and pancreas distention and digestion, and ultimately lead to substantial improvements in isolation outcomes. PMID:19077881

Harnessing the Foreign Body Reaction in Marginal Mass Device-less Subcutaneous Islet Transplantation in Mice.

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Pepper, Andrew R; Pawlick, Rena; Bruni, Antonio; Gala-Lopez, Boris; Wink, John; Rafiei, Yasmin; Bral, Mariusz; Abualhassan, Nasser; Shapiro, A M James

2016-07-01

Islet transplantation is a successful β-cell replacement therapy for selected patients with type 1 diabetes mellitus. However, despite early insulin independence, long-term graft attrition gradually reverts recipients to exogenous insulin dependency. Undoubtedly, as insulin producing stem cell therapies progress, a transplant site that is retrievable is desirable. This prerequisite is currently incompatible with intrahepatic islet transplantation. Herein, we evaluate the functional capacity of a prevascularized subcutaneous site to accommodate marginal islet mass transplantation in mice. Syngeneic mouse islets (150) were transplanted either under the kidney capsule (KC), into a prevascularized subcutaneous device-less (DL) site, or into the unmodified subcutaneous (SC) tissue. The DL site was created 4 weeks before diabetes induction and islet transplantation through the transient placement of a 5-Fr vascular catheter. Recipient mice were monitored for glycemic control and intraperitoneal glucose tolerance. A marginal islet mass transplanted into the DL site routinely reversed diabetes (n = 13 of 18) whereas all SC islet recipients failed to restore glycemic control (n = 0 of 10, P islet-KC mice (n = 15 of 16) became euglycemic posttransplant. The DL recipients' glucose profiles were comparable to KC islet grafts, postintrapertioneal glucose tolerance testing, whereas SC recipients remained hyperglycemic postglucose challenge. All normoglycemic mice maintained graft function for 100 days until graft retrieval. DL and KC islet grafts stained positively for insulin, microvessels, and a collagen scaffold. The device-less prevascularized approach supports marginal mass islet engraftment in mice.

Enhancing human islet transplantation by localized release of trophic factors from PLG scaffolds.

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Hlavaty, K A; Gibly, R F; Zhang, X; Rives, C B; Graham, J G; Lowe, W L; Luo, X; Shea, L D

2014-07-01

Islet transplantation represents a potential cure for type 1 diabetes, yet the clinical approach of intrahepatic delivery is limited by the microenvironment. Microporous scaffolds enable extrahepatic transplantation, and the microenvironment can be designed to enhance islet engraftment and function. We investigated localized trophic factor delivery in a xenogeneic human islet to mouse model of islet transplantation. Double emulsion microspheres containing exendin-4 (Ex4) or insulin-like growth factor-1 (IGF-1) were incorporated into a layered scaffold design consisting of porous outer layers for islet transplantation and a center layer for sustained factor release. Protein encapsulation and release were dependent on both the polymer concentration and the identity of the protein. Proteins retained bioactivity upon release from scaffolds in vitro. A minimal human islet mass transplanted on Ex4-releasing scaffolds demonstrated significant improvement and prolongation of graft function relative to blank scaffolds carrying no protein, and the release profile significantly impacted the duration over which the graft functioned. Ex4-releasing scaffolds enabled better glycemic control in animals subjected to an intraperitoneal glucose tolerance test. Scaffolds releasing IGF-1 lowered blood glucose levels, yet the reduction was insufficient to achieve euglycemia. Ex4-delivering scaffolds provide an extrahepatic transplantation site for modulating the islet microenvironment to enhance islet function posttransplant. © Copyright 2014 The American Society of Transplantation and the American Society of Transplant Surgeons.

Transplantation of co-aggregates of Sertoli cells and islet cells into liver without immunosuppression.

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Takemoto, Naohiro; Liu, Xibao; Takii, Kento; Teramura, Yuji; Iwata, Hiroo

2014-02-15

Transplantation of islets of Langerhans (islets) was used to treat insulin-dependent diabetes mellitus. However, islet grafts must be maintained by administration of immunosuppressive drugs, which can lead to complications in the long term. An approach that avoids immunosuppressive drug use is desirable. Co-aggregates of Sertoli cells and islet cells from BALB/c mice that were prepared by the hanging drop method were transplanted into C57BL/6 mouse liver through the portal vein as in human clinical islet transplantation. The core part of the aggregates contained mainly Sertoli cells, and these cells were surrounded by islet cells. The co-aggregates retained the functions of both Sertoli and islet cells. When 800 co-aggregates were transplanted into seven C57BL/6 mice via the portal vein, six of seven recipient mice demonstrated quasi-normoglycemia for more than 100 days. The hanging drop method is suitable for preparing aggregates of Sertoli and islet cells for transplantation. Notably, transplantation of these allogeneic co-aggregates into mice with chemically induced diabetes via the portal vein resulted in long-term graft survival without systemic immunosuppression.

Ontogeny of neuro-insular complexes and islets innervation in the human pancreas.

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Alexandra E. Proshchina

2014-04-01

Full Text Available The ontogeny of the neuro-insular complexes (NIC and the islets innervation in human pancreas has not been studied in detail. Our aim was to describe the developmental dynamics and distribution of the nervous system structures in the endocrine part of human pancreas. We used doublestaining with antibodies specific to pan-neural markers (neuron-specific enolase (NSE and S100 protein and to hormones of pancreatic endocrine cells. NSE and S100-positive nerves and ganglia were identified in the human fetal pancreas from gestation week (gw 10 onwards. Later the density of S100 and NSE-positive fibers increased. In adults this network was sparse. The islets innervation started to form from gw 14. NSE-containing endocrine cells were identified from gw 12 onwards. Additionally, S100-positive cells were detected both in the periphery and within some of the islets starting at gw 14. The analysis of islets innervation has shown that the fetal pancreas contained neuro-insular complexes and the number of these complexes was reduced in adults. The highest density of neuro-insular complexes is detected during middle and late fetal periods, when the mosaic islets, typical for adults, form. The close integration between the developing pancreatic islets and the nervous system structures may play an important role not only in the hormone secretion, but also in the islets morphogenesis.

Ontogeny of neuro-insular complexes and islets innervation in the human pancreas.

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Proshchina, Alexandra E; Krivova, Yulia S; Barabanov, Valeriy M; Saveliev, Sergey V

2014-01-01

The ontogeny of the neuro-insular complexes (NIC) and the islets innervation in human pancreas has not been studied in detail. Our aim was to describe the developmental dynamics and distribution of the nervous system structures in the endocrine part of human pancreas. We used double-staining with antibodies specific to pan-neural markers [neuron-specific enolase (NSE) and S100 protein] and to hormones of pancreatic endocrine cells. NSE and S100-positive nerves and ganglia were identified in the human fetal pancreas from gestation week (gw) 10 onward. Later the density of S100 and NSE-positive fibers increased. In adults, this network was sparse. The islets innervation started to form from gw 14. NSE-containing endocrine cells were identified from gw 12 onward. Additionally, S100-positive cells were detected both in the periphery and within some of the islets starting at gw 14. The analysis of islets innervation has shown that the fetal pancreas contained NIC and the number of these complexes was reduced in adults. The highest density of NIC is detected during middle and late fetal periods, when the mosaic islets, typical for adults, form. The close integration between the developing pancreatic islets and the nervous system structures may play an important role not only in the hormone secretion, but also in the islets morphogenesis.

Xenotransplantation of piscine islets into hyperglycemic rats

International Nuclear Information System (INIS)

Weber, C.; Weil, R. III; McIntosh, R.; Hogle, H.; Warden, G.; Reemtsma, K.

1975-01-01

Xenotransplantation of piscine islets into hyperglycemic rats usually lowers the blood sugar level of the recipient. The duration of this effect is prolonged by irradiation of the host or by enclosing donor tissue in synthetic envelopes. This prolongation appears to be related to interference with the host's ability to reject the graft; the duration of the prolongation may be limited by the host tissue reaction surrounding the envelope. The availability of anatomically separate piscine islet tissue makes it potentially useful for xenotransplantation into mammals

Assessment of DNA synthesis in Islet-1{sup +} cells in the adult murine heart

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Weinberger, Florian, E-mail: f.weinberger@uke.de; Mehrkens, Dennis, E-mail: dennis.mehrkens@uk-koeln.de; Starbatty, Jutta, E-mail: starbatty@uke.uni-hamburg.de; Nicol, Philipp, E-mail: Philipp.Nicol@gmx.de; Eschenhagen, Thomas, E-mail: t.eschenhagen@uke.de

2015-01-02

Highlights: • Islet-1 was expressed in the adult heart. • Islet-1-positive cells did not proliferate in the adult heart. • Sinoatrial node cells did not proliferate in the adult heart. - Abstract: Rationale: Islet-1 positive (Islet-1{sup +}) cardiac progenitor cells give rise to the right ventricle, atria and outflow tract during murine cardiac development. In the adult heart Islet-1 expression is limited to parasympathetic neurons, few cardiomyocytes, smooth muscle cells, within the proximal aorta and pulmonary artery and sinoatrial node cells. Its role in these cells is unknown. Here we tested the hypothesis that Islet-1{sup +} cells retain proliferative activity and may therefore play a role in regenerating specialized regions in the heart. Methods and results: DNA synthesis was analyzed by the incorporation of tritiated thymidine ({sup 3}H-thymidine) in Isl-1-nLacZ mice, a transgenic model with an insertion of a nuclear beta-galactosidase in the Islet-1 locus. Mice received daily injections of {sup 3}H-thymidine for 5 days. DNA synthesis was visualized throughout the heart by dipping autoradiography of cryosections. Colocalization of an nLacZ-signal and silver grains would indicate DNA synthesis in Islet-1{sup +} cells. Whereas Islet{sup −} non-myocyte nuclei were regularly marked by accumulation of silver grains, colocalization with nLacZ-signals was not detected in >25,000 cells analyzed. Conclusions: Islet-1{sup +} cells are quiescent in the adult heart, suggesting that, under normal conditions, even pacemaking cells do not proliferate at higher rates than normal cardiac myocytes.

Inactivation of p27kip1 Promoted Nonspecific Inflammation by Enhancing Macrophage Proliferation in Islet Transplantation.

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Li, Yang; Ding, Xiaoming; Fan, Ping; Guo, Jian; Tian, Xiaohui; Feng, Xinshun; Zheng, Jin; Tian, Puxun; Ding, Chenguang; Xue, Wujun

2016-11-01

Islet transplantation suffers from low efficiency caused by nonspecific inflammation-induced graft loss after transplantation. This study reports increased islet loss and enhanced inflammatory response in p27-deficient mice (p27-/-) and proposes a possible mechanism. Compared with wild type, p27-/- mice showed more severe functional injury of islet, with increased serum levels of inflammatory cytokines IL-1 and TNF-α, inducing macrophage proliferation. Furthermore, the increased number, proapoptotic proteins, and nuclear factor-kappa b (NF-κB) phosphorylation status of the infiltrating macrophages were accompanied by increased TNF-α mRNA level of islet graft site in p27-/- mice. Moreover, in vitro, we found that macrophages were still activated and cocultured with islet and promoted islet loss even blocking the direct effect of TNF-α on islets. Malondialdehyde (MDA, an end product of lipid peroxidation) in islet and media were increased after cocultured with macrophages. p27 deficiency also increased macrophage proliferation and islet injury. Therefore, p27 inactivation promotes injury islet graft loss via the elevation of proliferation and inflammatory cytokines secretion in infiltrating macrophages which induced nonspecific inflammation independent of TNF-α/nuclear factor-kappa b pathway. This potentially represents a promising therapeutic target in improving islet graft survival.

Microencapsulated 3-dimensional sensor for the measurement of oxygen in single isolated pancreatic islets.

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Wanyu Chen

Full Text Available Oxygen consumption reflects multiple processes in pancreatic islets including mechanisms contributing to insulin secretion, oxidative stress and viability, providing an important readout in studies of islet function, islet viability and drug testing. Due to the scarcity, heterogeneity, and intrinsic kinetic properties of individual islets, it would be of great benefit to detect oxygen consumption by single islets. We present a novel method we have developed to image oxygen in single islets.Using a microfluidics system, individual islets and a fluorescent oxygen-sensitive dye were encased within a thin alginate polymer layer. Insulin secretion by the encapsulated islets was normal. Fluorescent signal from the encased dye, detected using a standard inverted fluorescence microscope and digital camera, was stable and proportional to the amount of oxygen in the media. When integrated into a perifusion system, the sensing system detected changes in response to metabolic substrates, mitochondrial poisons, and induced-oscillations. Glucose responses averaged 30.1±7.1% of the response to a metabolic inhibitor (cyanide, increases were observed in all cases (n = 6, and the system was able to resolve changes in oxygen consumption that had a period greater than 0.5 minutes. The sensing system operated similarly from 2-48 hours following encapsulation, and viability and function of the islets were not significantly affected by the encapsulation process.An oxygen-dependent dye situated around and within a pancreatic islet encapsulated by a thin layer of alginate was sensitive to changes in oxygen consumption, and was not harmful to the function or viability of islets over the course of two days. The microcapsule-based sensing method is particularly suited to assessing the effects of compounds (dose responses and time courses and chronic changes occurring over the course of days. The approach should be applicable to other cell types and dyes sensitive to other

Electrofusion of mesenchymal stem cells and islet cells for diabetes therapy: a rat model.

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Goichi Yanai

Full Text Available Islet transplantation is a minimally invasive treatment for severe diabetes. However, it often requires multiple donors to accomplish insulin-independence and the long-term results are not yet satisfying. Therefore, novel ways to overcome these problems have been explored. Isolated islets are fragile and susceptible to pro-apoptotic factors and poorly proliferative. In contrast, mesenchymal stem cells (MSCs are highly proliferative, anti-apoptotic and pluripotent to differentiate toward various cell types, promote angiogenesis and modulate inflammation, thereby studied as an enhancer of islet function and engraftment. Electrofusion is an efficient method of cell fusion and nuclear reprogramming occurs in hybrid cells between different cell types. Therefore, we hypothesized that electrofusion between MSC and islet cells may yield robust islet cells for diabetes therapy. We establish a method of electrofusion between dispersed islet cells and MSCs in rats. The fusion cells maintained glucose-responsive insulin release for 20 days in vitro. Renal subcapsular transplantation of fusion cells prepared from suboptimal islet mass (1,000 islets that did not correct hyperglycemia even if co-transplanted with MSCs, caused slow but consistent lowering of blood glucose with significant weight gain within the observation period in streptozotocin-induced diabetic rats. In the fusion cells between rat islet cells and mouse MSCs, RT-PCR showed new expression of both rat MSC-related genes and mouse β-cell-related genes, indicating bidirectional reprogramming of both β-cell and MSCs nuclei. Moreover, decreased caspase3 expression and new expression of Ki-67 in the islet cell nuclei suggested alleviated apoptosis and gain of proliferative capability, respectively. These results show that electrofusion between MSCs and islet cells yield special cells with β-cell function and robustness of MSCs and seems feasible for novel therapeutic strategy for diabetes

Pancreatic Islet Cell Transplantation: A new era in transplantation

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Warnock, Garth L.; Rajotte, Ray V.

1992-01-01

Transplantation of insulin-producing tissue offers a physiologic approach to restoration of glycemic control. Whereas transplantation of vascularized pancreatic grafts has recently achieved encouraging results, pancreatic islet cell transplantation holds the promise of low morbidity and reduced requirements for agressive immunosuppression for recipients. Islet cell transplantation was recently demonstrated to induce euglycemia with insulin independence.

Current Status of Immunomodulatory and Cellular Therapies in Preclinical and Clinical Islet Transplantation

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Chhabra, Preeti; Brayman, Kenneth L.

2011-01-01

Clinical islet transplantation is a β-cell replacement strategy that represents a possible definitive intervention for patients with type 1 diabetes, offering substantial benefits in terms of lowering daily insulin requirements and reducing incidences of debilitating hypoglycemic episodes and unawareness. Despite impressive advances in this field, a limiting supply of islets, inadequate means for preventing islet rejection, and the deleterious diabetogenic and nephrotoxic side effects associated with chronic immunosuppressive therapy preclude its wide-spread applicability. Islet transplantation however allows a window of opportunity for attempting various therapeutic manipulations of islets prior to transplantation aimed at achieving superior transplant outcomes. In this paper, we will focus on the current status of various immunosuppressive and cellular therapies that promote graft function and survival in preclinical and clinical islet transplantation with special emphasis on the tolerance-inducing capacity of regulatory T cells as well as the β-cells regenerative capacity of stem cells. PMID:22046502

Islet cytotoxicity of interleukin 1. Influence of culture conditions and islet donor characteristics

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Mandrup-Poulsen, T; Spinas, G A; Prowse, S J

1987-01-01

We recently demonstrated that the macrophage product interleukin 1 (IL-1) is cytotoxic to isolated pancreatic islets and hypothesized that IL-1 is responsible for beta-cell destruction in insulin-dependent diabetes mellitus (IDDM). We studied whether the variation in IDDM preponderance with age, ...

Magnetic resonance imaging of mouse islet grafts labeled with novel chitosan-coated superparamagnetic iron oxide nanoparticles.

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Jyuhn-Huarng Juang

Full Text Available To better understand the fate of islet isografts and allografts, we utilized a magnetic resonance (MR imaging technique to monitor mouse islets labeled with a novel MR contrast agent, chitosan-coated superparamagnetic iron oxide (CSPIO nanoparticles.After being incubated with and without CSPIO (10 µg/ml, C57BL/6 mouse islets were examined under transmission electron microscope (TEM and their insulin secretion was measured. Cytotoxicity was examined in α (αTC1 and β (NIT-1 and βTC cell lines as well as islets. C57BL/6 mice were used as donors and inbred C57BL/6 and Balb/c mice were used as recipients of islet transplantation. Three hundred islets were transplanted under the left kidney capsule of each mouse and then MR was performed in the recipients periodically. At the end of study, the islet graft was removed for histology and TEM studies.After incubation of mouse islets with CSPIO (10 µg/mL, TEM showed CSPIO in endocytotic vesicles of α- and β-cells at 8 h. Incubation with CSPIO did not affect insulin secretion from islets and death rates of αTC1, NIT-1 and βTC cell lines as well as islets. After syngeneic and allogeneic transplantation, grafts of CSPIO-labeled islets were visualized on MR scans as persistent hypointense areas. At 8 weeks after syngeneic transplantation and 31 days after allogeneic transplantation, histology of CSPIO-labeled islet grafts showed colocalized insulin and iron staining in the same areas but the size of allografts decreased with time. TEM with elementary iron mapping demonstrated CSPIO distributed in the cytoplasm of islet cells, which maintained intact ultrastructure.Our results indicate that after syngeneic and allogeneic transplantation, islets labeled with CSPIO nanoparticles can be effectively and safely imaged by MR.

Evaluation of MicroRNA375 as a Novel Biomarker for Graft Damage in Clinical Islet Transplantation.

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Kanak, Mazhar A; Takita, Morihito; Shahbazov, Rauf; Lawrence, Michael C; Chung, Wen Yuan; Dennison, Ashley R; Levy, Marlon F; Naziruddin, Bashoo

2015-08-01

Early and sensitive detection of islet graft damage is essential for improving posttransplant outcomes. MicroRNA 375 (miR375) has been reported as a biomarker of pancreatic β-cell death in small animal models. The miR375 levels were measured in purified human islets, sera from patients with autologous and allogeneic islet transplantation as well as total pancreatectomy alone (nontransplanted group). The miR375 levels were also determined in a miniaturized in vitro tube model comprising human islets and autologous blood. The miR375 expression level in islets was dose-dependent (P islet damage in plasma in the in vitro model (P = 0.003). Clinical analysis revealed that circulating miR375 levels in both autologous and allogeneic islet recipients were significantly elevated for 7 days after islet infusion, compared with the nontransplanted group (P = 0.005 and islet graft damage among 3 different anti-inflammatory protocols for clinical autologous transplantation (P islet transplantation because serum C-peptide and proinsulin levels are difficult to interpret due to the influence of multiple factors, such as β-cell stress and physiological response.

Effects of spinach nitrate on insulin resistance, endothelial dysfunction markers and inflammation in mice with high-fat and high-fructose consumption

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Ting Li

2016-09-01

Full Text Available Background: Insulin resistance, which is associated with an increased risk of cardiovascular morbidity and mortality, has become a leading nutrition problem. Inorganic nitrate enriched in spinach has been demonstrated to reverse the pathological features of insulin resistance and endothelial dysfunction. However, the effects of a direct intake of nitrate-enriched spinach on insulin resistance and endothelial dysfunction have not been studied. Objective: To investigate the effects of spinach nitrate on insulin resistance, lipid metabolism, endothelial function, and inflammation in mice fed with a high-fat and high-fructose diet. Design: A diet intervention of spinach with or without nitrate was performed in mice. A high-fat and high-fructose diet was used to cause insulin resistance, endothelial dysfunction, and inflammation in mice. The impacts of spinach nitrate on lipid profile, insulin resistance, markers of endothelial function, and inflammation were determined in mice. Results: Spinach nitrate improved the vascular endothelial function of the mice with high-fat and high-fructose consumption, as evidenced by the elevated plasma nitrite level, increased serum nitric oxide (NO level and decreased serum ET-1 level after spinach nitrate intervention. Spinach nitrate also reduced serum triglycerides, total cholesterol, and low-density lipoprotein-cholesterol levels and elevated serum high-density lipoprotein-cholesterol levels in the mice fed with a high-fat and high-fructose diet. Mice receiving spinach with 60 mg/kg of nitrate (1.02±0.34 showed a significantly low homeostasis model assessment-insulin resistance index as compared with the model mice (2.05±0.58, which is indicating that spinach nitrate could effectively improve the insulin resistance. In addition, spinach nitrate remarkably decreased the elevated serum C-reactive protein, tumor necrosis factor α, and interleukin-6 levels induced by a high-fat and high-fructose diet

Ca(2+) mishandling and cardiac dysfunction in obesity and insulin resistance: role of oxidative stress.

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Carvajal, Karla; Balderas-Villalobos, Jaime; Bello-Sanchez, Ma Dolores; Phillips-Farfán, Bryan; Molina-Muñoz, Tzindilu; Aldana-Quintero, Hugo; Gómez-Viquez, Norma L

2014-11-01

Obesity and insulin resistance (IR) are strongly connected to the development of subclinical cardiac dysfunction and eventually can lead to heart failure, which is the main cause of morbidity and death in patients having these metabolic diseases. It has been considered that excessive fat tissue may play a critical role in producing systemic IR and enhancing reactive oxygen species (ROS) generation. This oxidative stress (OS) may elicit or exacerbate IR. On the other hand, evidence suggests that some of the cellular mechanisms involved in the pathophysiology of obesity and IR-related cardiomyopathy are excessive myocardial ROS production and abnormal Ca(2+) homeostasis. In addition, emerging evidence suggests that augmented ROS production may contribute to Ca(2+) mishandling by affecting the redox state of key proteins implicated in this process. In this review, we focus on the role of Ca(2+) mishandling in the development of cardiac dysfunction in obesity and IR and address the evidence suggesting that OS might also contribute to cardiac dysfunction by affecting Ca(2+) handling. Copyright © 2014 Elsevier Ltd. All rights reserved.

Microencapsulation of Pancreatic Islets for Use in a Bioartificial Pancreas

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Opara, Emmanuel C.; McQuilling, John P.; Farney, Alan C.

2013-01-01

Islet transplantation is the most exciting treatment option for individuals afflicted with Type 1 diabetes. However, the severe shortage of human pancreas and the need to use risky immunosuppressive drugs to prevent transplant rejection remain two major obstacles for the routine use of islet transplantation in diabetic patients. Successful development of a bioartificial pancreas using the approach of microencapsulation with perm-selective coating of islets with biopolymers for graft immunoisolation holds tremendous promise for diabetic patients because it has great potential to overcome these two barriers. In this chapter, we provide a detailed description of the microencapsulation process. PMID:23494435

Phase transitions in pancreatic islet cellular networks and implications for type-1 diabetes

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Stamper, I. J.; Jackson, Elais; Wang, Xujing

2014-01-01

In many aspects the onset of a chronic disease resembles a phase transition in a complex dynamic system: Quantitative changes accumulate largely unnoticed until a critical threshold is reached, which causes abrupt qualitative changes of the system. In this study we examine a special case, the onset of type-1 diabetes (T1D), a disease that results from loss of the insulin-producing pancreatic islet β cells. Within each islet, the β cells are electrically coupled to each other via gap-junctional channels. This intercellular coupling enables the β cells to synchronize their insulin release, thereby generating the multiscale temporal rhythms in blood insulin that are critical to maintaining blood glucose homeostasis. Using percolation theory we show how normal islet function is intrinsically linked to network connectivity. In particular, the critical amount of β-cell death at which the islet cellular network loses site percolation is consistent with laboratory and clinical observations of the threshold loss of β cells that causes islet functional failure. In addition, numerical simulations confirm that the islet cellular network needs to be percolated for β cells to synchronize. Furthermore, the interplay between site percolation and bond strength predicts the existence of a transient phase of islet functional recovery after onset of T1D and introduction of treatment, potentially explaining the honeymoon phenomenon. Based on these results, we hypothesize that the onset of T1D may be the result of a phase transition of the islet β-cell network.

Design of a vascularized synthetic poly(ethylene glycol) macroencapsulation device for islet transplantation.

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Weaver, Jessica D; Headen, Devon M; Hunckler, Michael D; Coronel, Maria M; Stabler, Cherie L; García, Andrés J

2018-07-01

The use of immunoisolating macrodevices in islet transplantation confers the benefit of safety and translatability by containing transplanted cells within a single retrievable device. To date, there has been limited development and characterization of synthetic poly(ethylene glycol) (PEG)-based hydrogel macrodevices for islet encapsulation and transplantation. Herein, we describe a two-component synthetic PEG hydrogel macrodevice system, designed for islet delivery to an extrahepatic islet transplant site, consisting of a hydrogel core cross-linked with a non-degradable PEG dithiol and a vasculogenic outer layer cross-linked with a proteolytically sensitive peptide to promote degradation and enhance localized vascularization. Synthetic PEG macrodevices exhibited equivalent passive molecular transport to traditional microencapsulation materials (e.g., alginate) and long-term stability in the presence of proteases in vitro and in vivo, out to 14 weeks in rats. Encapsulated islets demonstrated high viability within the device in vitro and the incorporation of RGD adhesive peptides within the islet encapsulating PEG hydrogel improved insulin responsiveness to a glucose challenge. In vivo, the implementation of a vasculogenic, degradable hydrogel layer at the outer interface of the macrodevice enhanced vascular density within the rat omentum transplant site, resulting in improved encapsulated islet viability in a syngeneic diabetic rat model. These results highlight the benefits of the facile PEG platform to provide controlled presentation of islet-supportive ligands, as well as degradable interfaces for the promotion of engraftment and overall graft efficacy. Copyright © 2018 Elsevier Ltd. All rights reserved.

Clock-controlled output gene Dbp is a regulator of Arnt/Hif-1β gene expression in pancreatic islet β-cells

International Nuclear Information System (INIS)

Nakabayashi, Hiroko; Ohta, Yasuharu; Yamamoto, Masayoshi; Susuki, Yosuke; Taguchi, Akihiko; Tanabe, Katsuya; Kondo, Manabu; Hatanaka, Masayuki; Nagao, Yuko; Tanizawa, Yukio

2013-01-01

Highlights: •Arnt mRNA expressed in a circadian manner in mouse pancreatic islets. •Expressions of Dbp and Arnt damped in the islets of a diabetic model mouse. •DBP and E4BP4 regulate Arnt promoter activity by direct binding. •Arnt may have a role in connecting circadian rhythm and metabolism. -- Abstract: Aryl hydrocarbon receptor nuclear translocator (ARNT)/hypoxia inducible factor-1β (HIF-1β) has emerged as a potential determinant of pancreatic β-cell dysfunction and type 2 diabetes in humans. An 82% reduction in Arnt expression was observed in islets from type 2 diabetic donors as compared to non-diabetic donors. However, few regulators of Arnt expression have been identified. Meanwhile, disruption of the clock components CLOCK and BMAL1 is known to result in hypoinsulinemia and diabetes, but the molecular details remain unclear. In this study, we identified a novel molecular connection between Arnt and two clock-controlled output genes, albumin D-element binding protein (Dbp) and E4 binding protein 4 (E4bp4). By conducting gene expression studies using the islets of Wfs1 −/− A y /a mice that develop severe diabetes due to β-cell apoptosis, we demonstrated clock-related gene expressions to be altered in the diabetic mice. Dbp mRNA decreased by 50%, E4bp4 mRNA increased by 50%, and Arnt mRNA decreased by 30% at Zeitgever Time (ZT) 12. Mouse pancreatic islets exhibited oscillations of clock gene expressions. E4BP4, a D-box negative regulator, oscillated anti-phase to DBP, a D-box positive regulator. We also found low-amplitude circadian expression of Arnt mRNA, which peaked at ZT4. Over-expression of DBP raised both mRNA and protein levels of ARNT in HEK293 and MIN6 cell lines. Arnt promoter-driven luciferase reporter assay in MIN6 cells revealed that DBP increased Arnt promoter activity by 2.5-fold and that E4BP4 competitively inhibited its activation. In addition, on ChIP assay, DBP and E4BP4 directly bound to D-box elements within the Arnt

Clock-controlled output gene Dbp is a regulator of Arnt/Hif-1β gene expression in pancreatic islet β-cells

Energy Technology Data Exchange (ETDEWEB)

Nakabayashi, Hiroko; Ohta, Yasuharu, E-mail: yohta@yamaguchi-u.ac.jp; Yamamoto, Masayoshi; Susuki, Yosuke; Taguchi, Akihiko; Tanabe, Katsuya; Kondo, Manabu; Hatanaka, Masayuki; Nagao, Yuko; Tanizawa, Yukio, E-mail: tanizawa@yamaguchi-u.ac.jp

2013-05-03

Highlights: •Arnt mRNA expressed in a circadian manner in mouse pancreatic islets. •Expressions of Dbp and Arnt damped in the islets of a diabetic model mouse. •DBP and E4BP4 regulate Arnt promoter activity by direct binding. •Arnt may have a role in connecting circadian rhythm and metabolism. -- Abstract: Aryl hydrocarbon receptor nuclear translocator (ARNT)/hypoxia inducible factor-1β (HIF-1β) has emerged as a potential determinant of pancreatic β-cell dysfunction and type 2 diabetes in humans. An 82% reduction in Arnt expression was observed in islets from type 2 diabetic donors as compared to non-diabetic donors. However, few regulators of Arnt expression have been identified. Meanwhile, disruption of the clock components CLOCK and BMAL1 is known to result in hypoinsulinemia and diabetes, but the molecular details remain unclear. In this study, we identified a novel molecular connection between Arnt and two clock-controlled output genes, albumin D-element binding protein (Dbp) and E4 binding protein 4 (E4bp4). By conducting gene expression studies using the islets of Wfs1{sup −/−} A{sup y}/a mice that develop severe diabetes due to β-cell apoptosis, we demonstrated clock-related gene expressions to be altered in the diabetic mice. Dbp mRNA decreased by 50%, E4bp4 mRNA increased by 50%, and Arnt mRNA decreased by 30% at Zeitgever Time (ZT) 12. Mouse pancreatic islets exhibited oscillations of clock gene expressions. E4BP4, a D-box negative regulator, oscillated anti-phase to DBP, a D-box positive regulator. We also found low-amplitude circadian expression of Arnt mRNA, which peaked at ZT4. Over-expression of DBP raised both mRNA and protein levels of ARNT in HEK293 and MIN6 cell lines. Arnt promoter-driven luciferase reporter assay in MIN6 cells revealed that DBP increased Arnt promoter activity by 2.5-fold and that E4BP4 competitively inhibited its activation. In addition, on ChIP assay, DBP and E4BP4 directly bound to D-box elements within the

Morphologic and morphometric evaluation of pancreatic islets in chronic Chagas' disease

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Saldanha João Carlos

2001-01-01

Full Text Available PURPOSE: Hyperglycemia and abnormal glucose tolerance tests observed in some patients with chronic Chagas' disease suggest the possibility of morphological changes in pancreatic islets and/or denervation. The purpose of this study was to describe the morphology and morphometry of pancreatic islets in chronic Chagas' disease. METHODS: Morphologic and computerized morphometric studies were performed in fragments of the head, body, and tail regions of the pancreas obtained at necropsies of 8 normal controls and 17 patients with chronic Chagas' disease: 8 with the digestive form (Megas and 9 with the congestive heart failure form. RESULTS: The Megas group had a larger (p < 0.05 pancreatic islet area in the tail of the pancreas (10649.3 ± 4408.8 µm² than the normal control (9481.8 ± 3242.4 µm² and congestive heart failure (9475.1 ± 2104.9 µm² groups; likewise, the density of the pancreatic islets (PI was greater (1.2 ± 0.7 vs. 0.9 ± 0.6 vs. 1.9 ± 1.0 PI/mm², respectively. In the tail region of the pancreas of patients with the Megas form, there was a significant and positive correlation (r = +0.73 between the area and density of pancreatic islets. Discrete fibrosis and leukocytic infiltrates were found in pancreatic ganglia and pancreatic islets of the patients with Chagas' disease. Trypanosoma cruzi nests were not observed in the examined sections. Individuals with the Megas form of Chagas' disease showed increased area and density of pancreatic islets in the tail of the pancreas. CONCLUSION: The observed morphometric and morphologic alterations are consistent with functional changes in the pancreas, including glycemia and insulin disturbances.

Immunohistochemical and morphometric study of the development of fetal and newborn rat pancreatic islets

International Nuclear Information System (INIS)

Badawoud, Mohammed H.

2003-01-01

Aim of this study is to perform a detailed morphometric immunohistochemichal study of develpment of fetal and newborn rat pancreatic islets. 24 pancreas were obtained from 19 and 21-day-old fetal rats,1 and 4-day-old newborn rats. They were fixed in a buffered neutral formalin ,dehydrated and embedded in paraplast. Sections were stained with anti-insulin antibodies. Study was performed at Department of Anatomy, King Abdul-Aziz University, Jeddah,Kingdom of Saudi Arabia, between 2001 and 2002. The volume density of B cells showed a grdual increase during the last days of gestation and a slight increase during the first 4 days after birth. All the other morphometric parameters showed a gradual increase during the last days of gestation and during the first days after birth.The B cell nuclear diameter and volume showed a slight increase after birth. B cells were stained and present in the central part of of fetal and new born islets,while the other islet cells were present in the periphery of the islets. The size of endocrine tissue, which was represented by the islet diameter, islet volume, islet volume density, total number of islet cells,number of B cells and volume density of B cells showed a progressive increase during the prenatal period. (author)

3H-cyclosporine internalization and secretion by human fetal pancreatic islets

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Formby, B.; Walker, L.; Peterson, C.M.

1988-01-01

Human fetal pancreatic islets were isolated from 16- to 20-week-old fetuses by a collagenase technique and cultured 48 hr in RPMI 1640 containing 10% human adult serum and unlabeled 0 to 5 micrograms cyclosporine A (CsA)/ml. Insulin secretory capacity of human fetal islets was expressed as a fractional stimulatory ratio FSR = F2/F1 of the fractional secretion rates during two successive 1 hr static incubations first with 2 mM glucose (F1) to stabilize secretion followed by maximal stimulus, i.e., 25 mM glucose plus 10 mM L-leucine and 10 mM L-arginine (F2). Unlabeled CsA at the above concentrations had no significant effects on the insulin secretory capacity expressed by FSR-values. Studies of net uptake of 3H-CsA by islets cultured for varying periods up to 40 hr and expressed as picomole 3H-CsA per picomole islet insulin content demonstrated that uptake rate was slow and did not reach isotopic equilibrium over the 40 hr of culture. When isolated fetal islets were cultured for 48 hr in the presence of 3H-CsA and varying concentrations of unlabeled CsA it was found during two successive 1 hr static incubations that fetal islets secrete insulin concomitantly with 3H-CsA following maximal stimulus for secretion. An optimal secretory molar ratio of 3H-CsA to insulin of 4.0 +/- 1.3 (n = 7) was found after islets were cultured 48 hr in the presence of a saturating 2.128 micrograms 3H-CsA per milliliter culture medium. In three successive 30-min static incubations of 3H-CsA loaded islets, first with low glucose, followed by high glucose plus L-arginine and L-leucine, and finally with high glucose plus L-arginine and L-leucine and 10 mM theophylline, the proportional fractional secretion rates of insulin and 3H-CsA were of the same magnitude

Pretargeting vs. direct targeting of human betalox5 islet cells subcutaneously implanted in mice using an anti-human islet cell antibody

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Liu Guozheng; Dou Shuping; Akalin, Ali; Rusckowski, Mary; Streeter, Philip R.; Shultz, Leonard D.; Greiner, Dale L.

2012-01-01

Introduction: We previously demonstrated MORF/cMORF pretargeting of human islets and betalox 5 cells (a human beta cell line) transplanted subcutaneously in mice with the anti-human islet antibody, HPi1. We now compare pretargeting with direct targeting in the beta cell transplant model to evaluate the degree to which target/non-target (T/NT) ratios may be improved by pretargeting. Methods: Specific binding of an anti-human islet antibody HPi1 to the beta cells transplanted subcutaneously in mice was examined against a negative control antibody. We then compared pretargeting by MORF-HPi1 plus 111 In-labeled cMORF to direct targeting by 111 In-labeled HPi1. Results: HPi1 binding to betalox5 human cells in the transplant was shown by immunofluorescence. Normal organ 111 In backgrounds by pretargeting were always lower, although target accumulations were similar. More importantly, the transplant to pancreas and liver ratios was, respectively, 26 and 10 by pretargeting as compared to 9 and 0.6 by direct targeting. Conclusions: Pretargeting greatly improves the T/NT ratios, and based on the estimated endocrine to exocrine ratio within a pancreas, pretargeting may be approaching the sensitivity required for successful imaging of human islets within this organ.

Activin receptor-like kinase 5 inhibition reverses impairment of endothelial cell viability by endogenous islet mesenchymal stromal cells.

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Clarkin, Claire E; King, Aileen J; Dhadda, Paramjeet; Chagastelles, Pedro; Nardi, Nance; Wheeler-Jones, Caroline P; Jones, Peter M

2013-03-01

Following islet transplantation, islet graft revascularization is compromised due to loss of endothelial cells (ECs) during islet culture. TGF-β signaling pathways are essential for vascular homeostasis but their importance for islet EC function is unclear. We have identified a population of multipotent mesenchymal stromal cells (MSCs) within islets and investigated how modulation of TGF-β signaling by these cells influences islet EC viability. Cultured islets exhibited reduced expression of EC markers (VEGFR2, VE-cadherin and CD31), which was associated with diminished but sustained expression of endoglin a marker of both ECs and MSCs. Double fluorescent labeling of islets in situ with the EC marker CD31 disclosed a population of CD31-negative cells which were positive for endoglin. In vitro coculture of microvascular ECs with endoglin-positive, CD31-negative islet MSCs reduced VEGFR2 protein expression, disrupted EC angiogenic behavior, and increased EC detachment. Medium conditioned by islet MSCs significantly decreased EC viability and increased EC caspase 3/7 activity. EC:MSC cocultures showed enhanced Smad2 phosphorylation consistent with altered ALK5 signaling. Pharmacological inhibition of ALK5 activity with SB431542 (SB) improved EC survival upon contact with MSCs, and SB-treated cultured islets retained EC marker expression and sensitivity to exogenous VEGF164 . Thus, endoglin-expressing islet MSCs influence EC ALK5 signaling in vitro, which decreases EC viability, and changes in ALK5 activity in whole cultured islets contribute to islet EC loss. Modifying TGF-β signaling may enable maintenance of islet ECs during islet isolation and thus improve islet graft revascularization post-transplantation. Copyright © 2013 AlphaMed Press.

Obesity Resistance Promotes Mild Contractile Dysfunction Associated with Intracellular Ca2+ Handling

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Sá, Felipe Gonçalves dos Santos de; Lima-Leopoldo, Ana Paula; Jacobsen, Bruno Barcellos; Ferron, Artur Junio Togneri; Estevam, Wagner Muller; Campos, Dijon Henrique Salomé; Castardeli, Edson; Cunha, Márcia Regina Holanda da; Cicogna, Antonio Carlos; Leopoldo, André Soares

2015-01-01

Diet-induced obesity is frequently used to demonstrate cardiac dysfunction. However, some rats, like humans, are susceptible to developing an obesity phenotype, whereas others are resistant to that. To evaluate the association between obesity resistance and cardiac function, and the impact of obesity resistance on calcium handling. Thirty-day-old male Wistar rats were distributed into two groups, each with 54 animals: control (C; standard diet) and obese (four palatable high-fat diets) for 15 weeks. After the experimental protocol, rats consuming the high-fat diets were classified according to the adiposity index and subdivided into obesity-prone (OP) and obesity-resistant (OR). Nutritional profile, comorbidities, and cardiac remodeling were evaluated. Cardiac function was assessed by papillary muscle evaluation at baseline and after inotropic maneuvers. The high-fat diets promoted increase in body fat and adiposity index in OP rats compared with C and OR rats. Glucose, lipid, and blood pressure profiles remained unchanged in OR rats. In addition, the total heart weight and the weight of the left and right ventricles in OR rats were lower than those in OP rats, but similar to those in C rats. Baseline cardiac muscle data were similar in all rats, but myocardial responsiveness to a post-rest contraction stimulus was compromised in OP and OR rats compared with C rats. Obesity resistance promoted specific changes in the contraction phase without changes in the relaxation phase. This mild abnormality may be related to intracellular Ca2+ handling

Automated Analysis of Microscopic Images of Isolated Pancreatic Islets

Czech Academy of Sciences Publication Activity Database

Habart, D.; Švihlík, J.; Schier, Jan; Cahová, M.; Girman, P.; Zacharovová, K.; Berková, Z.; Kříž, J.; Fabryová, E.; Kosinová, L.; Papáčková, Z.; Kybic, J.; Saudek, F.

2016-01-01

Roč. 25, č. 12 (2016), s. 2145-2156 ISSN 0963-6897 Grant - others:GA ČR(CZ) GA14-10440S Institutional support: RVO:67985556 Keywords : enumeration of islets * image processing * image segmentation * islet transplantation * machine-learning * quality control Subject RIV: IN - Informatics, Computer Science Impact factor: 3.006, year: 2016 http://library.utia.cas.cz/separaty/2016/ZOI/schier-0465945.pdf

Salvage Islet Auto Transplantation After Relaparatomy.

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Balzano, Gianpaolo; Nano, Rita; Maffi, Paola; Mercalli, Alessia; Melzi, Raffaelli; Aleotti, Francesca; Gavazzi, Francesca; Berra, Cesare; De Cobelli, Francesco; Venturini, Massimo; Magistretti, Paola; Scavini, Marina; Capretti, Giovanni; Del Maschio, Alessandro; Secchi, Antonio; Zerbi, Alessandro; Falconi, Massimo; Piemonti, Lorenzo

2017-10-01

To assess feasibility, safety, and metabolic outcome of islet auto transplantation (IAT) in patients undergoing completion pancreatectomy because of sepsis or bleeding after pancreatic surgery. From November 2008 to October 2016, approximately 22 patients were candidates to salvage IAT during emergency relaparotomy because of postpancreatectomy sepsis (n = 11) or bleeding (n = 11). Feasibility, efficacy, and safety of salvage IAT were compared with those documented in a cohort of 36 patients who were candidate to simultaneous IAT during nonemergency preemptive completion pancreatectomy through the pancreaticoduodenectomy. The percentage of candidates that received the infusion of islets was significantly lower in salvage IAT than simultaneous IAT (59.1% vs 88.9%, P = 0.008), mainly because of a higher rate of inadequate islet preparations. Even if microbial contamination of islet preparation was significantly higher in candidates to salvage IAT than in those to simultaneous IAT (78.9% vs 20%, P < 0.001), there was no evidence of a higher rate of complications related to the procedure. Median follow-up was 5.45 ± 0.52 years. Four (36%) of 11 patients reached insulin independence, 6 patients (56%) had partial graft function, and 1 patient (9%) had primary graft nonfunction. At the last follow-up visit, median fasting C-peptide was 0.43 (0.19-0.93) ng/mL; median insulin requirement was 0.38 (0.04-0.5) U/kg per day, and median HbA1c was 6.6% (5.9%-8.1%). Overall mortality, in-hospital mortality, metabolic outcome, graft survival, and insulin-free survival after salvage IAT were not different from those documented after simultaneous IAT. Our data demonstrate the feasibility, efficacy, and safety of salvage IAT after relaparotomy.

Engraftment versus immunosuppression: cost-benefit analysis of immunosuppression after intrahepatic murine islet transplantation.

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Marzorati, Simona; Melzi, Raffaella; Citro, Antonio; Cantarelli, Elisa; Mercalli, Alessia; Scavini, Marina; Piemonti, Lorenzo

2014-05-27

Immunosuppression (IS) in islet transplantation (Tx) is a double-edged sword: it prevents immunoreaction but has the potential to impair islet engraftment. The aim of this study was to identify in murine animal models the IS platform with the best balance between these two opposite effects. To study the impact of IS on islet engraftment diabetic C57BL/6 mice were transplanted with 350 syngeneic islets through the portal vein and treated once-daily with either rapamycin (RAPA; 0.1-0.5-1 mg/kg ip), tacrolimus (FK506; 0.1-0.5-1 mg/kg ip), mycophenolate mofetil (MMF; 60-120-300 mg/kg oral) or vehicle for 14 days. Islet function was evaluated by measuring not-fasting glycemia and by performing an IVGTT on days 15 and 30 post-Tx. RAPA ≥0.5 mg/Kg, FK506 ≥0.5 mg/Kg, and MMF ≥120 mg/kg had detrimental effects on islet engraftment but not on the function of islets already engrafted in the liver. The effect on engraftment was irreversible and persisted even after IS withdrawal. The lower dose of IS that did not affect engraftment was tested for preventing rejection in the full mismatch allogeneic Tx BALB/c to C57BL/6 model. RAPA and/or FK506 were inefficient in preventing rejection, even when anti-IL2R mAb was added to the IS regimen. On the other hand, MMF alone or in association with FK506 significantly prolonged the time to islet rejection. IS showed profound dose-dependent deleterious effects on islet cell engraftment. The MMF/FK506 combination proved the best balance with less toxicity at the time of engraftment and more efficacy in controlling graft rejection.

CT features of nonfunctioning islet cell carcinoma

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Eelkema, E.A.; Stephens, D.H.; Ward, E.M.; Sheedy, P.F. II

1984-11-01

To determine the computed tomographic (CT) characteristics of nonfunctioning islet cell carcinoma of the pancreas, the CT scans of 27 patients with that disease were reviewed. The pancreatic tumor was identified as a mass in 26 patients (96%) Of the 25 tumors evaluated with contrast enhancement, 20 became partially diffusely hyperdense relative to nearby normal pancreatic tissue. Hepatic metastases were identified in 15 patients (56%), regional lymphadenopathy in 10 (37%), atrophy of the gland proximal to the tumor in six (22%), dilatation of the biliary ducts in five (19%), and dilatation of the pancreatic duct in four (15%). The CT appearances of the nonfunctioning islet cell tumors were compared with those of 100 ordinary (ductal) pancreatic adenocarcinomas. Although the two types of tumors were sometimes indistinguishable, features found to be more characteristic of islet cell carcinoma included a pancreatic mass of unusually large size, calcification within the tumor, and contrast enhancement of either the primary tumor or hepatic metastases. Involvement of the celiac axis or proximal superior mesenteric artery was limited to ductal carcinoma.

Microencapsulated 3-Dimensional Sensor for the Measurement of Oxygen in Single Isolated Pancreatic Islets

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Khalil, Gamal; Sweet, Ian R.; Shen, Amy Q.

2012-01-01

Background Oxygen consumption reflects multiple processes in pancreatic islets including mechanisms contributing to insulin secretion, oxidative stress and viability, providing an important readout in studies of islet function, islet viability and drug testing. Due to the scarcity, heterogeneity, and intrinsic kinetic properties of individual islets, it would be of great benefit to detect oxygen consumption by single islets. We present a novel method we have developed to image oxygen in single islets. Methodology/Principal Findings Using a microfluidics system, individual islets and a fluorescent oxygen-sensitive dye were encased within a thin alginate polymer layer. Insulin secretion by the encapsulated islets was normal. Fluorescent signal from the encased dye, detected using a standard inverted fluorescence microscope and digital camera, was stable and proportional to the amount of oxygen in the media. When integrated into a perifusion system, the sensing system detected changes in response to metabolic substrates, mitochondrial poisons, and induced-oscillations. Glucose responses averaged 30.1±7.1% of the response to a metabolic inhibitor (cyanide), increases were observed in all cases (n = 6), and the system was able to resolve changes in oxygen consumption that had a period greater than 0.5 minutes. The sensing system operated similarly from 2–48 hours following encapsulation, and viability and function of the islets were not significantly affected by the encapsulation process. Conclusions/Significance An oxygen-dependent dye situated around and within a pancreatic islet encapsulated by a thin layer of alginate was sensitive to changes in oxygen consumption, and was not harmful to the function or viability of islets over the course of two days. The microcapsule-based sensing method is particularly suited to assessing the effects of compounds (dose responses and time courses) and chronic changes occurring over the course of days. The approach should be

The intrauterine metabolic environment modulates the gene expression pattern in fetal rat islets: prevention by maternal taurine supplementation

DEFF Research Database (Denmark)

Reusens, B; Sparre, T; Kalbe, L

2008-01-01

in gene expression in fetal islets affected by the LP diet and how taurine may prevent these changes. Methods  Pregnant Wistar rats were fed an LP diet (8% [wt/wt] protein) supplemented or not with taurine in the drinking water or a control diet (20% [wt/wt] protein). At 21.5 days of gestation, fetal......Aims/hypothesis  Events during fetal life may in critical time windows programme tissue development leading to organ dysfunction with potentially harmful consequences in adulthood such as diabetes. In rats, the beta cell mass of progeny from dams fed with a low-protein (LP) diet during gestation...

Single-donor islet transplantation in type 1 diabetes: patient selection and special considerations

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Tatum JA

2017-02-01

Full Text Available Jacob A Tatum,* Max O Meneveau,* Kenneth L Brayman Department of Surgery, Division of Transplantation, The University of Virginia Health System, Charlottesville, VA, USA *These authors contributed equally to this work. Abstract: Type 1 diabetes mellitus is an autoimmune disorder of the endocrine pancreas that currently affects millions of people in the United States. Although the disease can be managed with exogenous insulin administration, the ultimate cure for the condition lies in restoring a patient’s ability to produce their own insulin. Islet cell allotransplantation provides a means of endogenous insulin production. Though far from perfected, islet transplants are now a proven treatment for type 1 diabetics. However, proper patient selection is critical for achieving optimal outcomes. Given the shortage of transplantable organs, selecting appropriate candidates for whom the procedure will be of greatest benefit is essential. Although many of those who receive islets do not retain insulin independence, grafts do play a significant role in preventing hypoglycemic episodes that can be quite detrimental to quality of life and potentially fatal. Additionally, islet transplant requires lifelong immunosuppression. Antibodies, both preformed and following islet infusion, may play important roles in graft outcomes. Finally, no procedure is without inherent risk and islet transfusions can have serious consequences for recipients’ livers in the form of both vascular and metabolic complications. Therefore, patient-specific factors that should be taken into account before islet transplantation include aims of therapy, sensitization, and potential increased risk for hepatic and portal-venous sequelae. Keywords: islet transplantation, diabetes mellitus type 1, brittle diabetes, single donor, patient

Chemokine receptor expression in tumour islets and stroma in non-small cell lung cancer

International Nuclear Information System (INIS)

Ohri, Chandra M; Shikotra, Aarti; Green, Ruth H; Waller, David A; Bradding, Peter

2010-01-01

We have previously demonstrated that tumour islet infiltration by macrophages is associated with extended survival (ES) in NSCLC. We therefore hypothesised that patients with improved survival would have high tumour islet expression of chemokine receptors known to be associated with favourable prognosis in cancer. This study investigated chemokine receptor expression in the tumour islets and stroma in NSCLC. We used immunohistochemistry to identify cells expressing CXCR1, CXCR2, CXCR3, CXCR4, CXCR5 and CCR1 in the tumour islets and stroma in 20 patients with surgically resected NSCLC. Correlations were made with macrophage and mast cell expression. There was increased expression of CXCR2, CXCR3, and CCR1 in the tumour islets of ES compared with poor survival (PS) patients (p = 0.007, 0.01, and 0.002, respectively). There was an association between 5 year survival and tumour islet CXCR2, CXCR3 and CCR1 density (p = 0.02, 0.003 and <0.001, respectively) as well as stromal CXCR3 density (p = 0.003). There was a positive correlation between macrophage density and CXCR3 expression (r s = 0.520, p = 0.02) and between mast cell density and CXCR3 expression (r s = 0.499, p = 0.03) in the tumour islets. Above median expression of CXCR2, CXCR3 and CCR1 in the tumour islets is associated with increased survival in NSCLC, and expression of CXCR3 correlates with increased macrophage and mast cell infiltration in the tumour islets

Insulin resistance and progression to type 1 diabetes in the European Nicotinamide Diabetes Intervention Trial (ENDIT)

DEFF Research Database (Denmark)

Bingley, Polly J; Mahon, Jeffrey L; Gale, Edwin A M

2008-01-01

OBJECTIVE: Insulin resistance can modulate progression to type 1 diabetes in individuals with ongoing islet autoimmunity. We wanted to see whether measures of insulin resistance improved risk assessment in islet cell antibody (ICA)-positive relatives when added to other immune and metabolic markers......-up was 4.21 years, and 105 individuals developed diabetes. Oral and intravenous glucose tolerance tests were performed at baseline; antibodies to GAD, IA-2, and insulin were determined by radioimmunoassay; and insulin resistance was estimated by homeostasis model assessment. Risk was assessed by Cox...... glucose tolerance test (P insulin resistance (HOMA2-IR) achieved only borderline significance (P = 0.06). HOMA2-IR was an independent determinant in participants with loss of FPIR (P = 0...

Essential role of the small GTPase Ran in postnatal pancreatic islet development.

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Fang Xia

Full Text Available The small GTPase Ran orchestrates pleiotropic cellular responses of nucleo-cytoplasmic shuttling, mitosis and subcellular trafficking, but whether deregulation of these pathways contributes to disease pathogenesis has remained elusive. Here, we generated transgenic mice expressing wild type (WT Ran, loss-of-function Ran T24N mutant or constitutively active Ran G19V mutant in pancreatic islet β cells under the control of the rat insulin promoter. Embryonic pancreas and islet development, including emergence of insulin(+ β cells, was indistinguishable in control or transgenic mice. However, by one month after birth, transgenic mice expressing any of the three Ran variants exhibited overt diabetes, with hyperglycemia, reduced insulin production, and nearly complete loss of islet number and islet mass, in vivo. Deregulated Ran signaling in transgenic mice, adenoviral over-expression of WT or mutant Ran in isolated islets, or short hairpin RNA (shRNA silencing of endogenous Ran in model insulinoma INS-1 cells, all resulted in decreased expression of the pancreatic and duodenal homeobox transcription factor, PDX-1, and reduced β cell proliferation, in vivo. These data demonstrate that a finely-tuned balance of Ran GTPase signaling is essential for postnatal pancreatic islet development and glucose homeostasis, in vivo.

Ionic and secretory response of pancreatic islet cells to minoxidil sulfate

International Nuclear Information System (INIS)

Antoine, M.H.; Hermann, M.; Herchuelz, A.; Lebrun, P.

1991-01-01

Minoxidil sulfate is an antihypertensive agent belonging to the new class of vasodilators, the K+ channel openers. The present study was undertaken to characterize the effects of minoxidil sulfate on ionic and secretory events in rat pancreatic islets. The drug unexpectedly provoked a concentration-dependent decrease in 86Rb outflow. This inhibitory effect was reduced in a concentration-dependent manner by glucose and tolbutamide. Minoxidil sulfate did not affect 45Ca outflow from islets perfused in the presence of extracellular Ca++ and absence or presence of glucose. However, in islets exposed to a medium deprived of extracellular Ca++, the drug provoked a rise in 45Ca outflow. Whether in the absence or presence of extracellular Ca++, minoxidil sulfate increased the cytosolic free Ca++ concentration of islet cells. Lastly, minoxidil sulfate increased the release of insulin from glucose-stimulated pancreatic islets. These results suggest that minoxidil sulfate reduces the activity of the ATP-sensitive K+ channels and promotes an intracellular translocation of Ca++. The latter change might account for the effect of the drug on the insulin-releasing process. However, the secretory response to minoxidil sulfate could also be mediated, at least in part, by a modest Ca++ entry

The Peri-islet Basement Membrane, a Barrier to Infiltrating Leukocytes in Type 1 Diabetes in Mouse and Human

DEFF Research Database (Denmark)

Korpos, Eva; Kadri, Nadir; Kappelhoff, Reinhild

2013-01-01

We provide the first comprehensive analysis of the extracellular matrix (ECM) composition of peri-islet capsules, composed of the peri-islet basement membrane (BM) and subjacent interstitial matrix (IM), in development of type 1 diabetes in NOD mice and in human type 1 diabetes. Our data demonstr...... IM are reconstituted once inflammation subsides, indicating that the peri-islet BM-producing cells are not lost due to the inflammation, which has important ramifications to islet transplantation studies.......We provide the first comprehensive analysis of the extracellular matrix (ECM) composition of peri-islet capsules, composed of the peri-islet basement membrane (BM) and subjacent interstitial matrix (IM), in development of type 1 diabetes in NOD mice and in human type 1 diabetes. Our data...... demonstrate global loss of peri-islet BM and IM components only at sites of leukocyte infiltration into the islet. Stereological analyses reveal a correlation between incidence of insulitis and the number of islets showing loss of peri-islet BM versus islets with intact BMs, suggesting that leukocyte...

Long-Term Follow-Up of the Edmonton Protocol of Islet Transplantation in the United States.

Science.gov (United States)

Brennan, D C; Kopetskie, H A; Sayre, P H; Alejandro, R; Cagliero, E; Shapiro, A M J; Goldstein, J S; DesMarais, M R; Booher, S; Bianchine, P J

2016-02-01

We report the long-term follow-up of the efficacy and safety of islet transplantation in seven type 1 diabetic subjects from the United States enrolled in the multicenter international Edmonton Protocol who had persistent islet function after completion of the Edmonton Protocol. Subjects were followed up to 12 years with serial testing for sustained islet allograft function as measured by C-peptide. All seven subjects demonstrated continued islet function longer than a decade from the time of first islet transplantation. One subject remained insulin independent without the need for diabetic medications or supplemental transplants. One subject who was insulin-independent for over 8 years experienced graft failure 10.9 years after the first islet transplant. The remaining six subjects demonstrated continued islet function upon trial completion, although three had received a supplemental islet transplant each. At trial completion, five subjects were receiving insulin and two remained insulin independent, although one was treated with liraglutide. The median hemoglobin A1c was 6.3% (45 mmol/mol). All subjects experienced progressive decline in the C-peptide/glucose ratio. No patients experienced severe hypoglycemia, opportunistic infection, or lymphoma. Thus, although the rate and duration of insulin independence was low, the Edmonton Protocol was safe in the long term. Alternative approaches to islet transplantation are under investigation. © Copyright 2015 The American Society of Transplantation and the American Society of Transplant Surgeons.

Isolation, transplantation, and functional studies of adult porcine islets of Langerhans

DEFF Research Database (Denmark)

Nielsen, Thomas Buschmann; Yderstræde, Knud Bonnet; Beck-Nielsen, Henning

2002-01-01

that was only partially increased by additional challenge with arginine. More than 50% of DNA and 90% of the insulin content was lost during a one-week culture period. With some batch-to-batch variation, in 15 of 25 cases, 4,000 to 7,000 porcine islets cured streptozotocin diabetic nude mice within three weeks......Transplantation of islets of Langerhans is a possible treatment for type-I diabetes mellitus. However, there is a shortage of donors for such transplantations and the pig may be an alternative source of donor organs. The aims of the study reported here were to establish a method for adult porcine...... following transplantation. In conclusion, it is possible to isolate viable islets from adult pigs, using a semiautomatic set-up. With batch-to-batch variation, the islets are able to revert diabetes mellitus when transplanted to diabetic nude mice....

Intraocular in vivo imaging of pancreatic islet cell physiology/pathology

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Ingo B. Leibiger

2017-09-01

Major conclusions: Data provided by us and others demonstrate the high versatility of this imaging platform. The use of ‘reporter islets’ engrafted in the eye, reporting on the status of in situ endogenous islets in the pancreas of the same animal, allows the identification of key-events in the development and progression of diabetes. This will not only serve as a versatile research tool but will also lay the foundation for a personalized medicine approach and will serve as a screening platform for new drugs and/or treatment protocols. ‘Metabolic’ islet transplantation, in which islets engrafted in the eye replace the endogenous beta cells, will allow for the establishment of islet-specific transgenic models and ‘humanized’ mouse models as well as serving as the basis for a new clinical transplantation site for the cure of diabetes.

Gamma irradiation of isolated rat islets pretransplantation produces indefinite allograft survival in cyclosporine-treated recipients

International Nuclear Information System (INIS)

James, R.F.; Lake, S.P.; Chamberlain, J.; Thirdborough, S.; Bassett, P.D.; Mistry, N.; Bell, P.R.

1989-01-01

In this study we have examined the use of low-dose gamma-irradiation for the reduction of islet immunogenicity in the strong allogeneic combination of WAG rat islets transplanted into diabetic AUG recipients. First, we determined that gamma-irradiation reduced immunogenicity in vitro by use of a modified MLR with WAG islets as stimulators and AUG splenocytes as responders. We then determined the maximum dose of gamma-irradiation that could be used (250 rads) before islet function was affected. As 250 rads islet pretreatment alone was ineffective in prolonging allograft survival, we combined the pretreatment with a short course (days 0, 1, 2; 30 mg/kg) of cyclosporine. We found that CsA was only effective in significantly prolonging allograft survival when given subcutaneously in olive oil. The CsA treatment alone gave a significantly prolonged survival time for the islet allografts (median, 37 days vs. 6 days for controls), but when combined with the 250 rads islet pretreatment a synergistic effect was seen with 100% becoming long-term survivors (greater than 100 days). The long-term surviving AUG rats from both the CsA alone group and the CsA plus 250 rads pretreated islets group were challenged with WAG dendritic cells (DC). The islets from the 250 rads pretreated group were subsequently rejected (day 6) while the CsA alone group were not affected. The role of low dose gamma-irradiation when combined with CsA treatment of islet graft recipients in inducing specific unresponsiveness will be discussed

Gamma irradiation of isolated rat islets pretransplantation produces indefinite allograft survival in cyclosporine-treated recipients

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James, R.F.; Lake, S.P.; Chamberlain, J.; Thirdborough, S.; Bassett, P.D.; Mistry, N.; Bell, P.R.

1989-06-01

In this study we have examined the use of low-dose gamma-irradiation for the reduction of islet immunogenicity in the strong allogeneic combination of WAG rat islets transplanted into diabetic AUG recipients. First, we determined that gamma-irradiation reduced immunogenicity in vitro by use of a modified MLR with WAG islets as stimulators and AUG splenocytes as responders. We then determined the maximum dose of gamma-irradiation that could be used (250 rads) before islet function was affected. As 250 rads islet pretreatment alone was ineffective in prolonging allograft survival, we combined the pretreatment with a short course (days 0, 1, 2; 30 mg/kg) of cyclosporine. We found that CsA was only effective in significantly prolonging allograft survival when given subcutaneously in olive oil. The CsA treatment alone gave a significantly prolonged survival time for the islet allografts (median, 37 days vs. 6 days for controls), but when combined with the 250 rads islet pretreatment a synergistic effect was seen with 100% becoming long-term survivors (greater than 100 days). The long-term surviving AUG rats from both the CsA alone group and the CsA plus 250 rads pretreated islets group were challenged with WAG dendritic cells (DC). The islets from the 250 rads pretreated group were subsequently rejected (day 6) while the CsA alone group were not affected. The role of low dose gamma-irradiation when combined with CsA treatment of islet graft recipients in inducing specific unresponsiveness will be discussed.

Transplantation of micro- and macroencapsulated piglet islets into mice and monkeys.

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Elliott, R B; Escobar, L; Calafiore, R; Basta, G; Garkavenko, O; Vasconcellos, A; Bambra, C

2005-01-01

Neonatal porcine islets within alginate microcapsules transplanted intraperitoneally (IP) or within semi-permeable macrocapsules (TheraCyte) and transplanted subcutaneously (SC) survive and reverse diabetes for up to 16 weeks in diabetic autoimmune nonobese diabetic (NOD) mice. The islets in microcapsules transplanted IP into nondiabetic cynomolgus monkeys survived for 8 weeks. Similar results were shown with islets transplanted in TheraCytes. Neither species showed adverse effects or evidence of infection with porcine endogenous retroviruses or other endemic pig viruses. Proof of principle is illustrated for successful xenotransplantation in humans.

Resistance Training in Type II Diabetes Mellitus: Impact on Areas of Metabolic Dysfunction in Skeletal Muscle and Potential Impact on Bone

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Richard J. Wood

2012-01-01

Full Text Available The prevalence of Type II Diabetes mellitus (T2DM is increasing rapidly and will continue to be a major healthcare expenditure burden. As such, identification of effective lifestyle treatments is paramount. Skeletal muscle and bone display metabolic and functional disruption in T2DM. Skeletal muscle in T2DM is characterized by insulin resistance, impaired glycogen synthesis, impairments in mitochondria, and lipid accumulation. Bone quality in T2DM is decreased, potentially due to the effects of advanced glycation endproducts on collagen, impaired osteoblast activity, and lipid accumulation. Although exercise is widely recognized as an important component of treatment for T2DM, the focus has largely been on aerobic exercise. Emerging research suggests that resistance training (strength training may impose potent and unique benefits in T2DM. The purpose of this review is to examine the role of resistance training in treating the dysfunction in skeletal muscle and the potential role for resistance training in treating the associated dysfunction in bone.

Attenuation of primary nonfunction for syngeneic islet graft using sodium 4-phenylbutyrate.

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Fu, S-H; Chen, S-T; Hsu, B R-S

2005-05-01

Sodium 4-phenylbutyrate (4-SPB), an aromatic derivative of butyric acid, was examined to elucidate its effect on islet engraftment in a syngeneic transplantation model using C57BL/6 mice. Diabetic mice that received subrenal implantation of 150 islets on day 0 and oral administration of twice daily 4-SPB (500 mg/kg body weight) on days -2 through 28 displayed a significantly shorter duration of posttransplantation temporary hyperglycemia than diabetic mice that received islets in isotonic sodium chloride solution (NaCl), namely 16 +/- 2 (n = 12) vs 23 +/- 2 days (n = 7; P < .05). Four weeks after transplantation, the insulin content (IC) of grafts from mice treated with islets and 4-SPB was substantially higher than that of grafts from mice treated with islets and NaCl, namely 2.59 +/- 0.37 (n = 8) vs 1.36 +/- 0.36 mug (n = 13; P < .01). The IC of pancreatic remnants showed no significant difference between groups after 2 and 4 weeks of incubation. In vitro studies demonstrated that the net glucose-stimulated insulin secretion (GSIS) and the ratio of net GSIS to the IC of islets cultured with 4-SPB (1 mM) did not differ significantly from those cultured with NaCl. The lipopolysaccharide-stimulated secretions of IL-1beta, IL-10, and IFNgamma from peritoneal exudate monocytes were significantly reduced by co-incubation with 4-SPB (1 mM). In conclusion, our data suggest that daily administration of 4-SPB reduces primary nonfunction and enhances islet engraftment in a syngeneic mouse transplantation model.

Methylated trivalent arsenicals are potent inhibitors of glucose stimulated insulin secretion by murine pancreatic islets

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Douillet, Christelle [Department of Nutrition, Gillings School of Global Public Health, 2302 MHRC, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7461 (United States); Currier, Jenna [Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7461 (United States); Saunders, Jesse [Department of Nutrition, Gillings School of Global Public Health, 2302 MHRC, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7461 (United States); Bodnar, Wanda M. [Department of Environmental Sciences and Engineering, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7431 (United States); Matoušek, Tomáš [Institute of Analytical Chemistry of the ASCR, v.v.i., Veveří 97, 602 00 Brno (Czech Republic); Stýblo, Miroslav, E-mail: styblo@med.unc.edu [Department of Nutrition, Gillings School of Global Public Health, 2302 MHRC, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7461 (United States)

2013-02-15

Epidemiologic evidence has linked chronic exposure to inorganic arsenic (iAs) with an increased prevalence of diabetes mellitus. Laboratory studies have identified several mechanisms by which iAs can impair glucose homeostasis. We have previously shown that micromolar concentrations of arsenite (iAs{sup III}) or its methylated trivalent metabolites, methylarsonite (MAs{sup III}) and dimethylarsinite (DMAs{sup III}), inhibit the insulin-activated signal transduction pathway, resulting in insulin resistance in adipocytes. Our present study examined effects of the trivalent arsenicals on insulin secretion by intact pancreatic islets isolated from C57BL/6 mice. We found that 48-hour exposures to low subtoxic concentrations of iAs{sup III}, MAs{sup III} or DMAs{sup III} inhibited glucose-stimulated insulin secretion (GSIS), but not basal insulin secretion. MAs{sup III} and DMAs{sup III} were more potent than iAs{sup III} as GSIS inhibitors with estimated IC{sub 50} ≤ 0.1 μM. The exposures had little or no effects on insulin content of the islets or on insulin expression, suggesting that trivalent arsenicals interfere with mechanisms regulating packaging of the insulin transport vesicles or with translocation of these vesicles to the plasma membrane. Notably, the inhibition of GSIS by iAs{sup III}, MAs{sup III} or DMAs{sup III} could be reversed by a 24-hour incubation of the islets in arsenic-free medium. These results suggest that the insulin producing pancreatic β-cells are among the targets for iAs exposure and that the inhibition of GSIS by low concentrations of the methylated metabolites of iAs may be the key mechanism of iAs-induced diabetes. - Highlights: ► Trivalent arsenicals inhibit glucose stimulated insulin secretion by pancreatic islets. ► MAs{sup III} and DMAs{sup III} are more potent inhibitors than arsenite with IC{sub 50} ∼ 0.1 μM. ► The arsenicals have little or no effects on insulin expression in pancreatic islets. ► The inhibition of

Amelioration of Mitochondrial Dysfunction-Induced Insulin Resistance in Differentiated 3T3-L1 Adipocytes via Inhibition of NF-κB Pathways

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Mohamad Hafizi Abu Bakar

2014-12-01

Full Text Available A growing body of evidence suggests that activation of nuclear factor kappa B (NF-κB signaling pathways is among the inflammatory mechanism involved in the development of insulin resistance and chronic low-grade inflammation in adipose tissues derived from obese animal and human subjects. Nevertheless, little is known about the roles of NF-κB pathways in regulating mitochondrial function of the adipose tissues. In the present study, we sought to investigate the direct effects of celastrol (potent NF-κB inhibitor upon mitochondrial dysfunction-induced insulin resistance in 3T3-L1 adipocytes. Celastrol ameliorates mitochondrial dysfunction by altering mitochondrial fusion and fission in adipocytes. The levels of oxidative DNA damage, protein carbonylation and lipid peroxidation were down-regulated. Further, the morphology and quantification of intracellular lipid droplets revealed the decrease of intracellular lipid accumulation with reduced lipolysis. Moreover, massive production of the pro-inflammatory mediators tumor necrosis factor-α (TNF-α and interleukin-1β (IL-1β were markedly depleted. Insulin-stimulated glucose uptake activity was restored with the enhancement of insulin signaling pathways. This study signified that the treatments modulated towards knockdown of NF-κB transcription factor may counteract these metabolic insults exacerbated in our model of synergy between mitochondrial dysfunction and inflammation. These results demonstrate for the first time that NF-κB inhibition modulates mitochondrial dysfunction induced insulin resistance in 3T3-L1 adipocytes.

Anti-inflammatory thalidomide improves islet grafts survival and functions in a xenogenic environment.

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Chunguang Chen

Full Text Available Thalidomide possesses both anti-inflammatory and anti-angiogenic properties. This study investigates its potential application in islet transplantation with a xenogenic transplantation model. Transplantation was performed using C57Bl/6 mice and NMRI nu/nu mice as recipients of porcine islets. Moreover, islet graft vasculature and inflammation were investigated to identify the mechanisms of thalidomide action. In the immunocompetent environment of C57Bl/6 mice, a fast graft rejection was observed. The group treated with thalidomide 200 mg/kg BW per day achieved and maintained euglycemia in the complete observation period for 42 days. The treated mice had more functional islet graft mass with less leukocyte infiltration. The pro-inflammatory TNF-alpha and VEGF content in islet grafted kidneys was significantly lowered by the treatment. By comparison, thalidomide was not effective in improving graft survival in immunocompromised nude mice. It strongly inhibited the VEGF and TNF-alpha-induced endothelial proliferation of isolated pig islets in a dose dependent manner. The magnitude of thalidomide's inhibitory effect was nearly identical to the effect of VEGF- receptor 2 inhibitor SU416 and anti-TNF-receptor 1 neutralizing antibody, and was reversed by sphingosine-1-phosphate. In conclusion, the anti-inflammatory effect of thalidomide improved islet graft survival and function in a transplantation model with a maximum immune barrier.

Prolonged Survival of Subcutaneous Allogeneic Islet Graft by Donor Chimerism without Immunosuppressive Treatment

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Brend Ray-Sea Hsu

2017-01-01

Full Text Available The aim of this study was to investigate whether tolerance-induced protection of islets in the renal subcapsular space can also prevent subcutaneous allogeneic islets from being rejected. We used bone marrow stem cells from C57BL/6 (H2b mice to construct donor chimerism in conditioned diabetic BALB/c (H2d mice and investigated the effect of donor chimerism on engraftment and survival of subcutaneously transplanted allogeneic islets in streptozotocin-induced diabetic mice. We also studied the anti-inflammatory effect of mesenchymal stem cell on islet engraftment. Full but not low-grade or no donor chimerism was associated with successful engraftment of allogeneic islets and restoration of normoglycemia in the treated diabetic mice. The temporary hyperglycemia was 11 ± 1 versus 19 ± 5 days (p<0.05 for the mice with full donor chimerism with transplanted islets in the renal subcapsular space versus the subcutaneous space, respectively. Cotransplantation of mesenchymal stem cell did not enhance alloislet engraftment. Full multilineage donor chimerism was associated with a higher transient expansion of CD11b+ and Gr-1+ myeloid progenitor cells and effector memory CD4 and CD8 T cells. In conclusion, full donor chimerism protected both renal subcapsular and subcutaneous allogeneic islets in this rodent transplantation model.

[Xenogeneic cell therapeutics: Treatment of type 1 diabetes using porcine pancreatic islets and islet cells].

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Godehardt, Antonia W; Schilling-Leiß, Dagmar; Sanzenbacher, Ralf; Tönjes, Ralf R

2015-11-01

In view of the existing shortage of human donor organs and tissues, xenogeneic cell therapeutics (xCT) offer an alternative for adequate treatment. In particular, porcine pancreatic islets and islet cells have already entered the field of experimental therapy for type-1 diabetes mellitus (T1DM) patients. Thereby, xCT depict challenging products with a glance on medical, ethical, and regulatory questions. With cross-species transplantation (xenotransplantation), the risk of immunological graft rejection as well as the risk of infectious transmission of microbial and viral pathogens must be considered. This includes the bidirectional transmission of microorganisms from graft to host as well as from host to graft. Crossing the border of species requires a critical risk-benefit evaluation as well as a thorough longtime surveillance of transplant recipients after treatment. The international legal and regulatory requirements for xCT are inter alia based on the World Health Organization criteria summarized in the Changsha Communiqué (2008). In the European Union, they were reflected by the European Medicines Agency (EMA) Guideline on Xenogeneic Cell-based Medicinal Products following the implementation of the Regulation on Advanced Therapies (ATMP). On the basis of this regulation, the first non-clinical and clinical experiences were obtained for porcine islets. The results suggest that supportive treatment of T1DM risk patients with xCT may be an alternative to established allogeneic organ transplantation in the future.

LIGHT/TNFSF14 is increased in patients with type 2 diabetes mellitus and promotes islet cell dysfunction and endothelial cell inflammation in vitro.

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Halvorsen, Bente; Santilli, Francesca; Scholz, Hanne; Sahraoui, Afaf; Gulseth, Hanne L; Wium, Cecilie; Lattanzio, Stefano; Formoso, Gloria; Di Fulvio, Patrizia; Otterdal, Kari; Retterstøl, Kjetil; Holven, Kirsten B; Gregersen, Ida; Stavik, Benedicte; Bjerkeli, Vigdis; Michelsen, Annika E; Ueland, Thor; Liani, Rossella; Davi, Giovanni; Aukrust, Pål

2016-10-01

Activation of inflammatory pathways is involved in the pathogenesis of type 2 diabetes mellitus. On the basis of its role in vascular inflammation and in metabolic disorders, we hypothesised that the TNF superfamily (TNFSF) member 14 (LIGHT/TNFSF14) could be involved in the pathogenesis of type 2 diabetes mellitus. Plasma levels of LIGHT were measured in two cohorts of type 2 diabetes mellitus patients (191 Italian and 40 Norwegian). Human pancreatic islet cells and arterial endothelial cells were used to explore regulation and relevant effects of LIGHT in vitro. Our major findings were: (1) in both diabetic cohorts, plasma levels of LIGHT were significantly raised compared with sex- and age-matched healthy controls (n = 32); (2) enhanced release from activated platelets seems to be an important contributor to the raised LIGHT levels in type 2 diabetes mellitus; (3) in human pancreatic islet cells, inflammatory cytokines increased the release of LIGHT and upregulated mRNA and protein levels of the LIGHT receptors lymphotoxin β receptor (LTβR) and TNF receptor superfamily member 14 (HVEM/TNFRSF14); (4) in these cells, LIGHT attenuated the insulin release in response to high glucose at least partly via pro-apoptotic effects; and (5) in human arterial endothelial cells, glucose boosted inflammatory response to LIGHT, accompanied by an upregulation of mRNA levels of HVEM (also known as TNFRSF14) and LTβR (also known as LTBR). Our findings show that patients with type 2 diabetes mellitus are characterised by increased plasma LIGHT levels. Our in vitro findings suggest that LIGHT may contribute to the progression of type 2 diabetes mellitus by attenuating insulin secretion in pancreatic islet cells and by contributing to vascular inflammation.

The Study of Non-Viral Nanoscale Delivery Systems for Islet Transplantation

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Gutierrez, Diana

Due to safety concerns associated with using viral systems clinically to expand islet cells and make them available to many more patients, significant emphasis has been placed on producing a safe and effective non-viral delivery system for biological research and gene therapy. To obtain this goal, we propose the use of an innovative technology that utilizes gold nanoparticles (AuNPs) as a non-viral method of delivery. Our laboratory was one of the first to describe the use of AuNPs in human islets and observe AuNPs can penetrate into the core of islets to deliver a gene to the vast majority of the cells, without damaging the cell. Gold nanoparticles proved to be a biocompatible delivery system both in vitro and in vivo. Thus far, gene therapy and molecular biology have focused primarily on delivering DNA of a specific gene into cells. The risk of this approach is that the DNA can be permanently incorporated into the genome and lead to damages in the cell that could result in overexpression of cancerous tumor cells. This risk does not exist with the use of mRNA. Many researchers believe mRNA is too unstable to be used as a molecular tool to overexpress specific proteins. With advances in nanotechnology, and better understanding of the translation process, methods have been developed that allow for expression of specific proteins by intracellular delivery of protein-encoding mRNA. We used AuNPs conjugated to mCherry mRNA to establish a proof of concept of the feasibility of using AuNP-mRNA to achieve increased expression of a specific protein within cells. To do this, we conjugated mCherry mRNA to AuNPs and tested the feasibility for increasing delivery efficacy and preserve functionality of human pancreatic islets. We believe that with this novel technology we can create AuNPs that allow specific mRNA to enter islets and lead to the production of a specific protein within the cell, with the aim to induce beta cell proliferation. In a previous experiment with single

Factors influencing the properties and performance of microcapsules for immunoprotection of pancreatic islets.

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van Schilfgaarde, R; de Vos, P

1999-01-01

There are several approaches of immunoprotection of pancreatic islets for the purpose of successful allo- or xenotransplantation in the absence of immunosuppressive medication. Extravascular approaches are either macroencapsulation (large numbers of islets together in one device) or microencapsulation. The latter approach is to envelop each individual islet in a semipermeable immunoprotective capsule. Quite promising results have been achieved with polylysine-alginate microencapsulated islet grafts in rodents, but clinical application is still restricted to a very small number of cases. Relevant considerations regard the following aspects. The biocompatibility of the microcapsules is influenced by the chemical composition of the materials applied and by mechanical factors related to the production process. With purified instead of crude alginates, the percentage of capsules with fibrotic overgrowth is reduced to approximately ten percent, and the remaining overgrowth is mainly explained by mechanical factors, i.e. inadequate encapsulation of individual islets. Even with purified alginates, however, the duration of encapsulated graft function is limited to a period of six to twenty weeks. Obviously, other factors than bioincompatibility play a role, which factors have to be identified. The limited duration of graft survival cannot be explained by rejection since, in rats, survival times of encapsulated isografts are similar, if not identical, to those of encapsulated allografts. An important factor is probably insufficient nutrition as a consequence of insufficient blood supply of the encapsulated and thus isolated islet. This also influences the functional performance of encapsulated islet grafts. Although normoglycemia can be readily obtained in streptozotocin diabetic rat recipients, glucose tolerance remains severely impaired, as a consequence of an insufficient increase of insulin levels in response to intravenous or oral glucose challenge. Important factors

A VERSATILE ALGINATE DROPLET GENERATOR APPLICABLE FOR MICROENCAPSULATION OF PANCREATIC-ISLETS

NARCIS (Netherlands)

WOLTERS, GHJ; FRITSCHY, WM; GERRITS, D; VANSCHILFAGAARDE, R

1992-01-01

Alginate beads for immunoisolation of pancreatic islets by microencapsulation should be small, smooth, and spherical in order to ensure that around the islets a strong alginate-polylysine-alginate capsule will be formed with optimal biocompatibility and diffusion of nutrients and hormones. However,

Pancreatic islet allograft in spleen with immunosuppression with cyclosporine. Experimental model in dogs.

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Waisberg, Jaques; Neff, Charles Benjamin; Waisberg, Daniel Reis; Germini, Demetrius; Gonçalves, José Eduardo; Zanotto, Arnaldo; Speranzini, Manlio Basilio

2011-01-01

To study the functional behavior of the allograft with immunosuppression of pancreatic islets in the spleen. Five groups of 10 Mongrel dogs were used: Group A (control) underwent biochemical tests; Group B underwent total pancreatectomy; Group C underwent total pancreatectomy and pancreatic islet autotransplant in the spleen; Group D underwent pancreatic islet allograft in the spleen without immunosuppressive therapy; Group E underwent pancreatic islet allograft in the spleen and immunosuppression with cyclosporine. All of the animals with grafts received pancreatic islets prepared by the mechanical-enzymatic method - stationary collagenase digestion and purification with dextran discontinuous density gradient, implanted in the spleen. The animals with autotransplant and those with allografts with immunosuppression that became normoglycemic showed altered results of intravenous tolerance glucose (p < 0.001) and peripheral and splenic vein plasmatic insulin levels were significantly lower (p < 0.001) in animals that had allografts with immunosuppression than in those with just autotransplants. In the animals with immunosuppression with cyclosporine subjected to allograft of pancreatic islets prepared with the mechanical-enzymatic preparation method (stationary collagenase digestion and purification with dextran discontinuous density gradient), the production of insulin is decreased and the response to intravenous glucose is altered.

Identification of transplanted pancreatic islet cells by radioactive Dithizone-[131I]-Histamine conjugate. Preliminary report

International Nuclear Information System (INIS)

Garnuszek, P.; Licinska, I.; Mazurek, A.P.; Mrozek, A.; Wardawa, A.; Fiedor, P.S.

2000-01-01

Background: The unique mechanism of dithizone action in the interior of the viable pancreatic islet suggests the possible development of a specific radiopharmaceutical that may have a potential clinical application in the diagnosis of the pancreatic organ allografts or islets rejection. The radiodiagnostic properties of the newly developed radioactive analogue of dithizone, i.e. Dithizone-[131I]-Histamine conjugate have been evaluated in the present study. METHODS: The four islet cells transplantation models were chosen for this purpose. The most important feature of the Dithizone-[131I]-Histamine conjugate is its possessed ability of zinc chelation. As was presented in the recent study, the conjugate stains pink-reddish the isolated pancreatic islets in vitro. Among the studied transplantation models, only the islets grafting under testis capsule enabled determination of the pancreatic islets in rats by radioactive Dithizone-[131I]-Histamine conjugate. The level of the radioactivity in the recipient testis (right) was almost two times higher compared to the controls (0.24 v. 0.13% ID/g, respectively). CONCLUSIONS: These preliminary data demonstrate the ability of the developed radioactive analogue of dithizone for in vivo identification of transplanted pancreatic islets, and suggests a potential clinical application of the radiodithizone in the diagnosis of the pancreatic islet rejection. (author)

Islet graft survival and function: concomitant culture and transplantation with vascular endothelial cells in diabetic rats.

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Pan, Xiaoming; Xue, Wujun; Li, Yang; Feng, Xinshun; Tian, Xiaohui; Ding, Chenguang

2011-12-15

Human islet transplantation is a great potential therapy for type I diabetes. To investigate islet graft survival and function, we recently showed the improved effects after co-culture and co-transplantation with vascular endothelial cells (ECs) in diabetic rats. ECs were isolated, and the viability of isolated islets was assessed in two groups (standard culture group and co-culture group with ECs). Then streptozotocin-induced diabetic rats were divided into four groups before islet transplantation as follows: group A with infusion of islet grafts; group B with combined vascular ECs and islet grafts; groups C and D as controls with single ECs infusion and phosphate-buffered saline injection, respectively. Blood glucose and insulin concentrations were measured daily. Expression of vascular endothelial growth factor was investigated by immunohistochemical staining. The mean microvascular density was also calculated. More than 90% of acridine orange-propidium iodide staining positive islets demonstrated normal morphology while co-cultured with ECs for 7 days. Compared with standard control, insulin release assays showed a significantly higher simulation index in co-culture group except for the first day (Ptransplantation, there was a significant difference in concentrations of blood glucose and insulin among these groups after 3 days (Pislet group (P=0.04). Co-culture with ECs in vitro could improve the survival and function of isolated rat islet, and co-transplantation of islets with ECs could effectively prolong the islet graft survival in diabetic rats.

Portal vein thrombosis is a potentially preventable complication in clinical islet transplantation

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Kawahara, Toshiyasu; Kin, Tatsuya; Kashkoush, Samy; Gala-Lopez, Boris; Bigam, David L.; Kneteman, Norman M.; Koh, Angela; Senior, Peter A.; Shapiro, A.M. James

2011-01-01

Percutaneous transhepatic portal access avoids surgery, but is rarely associated with bleeding or portal venous thrombosis. We herein report our large, single-center experience of percutaneous islet implantation, and evaluate risk factors of portal vein thrombosis and graft function. Prospective data was collected on 268 intraportal islet transplants (122 subjects). A portal venous Doppler ultrasound was obtained on Days 1 and 7 days posttransplant. Therapeutic heparinization, complete ablation of the portal catheter tract with Avitene paste, and limiting packed cell volume to islet transplant procedures over the past 5 years. In the previous cumulative experience, partial thrombosis did not affect islet function. Standard liver volume correlated negatively (r=−0.257, Pislet transplantation, provided therapeutic anticoagulation is maintained, and packed cell volume is limited to <5 ml. PMID:21883914

Islet Transplantation in Type 1 Diabetes: Ongoing Challenges, Refined Procedures, and Long-Term Outcome

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Shapiro, A.M. James

2012-01-01

Remarkable progress has been made in islet transplantation over a span of 40 years. Once just an experimental curiosity in mice, this therapy has moved forward, and can now provide robust therapy for highly selected patients with type 1 diabetes (T1D), refractory to stabilization by other means. This progress could not have occurred without extensive dynamic international collaboration. Currently, 1,085 patients have undergone islet transplantation at 40 international sites since the Edmonton Protocol was reported in 2000 (752 allografts, 333 autografts), according to the Collaborative Islet Transplant Registry. The long-term results of islet transplantation in selected centers now match registry data of pancreas-alone transplantation, with 6 sites reporting five-year insulin independence rates ≥50%. Islet transplantation has been criticized for the use of multiple donor pancreas organs, but progress has also occurred in single-donor success, with 10 sites reporting increased single-donor engraftment. The next wave of innovative clinical trial interventions will address instant blood-mediated inflammatory reaction (IBMIR), apoptosis, and inflammation, and will translate into further marked improvements in single-donor success. Effective control of auto- and alloimmunity is the key to long-term islet function, and high-resolution cellular and antibody-based assays will add considerable precision to this process. Advances in immunosuppression, with new antibody-based targeting of costimulatory blockade and other T-B cellular signaling, will have further profound impact on the safety record of immunotherapy. Clinical trials will move forward shortly to test out new human stem cell derived islets, and in parallel trials will move forward, testing pig islets for compatibility in patients. Induction of immunological tolerance to self-islet antigens and to allografts is a difficult challenge, but potentially within our grasp. PMID:23804275

Glucose activates prenyltransferases in pancreatic islet {beta}-cells

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Goalstone, Marc [Department of Medicine, University of Colorado, VA Medical Center, Denver, CO 80220 (United States); Kamath, Vasudeva [Department of Pharmaceutical Sciences, Wayne State University, VA Medical Center, Detroit, MI 48201 (United States); Kowluru, Anjaneyulu, E-mail: akowluru@med.wayne.edu [Department of Pharmaceutical Sciences, Wayne State University, VA Medical Center, Detroit, MI 48201 (United States)

2010-01-01

A growing body of evidence implicates small G-proteins [e.g., Cdc42 and Rac1] in glucose-stimulated insulin secretion [GSIS] in the islet {beta}-cell. These signaling proteins undergo post-translational modifications [e.g., prenylation] at their C-terminal cysteine residue and appear to be essential for the transport and fusion of insulin-containing secretory granules with the plasma membrane and the exocytotic secretion of insulin. However, potential regulation of the prenylating enzymes by physiological insulin secretogues [e.g., glucose] has not been investigated thus far. Herein, we report immunological localization, sub-cellular distribution and regulation of farnesyltransferases [FTases] and geranylgeranyltransferase [GGTase] by glucose in insulin-secreting INS 832/13 {beta}-cells and normal rat islets. Our findings suggest that an insulinotropic concentration of glucose [20 mM] markedly stimulated the expression of the {alpha}-subunits of FTase/GGTase-1, but not the {beta}-subunits of FTase or GGTase-1 without significantly affecting the predominantly cytosolic distribution of these holoenzymes in INS 832/13 cells and rodent islets. Under these conditions, glucose significantly stimulated [2.5- to 4.0-fold over basal] the activities of both FTase and GGTase-1 in both cell types. Together, these findings provide the first evidence to suggest that GSIS involves activation of the endogenous islet prenyltransferases by glucose, culminating in the activation of their respective G-protein substrates, which is necessary for cytoskeletal rearrangement, vesicular transport, fusion and secretion of insulin.

Glucose activates prenyltransferases in pancreatic islet β-cells

International Nuclear Information System (INIS)

Goalstone, Marc; Kamath, Vasudeva; Kowluru, Anjaneyulu

2010-01-01

A growing body of evidence implicates small G-proteins [e.g., Cdc42 and Rac1] in glucose-stimulated insulin secretion [GSIS] in the islet β-cell. These signaling proteins undergo post-translational modifications [e.g., prenylation] at their C-terminal cysteine residue and appear to be essential for the transport and fusion of insulin-containing secretory granules with the plasma membrane and the exocytotic secretion of insulin. However, potential regulation of the prenylating enzymes by physiological insulin secretogues [e.g., glucose] has not been investigated thus far. Herein, we report immunological localization, sub-cellular distribution and regulation of farnesyltransferases [FTases] and geranylgeranyltransferase [GGTase] by glucose in insulin-secreting INS 832/13 β-cells and normal rat islets. Our findings suggest that an insulinotropic concentration of glucose [20 mM] markedly stimulated the expression of the α-subunits of FTase/GGTase-1, but not the β-subunits of FTase or GGTase-1 without significantly affecting the predominantly cytosolic distribution of these holoenzymes in INS 832/13 cells and rodent islets. Under these conditions, glucose significantly stimulated [2.5- to 4.0-fold over basal] the activities of both FTase and GGTase-1 in both cell types. Together, these findings provide the first evidence to suggest that GSIS involves activation of the endogenous islet prenyltransferases by glucose, culminating in the activation of their respective G-protein substrates, which is necessary for cytoskeletal rearrangement, vesicular transport, fusion and secretion of insulin.

Survival of Free and Encapsulated Human and Rat Islet Xenografts Transplanted into the Mouse Bone Marrow

Science.gov (United States)

Meier, Raphael P. H.; Seebach, Jörg D.; Morel, Philippe; Mahou, Redouan; Borot, Sophie; Giovannoni, Laurianne; Parnaud, Geraldine; Montanari, Elisa; Bosco, Domenico; Wandrey, Christine; Berney, Thierry; Bühler, Leo H.; Muller, Yannick D.

2014-01-01

Bone marrow was recently proposed as an alternative and potentially immune-privileged site for pancreatic islet transplantation. The aim of the present study was to assess the survival and rejection mechanisms of free and encapsulated xenogeneic islets transplanted into the medullary cavity of the femur, or under the kidney capsule of streptozotocin-induced diabetic C57BL/6 mice. The median survival of free rat islets transplanted into the bone marrow or under the kidney capsule was 9 and 14 days, respectively, whereas that of free human islets was shorter, 7 days (bone marrow) and 10 days (kidney capsule). Infiltrating CD8+ T cells and redistributed CD4+ T cells, and macrophages were detected around the transplanted islets in bone sections. Recipient mouse splenocytes proliferated in response to donor rat stimulator cells. One month after transplantation under both kidney capsule or into bone marrow, encapsulated rat islets had induced a similar degree of fibrotic reaction and still contained insulin positive cells. In conclusion, we successfully established a small animal model for xenogeneic islet transplantation into the bone marrow. The rejection of xenogeneic islets was associated with local and systemic T cell responses and macrophage recruitment. Although there was no evidence for immune-privilege, the bone marrow may represent a feasible site for encapsulated xenogeneic islet transplantation. PMID:24625569

Current principles and practice in autologous intraportal islet transplantation: a meta-analysis of the technical considerations.

Science.gov (United States)

Kumar, Rohan; Chung, Wen Yuan; Dennison, Ashley Robert; Garcea, Giuseppe

2016-04-01

Autologous islet transplantation (IAT) following pancreatectomy is now a recognized, albeit highly specialized procedure carried out in a small number of centers worldwide. Current clinical principles and best practice with emphasis on examining the technical aspects of surgery in centers with significant IAT experience are reviewed. Literature search for studies discussing any technical aspect of pancreatectomy with intraportal IAT was included. Thirty-five papers were included; all were single-center case series. The indications, surgical approach to pancreatectomy with IAT, islet yield, static pancreas preservation prior to islet digestion, portal vein access, absolute islet infusion volumes, and portal venous pressure changes during transfusion evaluated. IAT is considered a "last resort" when alternative approaches have been exhausted. Pre-morbid histology and prior surgical drainage adversely influence islet yields and may influence the clinical decision to perform pancreatectomy and IAT. Following pancreas digestion, absolute numbers of islets recovered and smaller islet size predict rates of insulin independence following IAT. Islet volumes and portal venous pressure changes are important factors for the development of complications. Surgical access for IAT includes intra-operative, immediate or delayed infusion via an "exteriorized" vein, and radiological percutaneous approaches. Delayed infusion can be combined with pancreas preservation techniques prior to islet isolation. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Survival of free and encapsulated human and rat islet xenografts transplanted into the mouse bone marrow.

Directory of Open Access Journals (Sweden)

Raphael P H Meier

Full Text Available Bone marrow was recently proposed as an alternative and potentially immune-privileged site for pancreatic islet transplantation. The aim of the present study was to assess the survival and rejection mechanisms of free and encapsulated xenogeneic islets transplanted into the medullary cavity of the femur, or under the kidney capsule of streptozotocin-induced diabetic C57BL/6 mice. The median survival of free rat islets transplanted into the bone marrow or under the kidney capsule was 9 and 14 days, respectively, whereas that of free human islets was shorter, 7 days (bone marrow and 10 days (kidney capsule. Infiltrating CD8+ T cells and redistributed CD4+ T cells, and macrophages were detected around the transplanted islets in bone sections. Recipient mouse splenocytes proliferated in response to donor rat stimulator cells. One month after transplantation under both kidney capsule or into bone marrow, encapsulated rat islets had induced a similar degree of fibrotic reaction and still contained insulin positive cells. In conclusion, we successfully established a small animal model for xenogeneic islet transplantation into the bone marrow. The rejection of xenogeneic islets was associated with local and systemic T cell responses and macrophage recruitment. Although there was no evidence for immune-privilege, the bone marrow may represent a feasible site for encapsulated xenogeneic islet transplantation.

Preclinical evaluation of a 68Ga-labeled biotin analogue for applications in islet transplantation

International Nuclear Information System (INIS)

Eriksson, Olof; Carlsson, Fredrik; Blom, Elisabeth; Sundin, Anders; Långström, Bengt; Korsgren, Olle; Velikyan, Irina

2012-01-01

Introduction: Islet transplantation is a promising treatment for type 1 diabetes mellitus, but the fate of the cells after intraportal infusion is unclear. It is therefore imperative to develop novel techniques for noninvasive imaging and quantification of events following islet transplantation. Methods: Small islet-like microbeads, avidin-covered agarose resins (AARs), were used as a model system for islet transplantation. Capability for specific [ 68 Ga]Ga-DOTA-(PEG) 2 -biotin uptake and retention for either AARs or human islets conjugated with avidin by means of a heparin scaffold was studied in vitro. Biodistribution of the novel positron emission tomography (PET) tracer [ 68 Ga]Ga-DOTA-(PEG) 2 -biotin was evaluated in mice treated by intraportal transplantation of AARs by μPET/computed tomography and ex vivo organ distribution and compared with control mice. Results: AARs had high capability to bind [ 68 Ga]Ga-DOTA-(PEG) 2 -biotin, close to 50% of administrated tracer/μl in vitro (>0.25 MBq/μl). Avidin-tagged human islets could bind on average 2.2% of administered tracer/μl. Specificity (>90%) and retention (>90% after 1 h) were high for both AARs and avidin-tagged islets. Hepatic tracer uptake and retention were increased in mice transplanted with AARs [standardized uptake value (SUV)=2.6] compared to the untreated group (SUV=1.4). In vivo uptake of tracer to AARs was blocked by preadministration of unlabeled biotin. Conclusions: Avidin-tagged islet-like objects can be tracked in hepatic volume after intraportal transplantation by using [ 68 Ga]Ga-DOTA-(PEG) 2 -biotin and PET.

Pancreas Islet Transplantation for Patients With Type 1 Diabetes Mellitus: A Clinical Evidence Review.

Science.gov (United States)

2015-01-01

Type 1 diabetes mellitus is caused by the autoimmune destruction of pancreatic beta (β) cells, resulting in severe insulin deficiency. Islet transplantation is a β-cell replacement therapeutic option that aims to restore glycemic control in patients with type 1 diabetes. The objective of this study was to determine the clinical effectiveness of islet transplantation in patients with type 1 diabetes, with or without kidney disease. We conducted a systematic review of the literature on islet transplantation for type 1 diabetes, including relevant health technology assessments, systematic reviews, meta-analyses, and observational studies. We used a two-step process: first, we searched for systematic reviews and health technology assessments; second, we searched primary studies to update the chosen health technology assessment. The Assessment of Multiple Systematic Reviews measurement tool was used to examine the methodological quality of the systematic reviews and health technology assessments. We assessed the quality of the body of evidence and the risk of bias according to the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) Working Group criteria. Our searched yielded 1,354 citations. One health technology assessment, 11 additional observational studies to update the health technology assessment, one registry report, and four guidelines were included; the observational studies examined islet transplantation alone, islet-after-kidney transplantation, and simultaneous islet-kidney transplantation. In general, low to very low quality of evidence exists for islet transplantation in patients with type 1 diabetes with difficult-to-control blood glucose levels, with or without kidney disease, for these outcomes: health-related quality of life, secondary complications of diabetes, glycemic control, and adverse events. However, high quality of evidence exists for the specific glycemic control outcome of insulin independence compared with

A possible link between endothelial dysfunction and insulin resistance in hypertension. A LIFE substudy. Losartan Intervention For Endpoint-Reduction in Hypertension

DEFF Research Database (Denmark)

Olsen, M H; Andersen, U B; Wachtell, K

2000-01-01

We wanted to investigate whether insulin resistance and time to steady state during isoglycemic clamp were associated with endothelial dysfunction, peripheral vascular remodeling and forearm blood flow (FBF) in patients with longstanding hypertension....

Obesity Resistance Promotes Mild Contractile Dysfunction Associated with Intracellular Ca{sup 2+} Handling

Energy Technology Data Exchange (ETDEWEB)

Sá, Felipe Gonçalves dos Santos de; Lima-Leopoldo, Ana Paula; Jacobsen, Bruno Barcellos; Ferron, Artur Junio Togneri; Estevam, Wagner Muller [Centro de Educação Física e Desportos - Departamento de Desportos - Universidade Federal do Espírito Santo, Vitória, ES (Brazil); Campos, Dijon Henrique Salomé [Departamento de Clínica Médica - Faculdade de Medicina - Universidade Estadual Paulista, Botucatu, São Paulo (Brazil); Castardeli, Edson; Cunha, Márcia Regina Holanda da [Centro de Educação Física e Desportos - Departamento de Desportos - Universidade Federal do Espírito Santo, Vitória, ES (Brazil); Cicogna, Antonio Carlos [Departamento de Clínica Médica - Faculdade de Medicina - Universidade Estadual Paulista, Botucatu, São Paulo (Brazil); Leopoldo, André Soares, E-mail: andresoaresleopoldo@gmail.com [Centro de Educação Física e Desportos - Departamento de Desportos - Universidade Federal do Espírito Santo, Vitória, ES (Brazil)

2015-12-15

Diet-induced obesity is frequently used to demonstrate cardiac dysfunction. However, some rats, like humans, are susceptible to developing an obesity phenotype, whereas others are resistant to that. To evaluate the association between obesity resistance and cardiac function, and the impact of obesity resistance on calcium handling. Thirty-day-old male Wistar rats were distributed into two groups, each with 54 animals: control (C; standard diet) and obese (four palatable high-fat diets) for 15 weeks. After the experimental protocol, rats consuming the high-fat diets were classified according to the adiposity index and subdivided into obesity-prone (OP) and obesity-resistant (OR). Nutritional profile, comorbidities, and cardiac remodeling were evaluated. Cardiac function was assessed by papillary muscle evaluation at baseline and after inotropic maneuvers. The high-fat diets promoted increase in body fat and adiposity index in OP rats compared with C and OR rats. Glucose, lipid, and blood pressure profiles remained unchanged in OR rats. In addition, the total heart weight and the weight of the left and right ventricles in OR rats were lower than those in OP rats, but similar to those in C rats. Baseline cardiac muscle data were similar in all rats, but myocardial responsiveness to a post-rest contraction stimulus was compromised in OP and OR rats compared with C rats. Obesity resistance promoted specific changes in the contraction phase without changes in the relaxation phase. This mild abnormality may be related to intracellular Ca2+ handling.

The role of interventional radiology and imaging in pancreatic islet cell transplantation

International Nuclear Information System (INIS)

Dixon, S.; Tapping, C.R.; Walker, J.N.; Bratby, M.; Anthony, S.; Boardman, P.; Phillips-Hughes, J.; Uberoi, R.

2012-01-01

Pancreatic islet cell transplantation (PICT) is a novel treatment for patients with insulin-dependent diabetes who have inadequate glycaemic control or hypoglycaemic unawareness, and who suffer from the microvascular/macrovascular complications of diabetes despite aggressive medical management. Islet transplantation primarily aims to improve the quality of life for type 1 diabetic patients by achieving insulin independence, preventing hypoglycaemic episodes, and reversing hypoglycaemic unawareness. The islet cells for transplantation are extracted and purified from the pancreas of brain-stem dead, heart-beating donors. They are infused into the recipient's portal vein, where they engraft into the liver to release insulin in order to restore euglycaemia. Initial strategies using surgical access to the portal vein have been superseded by percutaneous access using interventional radiology techniques, which are relatively straightforward to perform. It is important to be vigilant during the procedure in order to prevent major complications, such as haemorrhage, which can be potentially life-threatening. In this article we review the history of islet cell transplantation, present an illustrated review of our experience with islet cell transplantation by describing the role of imaging and interventional radiology, and discuss current research into imaging techniques for monitoring graft function.

Intra-islet glucagon secretion and action in the regulation of glucose homeostasis.

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Qinghua eWang

2013-01-01

Full Text Available Glucagon, a key hormone in the regulation of glucose homeostasis, acts as a counter-regulatory hormone to insulin by promoting hepatic glucose output. Under normal conditions, insulin and glucagon operate in concert to maintain the glucose level within a narrow physiological range. In diabetes, however, while insulin secretion or action is insufficient, the production and secretion of glucagon are excessive, contributing to the development of diabetic hyperglycemia. Within an islet, intra-islet insulin, in cooperation with intra-islet GABA, suppresses glucagon secretion via direct modulation of -cell intracellular signaling pathways involving Akt activation, GABA receptor phosphorylation and the receptor plasma membrane translocation, while intra-islet glucagon plays an important role in modulating β-cell function and insulin secretion. Defects in the insulin-glucagon fine-tuning machinery may result in β-cell glucose incompetence, leading to unsuppressed glucagon secretion and subsequent hyperglycemia, which often occur under extreme conditions of glucose influx or efflux. Therefore, deciphering the precise molecular mechanisms underlying glucagon secretion and action will facilitate our understanding of glucagon physiology, in particular, its role in regulating islet β-cell function, and hence the mechanisms behind body glucose homeostasis.

Exercise Increases Insulin Content and Basal Secretion in Pancreatic Islets in Type 1 Diabetic Mice

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Han-Hung Huang

2011-01-01

Full Text Available Exercise appears to improve glycemic control for people with type 1 diabetes (T1D. However, the mechanism responsible for this improvement is unknown. We hypothesized that exercise has a direct effect on the insulin-producing islets. Eight-week-old mice were divided into four groups: sedentary diabetic, exercised diabetic, sedentary control, and exercised control. The exercised groups participated in voluntary wheel running for 6 weeks. When compared to the control groups, the islet density, islet diameter, and β-cell proportion per islet were significantly lower in both sedentary and exercised diabetic groups and these alterations were not improved with exercise. The total insulin content and insulin secretion were significantly lower in sedentary diabetics compared to controls. Exercise significantly improved insulin content and insulin secretion in islets in basal conditions. Thus, some improvements in exercise-induced glycemic control in T1D mice may be due to enhancement of insulin content and secretion in islets.

Effects of Energy Dissipation Rate on Islets of Langerhans: Implications for Isolation and Transplantation

Science.gov (United States)

Shenkman, Rustin M.; Godoy-Silva, Ruben; Papas, Klearchos K.; Chalmers, Jeffrey J.

2010-01-01

Acute physical stresses can occur in the procurement and isolation process and potentially can contribute to islet death or malfunction upon transplantation. A contractional flow device, previously used to subject suspended cells to well-defined hydrodynamic forces, has been modified and used to assess the vulnerability of porcine islets of Langerhans to hydrodynamic forces. The flow profiles and velocity gradients in this modified device were modeled using commercial CFD software and characterized, as in previous studies, with the scalar parameter, energy dissipation rate (EDR). Porcine islets were stressed in a single pass at various stress levels (i.e., values of EDR). Membrane integrity, oxygen uptake rate, caspase 3/7 activity, and insulin release were not affected by the levels of fluid stress tested up to an EDR of 2 × 103 W/m3. Visual observation of the stressed islets suggested that cells at the islet exterior were peeled away at EDR greater than 10,000 W/m3, however, this observation could not be confirmed using image analysis software, which determined the ratio of surface perimeter to total area. The result of this study suggests an upper limit in fluid stress to which islets can be subjected. Such upper limits assist in the design and operation of future islet processing equipment and processes. PMID:19191351

Effect of endurance versus resistance training on quadriceps muscle dysfunction in COPD

DEFF Research Database (Denmark)

Iepsen, Ulrik Winning; Munch, Gregers Druedal Wibe; Rugbjerg, Mette

2016-01-01

INTRODUCTION: Exercise is an important countermeasure to limb muscle dysfunction in COPD. The two major training modalities in COPD rehabilitation, endurance training (ET) and resistance training (RT), may both be efficient in improving muscle strength, exercise capacity, and health-related quality...... and after the training intervention to assess muscle morphology and metabolic and angiogenic factors. Symptom burden, exercise capacity (6-minute walking and cycle ergometer tests), and vascular function were also assessed. RESULTS: Both training modalities improved symptom burden and exercise capacity...... with no difference between the two groups. The mean (SD) proportion of glycolytic type IIa muscle fibers was reduced after ET (from 48% [SD 11] to 42% [SD 10], Ptraining modality on muscle...

Successful suppression of the early rejection of pig islets in monkeys

NARCIS (Netherlands)

Rijkelijkhuizen, JKRA; Bouwman, E; van der Burg, MPM; Ringers, J; Ossevoort, MA; Kuhn, EM; Frost, P; Jonker, M

2000-01-01

Primary nonfunction (PNF) is seen very frequently after xenogeneic transplantation of islets of Langerhans. In a pig-to-rat model we recently observed that no PNF occurs when the islets are kept in culture at 37 degreesC fur 1-2 weeks prior to transplantation. In order to investigate the rejection

Mesenchymal stem cell and derived exosome as small RNA carrier and Immunomodulator to improve islet transplantation.

Science.gov (United States)

Wen, Di; Peng, Yang; Liu, Di; Weizmann, Yossi; Mahato, Ram I

2016-09-28

Human bone marrow mesenchymal stem cells (hBMSCs) and their exosomes can suppress immune reaction and deliver small RNAs. Thus, they may improve islet transplantation by delivering small RNAs for promoting islet function and inhibiting immune rejection. Here, we proposed an hBMSC and its exosome-based therapy to overcome immune rejection and poor islet function, both of which hinder the success of islet transplantation. We found overexpressed siFas and anti-miR-375 in plasmid encoding shFas and anti-miR-375 transfected hBMSC-derived exosomes, which silenced Fas and miR-375 of human islets and improved their viability and function against inflammatory cytokines. This plasmid transfected hBMSCs downregulated Fas and miR-375 of human islets in a humanized NOD scid gamma (NSG) mouse model, whose immune reaction was inhibited by injecting hBMSC and peripheral blood mononuclear cell (PBMC) co-cultured exosomes. These exosomes suppressed immune reaction by inhibiting PBMC proliferation and enhancing regulatory T cell (Treg) function. Collectively, our studies elucidated the mechanisms of RNA delivery from hBMSCs to human islets and the immunosuppressive effect of hBMSC and peripheral blood mononuclear cell co-cultured exosomes for improving islet transplantation. Copyright © 2016 Elsevier B.V. All rights reserved.

Feasibility of baculovirus-mediated reporter gene delivery for efficient monitoring of islet transplantation in vivo

International Nuclear Information System (INIS)

Liu, Shuai; Pan, Yu; Lv, Jing; Wu, Haifei; Tian, Jingyan; Zhang, Yifan

2014-01-01

Objective: The objective of this study was to explore the feasibility of baculovirus vector-mediated sodium iodide symporter (NIS) gene delivery to monitor islet transplantation. Methods: Baculovirus vectors expressing green fluorescent protein (GFP) or NIS (Bac-GFP and Bac-NIS) were established using the Bac-to-Bac baculovirus expression system. The GFP expression of Bac-GFP-infected rat islets was observed in vitro by fluorescence microscopy. Iodine uptake and inhibition of iodine uptake by NaClO 4 in Bac-NIS-infected islets were dynamically monitored in vitro. Bac-GFP- or Bac-NIS-infected islets were implanted into the left axillary cavity of NOD-SCID mice, and fluorescence imaging and 125 I NanoSPECT/CT imaging were subsequently performed in vivo. Results: Bac-GFP efficiently infected rat islets (over 95% infected at MOI = 40), and the expression of GFP lasted approximately two weeks. NaClO 4 could inhibit iodine uptake by Bac-NIS-infected islets. In vivo imaging revealed that the fluorescence intensity of the transplant sites in Bac-GFP-infected groups was significantly higher than in the non-infected group. Grafts could be clearly observed by 125 I NanoSPECT/CT imaging for up to 8 h. Conclusion: Baculovirus vectors are powerful vehicles for studying rat islets in gene delivery. It is feasible to use a baculovirus vector to delivery an NIS gene for non-invasive monitoring transplanted islets in vivo by the expression of the target gene

Decrease of glucose-induced insulin secretion of rat pancreatic islets after irradiation in vitro

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Heinzmann, D; Nadrowitz, R; Besch, W; Schmidt, W; Hahn, H J [Zentralinstitut fuer Diabetes, Karlsburg (German Democratic Republic); Ernst-Moritz-Arndt-Universitaet, Greifswald (German Democratic Republic). Radiologische Klinik)

1983-01-01

In vitro irradiation of rat pancreatic islets up to a dose of 2.5 Gy did neither alter glucose- nor isobutylmethyl xanthine (IBMX)-induced insulin secretion. Insulin as well as glucagon content of irradiated islets corresponded to that of the control tissue. So it was in islets irradiated with 25 Gy which were characterized by a decreased insulin secretion in the presence of glucose and IBMX, respectively. There was no indication of an enhanced hormone output in the radiation medium and it is to be suggested that higher radiation doses affect the insulin release of pancreatic islets in vitro. This must be taken into consideration for radioimmunosuppression experiments.

Identifying Effective Enzyme Activity Targets for Recombinant Class I and Class II Collagenase for Successful Human Islet Isolation.

Science.gov (United States)

Balamurugan, Appakalai N; Green, Michael L; Breite, Andrew G; Loganathan, Gopalakrishnan; Wilhelm, Joshua J; Tweed, Benjamin; Vargova, Lenka; Lockridge, Amber; Kuriti, Manikya; Hughes, Michael G; Williams, Stuart K; Hering, Bernhard J; Dwulet, Francis E; McCarthy, Robert C

2016-01-01

Isolation following a good manufacturing practice-compliant, human islet product requires development of a robust islet isolation procedure where effective limits of key reagents are known. The enzymes used for islet isolation are critical but little is known about the doses of class I and class II collagenase required for successful islet isolation. We used a factorial approach to evaluate the effect of high and low target activities of recombinant class I (rC1) and class II (rC2) collagenase on human islet yield. Consequently, 4 different enzyme formulations with divergent C1:C2 collagenase mass ratios were assessed, each supplemented with the same dose of neutral protease. Both split pancreas and whole pancreas models were used to test enzyme targets (n = 20). Islet yield/g pancreas was compared with historical enzymes (n = 42). Varying the Wunsch (rC2) and collagen degradation activity (CDA, rC1) target dose, and consequently the C1:C2 mass ratio, had no significant effect on tissue digestion. Digestions using higher doses of Wunsch and CDA resulted in comparable islet yields to those obtained with 60% and 50% of those activities, respectively. Factorial analysis revealed no significant main effect of Wunsch activity or CDA for any parameter measured. Aggregate results from 4 different collagenase formulations gave 44% higher islet yield (>5000 islet equivalents/g) in the body/tail of the pancreas (n = 12) when compared with those from the same segment using a standard natural collagenase/protease mixture (n = 6). Additionally, islet yields greater than 5000 islet equivalents/g pancreas were also obtained in whole human pancreas. A broader C1:C2 ratio can be used for human islet isolation than has been used in the past. Recombinant collagenase is an effective replacement for the natural enzyme and we have determined that high islet yield can be obtained even with low doses of rC1:rC2, which is beneficial for the survival of islets.

Hair Follicle Dermal Sheath Derived Cells Improve Islet Allograft Survival without Systemic Immunosuppression

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Xiaojie Wang

2015-01-01

Full Text Available Immunosuppressive drugs successfully prevent rejection of islet allografts in the treatment of type I diabetes. However, the drugs also suppress systemic immunity increasing the risk of opportunistic infection and cancer development in allograft recipients. In this study, we investigated a new treatment for autoimmune diabetes using naturally immune privileged, hair follicle derived, autologous cells to provide localized immune protection of islet allotransplants. Islets from Balb/c mouse donors were cotransplanted with syngeneic hair follicle dermal sheath cup cells (DSCC, group 1 or fibroblasts (FB, group 2 under the kidney capsule of immune-competent, streptozotocin induced, diabetic C57BL/6 recipients. Group 1 allografts survived significantly longer than group 2 (32.2 ± 12.2 versus 14.1 ± 3.3 days, P<0.001 without administration of any systemic immunosuppressive agents. DSCC reduced T cell activation in the renal lymph node, prevented graft infiltrates, modulated inflammatory chemokine and cytokine profiles, and preserved better beta cell function in the islet allografts, but no systemic immunosuppression was observed. In summary, DSCC prolong islet allograft survival without systemic immunosuppression by local modulation of alloimmune responses, enhancing of beta cell survival, and promoting of graft revascularization. This novel finding demonstrates the capacity of easily accessible hair follicle cells to be used as local immunosuppression agents in islet transplantation.

Posttransplant Lymphoproliferative Disorder After Clinical Islet Transplantation: Report of the First Two Cases.

Science.gov (United States)

Peters, A; Olateju, T; Deschenes, J; Shankarnarayan, S H; Chua, N; Shapiro, A M J; Senior, P

2017-09-01

We report the first two cases of posttransplant lymphoproliferative disorder (PTLD) in recipients of islet transplants worldwide. First, a 44-year-old recipient of three islet infusions developed PTLD 80 months after his initial transplantation, presenting with abdominal pain and diffuse terminal ileum thickening on imaging. He was treated with surgical excision, reduction of immunosuppression, and rituximab. Seven months later, he developed central nervous system PTLD, presenting with vertigo and diplopia; immunosuppression was discontinued, resulting in graft loss, and he was given high-dose methotrexate and underwent consolidative autologous stem cell transplantation. He remains in remission 37 months after the initial diagnosis. Second, a 58-year-old female recipient of two islet infusions developed PTLD 24 months after initial islet infusion, presenting with pancytopenia secondary to extensive bone marrow involvement. Immunosuppression was discontinued, resulting in graft loss, and she received rituximab and chemotherapy, achieving complete remission. Both patients were monomorphic B cell PTLD subtype by histology and negative for Epstein-Barr virus in tissue or blood. These cases document the first occurrences of this rare complication in islet transplantation, likely secondary to prolonged, intensive immunosuppression, and highlight the varying clinical manifestations of PTLD. Further studies are needed to determine incidence rate and risk factors in islet transplantation. © 2017 The American Society of Transplantation and the American Society of Transplant Surgeons.

Dissociation between insulin secretion and DNA synthesis in cultured pancreatic islets

DEFF Research Database (Denmark)

Nielsen, Jens Høiriis

1985-01-01

-Tdr incorporation. However, long-term exposure to IBMX did not result in increased DNA content of the islets. Inhibition of the DNA synthesis by 5 mM hydroxyurea resulted in a marked reduction in DNA content of the islets but no decrease in either insulin release or insulin content when expressed per ng DNA...

Supplements in human islet culture: human serum albumin is inferior to fetal bovine serum.

Science.gov (United States)

Avgoustiniatos, Efstathios S; Scott, William E; Suszynski, Thomas M; Schuurman, Henk-Jan; Nelson, Rebecca A; Rozak, Phillip R; Mueller, Kate R; Balamurugan, A N; Ansite, Jeffrey D; Fraga, Daniel W; Friberg, Andrew S; Wildey, Gina M; Tanaka, Tomohiro; Lyons, Connor A; Sutherland, David E R; Hering, Bernhard J; Papas, Klearchos K

2012-01-01

Culture of human islets before clinical transplantation or distribution for research purposes is standard practice. At the time the Edmonton protocol was introduced, clinical islet manufacturing did not include culture, and human serum albumin (HSA), instead of fetal bovine serum (FBS), was used during other steps of the process to avoid the introduction of xenogeneic material. When culture was subsequently introduced, HSA was also used for medium supplementation instead of FBS, which was typically used for research islet culture. The use of HSA as culture supplement was not evaluated before this implementation. We performed a retrospective analysis of 103 high-purity islet preparations (76 research preparations, all with FBS culture supplementation, and 27 clinical preparations, all with HSA supplementation) for oxygen consumption rate per DNA content (OCR/DNA; a measure of viability) and diabetes reversal rate in diabetic nude mice (a measure of potency). After 2-day culture, research preparations exhibited an average OCR/DNA 51% higher (p < 0.001) and an average diabetes reversal rate 54% higher (p < 0.05) than clinical preparations, despite 87% of the research islet preparations having been derived from research-grade pancreata that are considered of lower quality. In a prospective paired study on islets from eight research preparations, OCR/DNA was, on average, 27% higher with FBS supplementation than that with HSA supplementation (p < 0.05). We conclude that the quality of clinical islet preparations can be improved when culture is performed in media supplemented with serum instead of albumin.

A Stirred Microchamber for Oxygen Consumption Rate Measurements With Pancreatic Islets

Science.gov (United States)

Papas, Klearchos K.; Pisania, Anna; Wu, Haiyan; Weir, Gordon C.; Colton, Clark K.

2010-01-01

Improvements in pancreatic islet transplantation for treatment of diabetes are hindered by the absence of meaningful islet quality assessment methods. Oxygen consumption rate (OCR) has previously been used to assess the quality of organs and primary tissue for transplantation. In this study, we describe and characterize a stirred microchamber for measuring OCR with small quantities of islets. The device has a titanium body with a chamber volume of about 200 µL and is magnetically stirred and water jacketed for temperature control. Oxygen partial pressure (pO2) is measured by fluorescence quenching with a fiber optic probe, and OCR is determined from the linear decrease of pO2 with time. We demonstrate that measurements can be made rapidly and with high precision. Measurements with βTC3 cells and islets show that OCR is directly proportional to the number of viable cells in mixtures of live and dead cells and correlate linearly with membrane integrity measurements made with cells that have been cultured for 24 h under various stressful conditions. PMID:17497731

Transplantation of islet allografts and xenografts in totally pancreatectomized diabetic dogs using the hybrid artificial pancreas.

Science.gov (United States)

Monaco, A P; Maki, T; Ozato, H; Carretta, M; Sullivan, S J; Borland, K M; Mahoney, M D; Chick, W L; Muller, T E; Wolfrum, J

1991-01-01

Previously the authors reported on a Hybrid Artificial Pancreas device that maintained patent vascular anastomoses in normal dogs and, when seeded with allogeneic canine islets, maintained normal fasting blood sugars (FBS) in diabetic pancreatectomized dogs. Eventual failure of these devices was believed to be related to loss of islet viability and/or insufficient islet mass. The current study evaluates the effect of increased islet mass produced by implantation of two islet-seeded devices in pancreatectomized dogs and compares the results with those from dogs that received a single device. Twelve of fifteen dogs receiving single devices showed initial function as determined by elimination or reduction of exogenous insulin requirement; four showed initial function and seven showed extended function (100 to 284 days). Excessive weight loss (more than 20%), despite normal FBS and insulin dependence, required that four animals in this latter group be killed. Devices seeded with xenogeneic islets have met with limited success. One dog that received two bovine islet-seeded devices achieved function for more than 100 days; the remaining bovine-seeded devices (n = 8) functioned for only 3 to 16 days. Porcine islet-seeded devices were assessed by intravenous glucose tolerance tests (IVGTT). Recipients of two devices seeded with allogeneic islets demonstrated improved IVGTT results when compared to those from pancreatectomized dogs and recipients of single devices but were abnormal when compared to intact animals. Histologic examination of device and autopsy material from all failed experiments was performed and showed no mononuclear cell infiltration of the islet chamber or vascular graft material, only a few incidence of device thrombosis, and varying degrees of islet viability as judged by morphologic and immunohistochemical evaluation. The authors believe they have demonstrated progress toward the development and clinical applicability of the Hybrid Artificial Pancreas

A rapid, efficient, and economic device and method for the isolation and purification of mouse islet cells.

Directory of Open Access Journals (Sweden)

Yin Zongyi

Full Text Available Rapid, efficient, and economic method for the isolation and purification of islets has been pursued by numerous islet-related researchers. In this study, we compared the advantages and disadvantages of our developed patented method with those of commonly used conventional methods (Ficoll-400, 1077, and handpicking methods. Cell viability was assayed using Trypan blue, cell purity and yield were assayed using diphenylthiocarbazone, and islet function was assayed using acridine orange/ethidium bromide staining and enzyme-linked immunosorbent assay-glucose stimulation testing 4 days after cultivation. The results showed that our islet isolation and purification method required 12 ± 3 min, which was significantly shorter than the time required in Ficoll-400, 1077, and HPU groups (34 ± 3, 41 ± 4, and 30 ± 4 min, respectively; P 1000 islets. In summary, the MCT method is a rapid, efficient, and economic method for isolating and purifying murine islet cell clumps. This method overcomes some of the shortcomings of conventional methods, showing a relatively higher quality and yield of islets within a shorter duration at a lower cost. Therefore, the current method provides researchers with an alternative option for islet isolation and should be widely generalized.

Long-term inhibition of dipeptidyl peptidase IV improves glucose tolerance and preserves islet function in mice

DEFF Research Database (Denmark)

Reimer, M Kvist; Holst, Jens Juul; Ahrén, B

2002-01-01

DPPIV activity to 0.01+/-0.03 mU/ml vs 3.26+/-0.19 mU/ml in controls (PGlucose tolerance after gastric glucose gavage, as judged by the area under the curve for plasma glucose levels over the 120 min study period, was increased after 8 weeks by NVP DPP728 in mice fed normal diet (P=0.......029) and in mice fed a high-fat diet (P=0.036). This was accompanied by increased plasma levels of insulin and intact GLP-1. Glucose-stimulated insulin secretion from islets isolated from NVP DPP728-treated animals after 8 weeks of treatment was increased as compared with islets from control animals at 5.6, 8...... and such mice rendered glucose-intolerant and insulin-resistant by feeding a high-fat diet. DESIGN: In mice fed a standard diet (11% fat) or a high-fat diet (58% fat), NVP DPP728 (0.12 micromol/g body weight) was administered in the drinking water for an 8 week period. RESULTS: DPPIV inhibition reduced plasma...

Asymmetric dimethylarginine (ADMA) elevation and arginase up-regulation contribute to endothelial dysfunction related to insulin resistance in rats and morbidly obese humans.

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El Assar, Mariam; Angulo, Javier; Santos-Ruiz, Marta; Ruiz de Adana, Juan Carlos; Pindado, María Luz; Sánchez-Ferrer, Alberto; Hernández, Alberto; Rodríguez-Mañas, Leocadio

2016-06-01

The presence of insulin resistance (IR) is determinant for endothelial dysfunction associated with obesity. Although recent studies have implicated the involvement of mitochondrial superoxide and inflammation in the defective nitric oxide (NO)-mediated responses and subsequent endothelial dysfunction in IR, other mechanisms could compromise this pathway. In the present study, we assessed the role of asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of endothelium-dependent vasodilatation in human morbid obesity and in a non-obese rat model of IR. We show that both increased ADMA and up-regulated arginase are determinant factors in the alteration of the l-arginine/NO pathway associated with IR in both models and also that acute treatment of arteries with arginase inhibitor or with l-arginine significantly alleviate endothelial dysfunction. These results help to expand our knowledge regarding the mechanisms of endothelial dysfunction that are related to obesity and IR and establish potential therapeutic targets for intervention. Insulin resistance (IR) is determinant for endothelial dysfunction in human obesity. Although we have previously reported the involvement of mitochondrial superoxide and inflammation, other mechanisms could compromise NO-mediated responses in IR. We evaluated the role of the endogenous NOS inhibitor asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of l-arginine/NO-mediated vasodilatation in human morbid obesity and in a non-obese rat model of IR. Bradykinin-induced vasodilatation was evaluated in microarteries derived from insulin-resistant morbidly obese (IR-MO) and non-insulin-resistant MO (NIR-MO) subjects. Defective endothelial vasodilatation in IR-MO was improved by l-arginine supplementation. Increased levels of ADMA were detected in serum and adipose tissue from IR-MO. Serum ADMA positively correlated with IR score and negatively with pD2 for bradykinin. Gene

Asymmetric dimethylarginine (ADMA) elevation and arginase up‐regulation contribute to endothelial dysfunction related to insulin resistance in rats and morbidly obese humans

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El Assar, Mariam; Angulo, Javier; Santos‐Ruiz, Marta; Ruiz de Adana, Juan Carlos; Pindado, María Luz; Sánchez‐Ferrer, Alberto; Hernández, Alberto

2016-01-01

Key points The presence of insulin resistance (IR) is determinant for endothelial dysfunction associated with obesity.Although recent studies have implicated the involvement of mitochondrial superoxide and inflammation in the defective nitric oxide (NO)‐mediated responses and subsequent endothelial dysfunction in IR, other mechanisms could compromise this pathway.In the present study, we assessed the role of asymmetric dimethylarginine (ADMA) and arginase with respect to IR‐induced impairment of endothelium‐dependent vasodilatation in human morbid obesity and in a non‐obese rat model of IR.We show that both increased ADMA and up‐regulated arginase are determinant factors in the alteration of the l‐arginine/NO pathway associated with IR in both models and also that acute treatment of arteries with arginase inhibitor or with l‐arginine significantly alleviate endothelial dysfunction.These results help to expand our knowledge regarding the mechanisms of endothelial dysfunction that are related to obesity and IR and establish potential therapeutic targets for intervention. Abstract Insulin resistance (IR) is determinant for endothelial dysfunction in human obesity. Although we have previously reported the involvement of mitochondrial superoxide and inflammation, other mechanisms could compromise NO‐mediated responses in IR. We evaluated the role of the endogenous NOS inhibitor asymmetric dimethylarginine (ADMA) and arginase with respect to IR‐induced impairment of l‐arginine/NO‐mediated vasodilatation in human morbid obesity and in a non‐obese rat model of IR. Bradykinin‐induced vasodilatation was evaluated in microarteries derived from insulin‐resistant morbidly obese (IR‐MO) and non‐insulin‐resistant MO (NIR‐MO) subjects. Defective endothelial vasodilatation in IR‐MO was improved by l‐arginine supplementation. Increased levels of ADMA were detected in serum and adipose tissue from IR‐MO. Serum ADMA positively correlated with

Pediatric pancreas transplantation, including total pancreatectomy with islet autotransplantation.

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Bondoc, Alexander J; Abu-El-Haija, Maisam; Nathan, Jaimie D

2017-08-01

Unlike other solid-organ transplants, whole pancreas transplantation in children is relatively rare, and it occurs more frequently in the context of multivisceral or composite organ transplantation. Because children only infrequently suffer severe sequelae of type 1 diabetes mellitus, pancreas transplantation is rarely indicated in the pediatric population. More commonly, pediatric pancreas transplant occurs in the setting of incapacitating acute recurrent or chronic pancreatitis, specifically islet autotransplantation after total pancreatectomy. In this clinical scenario, total pancreatectomy removes the nidus of chronic pain and debilitation, while autologous islet transplantation aims to preserve endocrine function. The published experiences with pediatric total pancreatectomy with islet autotransplantation (TPIAT) in children has demonstrated excellent outcomes including liberation from chronic opioid use, as well as improved mental and physical quality of life with good glycemic control. Given the complexity of the operation, risk of postoperative complication, and long-term physiologic changes, appropriate patient selection and comprehensive multidisciplinary care teams are critical to ensuring optimal outcomes. Copyright © 2017 Elsevier Inc. All rights reserved.

The fractal spatial distribution of pancreatic islets in three dimensions: a self-avoiding growth model

International Nuclear Information System (INIS)

Jo, Junghyo; Periwal, Vipul; Hörnblad, Andreas; Ahlgren, Ulf; Kilimnik, German; Hara, Manami

2013-01-01

The islets of Langerhans, responsible for controlling blood glucose levels, are dispersed within the pancreas. A universal power law governing the fractal spatial distribution of islets in two-dimensional pancreatic sections has been reported. However, the fractal geometry in the actual three-dimensional pancreas volume, and the developmental process that gives rise to such a self-similar structure, has not been investigated. Here, we examined the three-dimensional spatial distribution of islets in intact mouse pancreata using optical projection tomography and found a power law with a fractal dimension of 2.1. Furthermore, based on two-dimensional pancreatic sections of human autopsies, we found that the distribution of human islets also follows a universal power law with a fractal dimension of 1.5 in adult pancreata, which agrees with the value previously reported in smaller mammalian pancreas sections. Finally, we developed a self-avoiding growth model for the development of the islet distribution and found that the fractal nature of the spatial islet distribution may be associated with the self-avoidance in the branching process of vascularization in the pancreas. (paper)

PEGylated bilirubin nanoparticle as an anti-oxidative and anti-inflammatory demulcent in pancreatic islet xenotransplantation.

Science.gov (United States)

Kim, Min Jun; Lee, Yonghyun; Jon, Sangyong; Lee, Dong Yun

2017-07-01

Transplanted islets suffer hypoxic stress, which leads to nonspecific inflammation. This is the major cause of islet graft failure during the early stage of intrahepatic islet transplantation. Although bilirubin has shown potent anti-oxidative and anti-inflammatory functions, its clinical applications have been limited due to its insolubility and short half-life. To overcome this problem, novel amphiphilic bilirubin nanoparticles are designed. Hydrophilic poly(ethylene glycol) (PEG) is conjugated to the hydrophobic bilirubin molecule. Then, the PEG-bilirubin conjugates form nanoparticles via self-assembly, i.e., so-called to BRNPs. BRNPs can protect islet cells not only from chemically induced oxidative stress by scavenging reactive oxygen species molecules, but also from activated macrophages by suppressing cytokine release. Importantly, in vivo experiments demonstrate that BRNP treatment can dramatically and significantly prolong islet graft survival compared to bilirubin treatment. In addition, immunohistochemical analysis shows BRNPs have potent anti-oxidative and anti-inflammatory capabilities. Collectively, novel BRNPs can be a new potent remedy for successful islet transplantation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Wave forcing and morphological changes of New Caledonia lagoon islets: Insights on their possible relations

Science.gov (United States)

Aucan, Jérôme; Vendé-Leclerc, Myriam; Dumas, Pascal; Bricquir, Marianne

2017-10-01

In the present study, we examine how waves may contribute to the morphological changes of islets in the New Caledonia lagoon. We collected in situ wave data to investigate their characteristics. Three types of waves are identified and quantified: (1) high-frequency waves generated within the lagoon, (2) low-frequency waves originating from swells in the Tasman Sea, and (3) infragravity waves. We found out that high-frequency waves are the dominant forcing on the islets during typical wind events throughout the year, while infragravity waves, likely generated by the breaking of low-frequency waves, dominate during seasonal swell events. During swell events, low-frequency waves can also directly propagate to the islets through channels across the barrier reef, or be tidally modulated across the barrier reef before reaching the islets. Topographic surveys and beach profiles on one islet indicate areas with seasonal morphological changes and other areas with longer, interannual or decadal, erosion patterns. Although more data are needed to validate this hypothesis, we suspect that a relation exists between wave forcing and morphological changes of the islets.

Repurposing Lesogaberan to Promote Human Islet Cell Survival and β-Cell Replication

Directory of Open Access Journals (Sweden)

Jide Tian

2017-01-01

Full Text Available The activation of β-cell’s A- and B-type gamma-aminobutyric acid receptors (GABAA-Rs and GABAB-Rs can promote their survival and replication, and the activation of α-cell GABAA-Rs promotes their conversion into β-cells. However, GABA and the most clinically applicable GABA-R ligands may be suboptimal for the long-term treatment of diabetes due to their pharmacological properties or potential side-effects on the central nervous system (CNS. Lesogaberan (AZD3355 is a peripherally restricted high-affinity GABAB-R-specific agonist, originally developed for the treatment of gastroesophageal reflux disease (GERD that appears to be safe for human use. This study tested the hypothesis that lesogaberan could be repurposed to promote human islet cell survival and β-cell replication. Treatment with lesogaberan significantly enhanced replication of human islet cells in vitro, which was abrogated by a GABAB-R antagonist. Immunohistochemical analysis of human islets that were grafted into immune-deficient mice revealed that oral treatment with lesogaberan promoted human β-cell replication and islet cell survival in vivo as effectively as GABA (which activates both GABAA-Rs and GABAB-Rs, perhaps because of its more favorable pharmacokinetics. Lesogaberan may be a promising drug candidate for clinical studies of diabetes intervention and islet transplantation.

Delta-like Ligand-4-Notch Signaling Inhibition Regulates Pancreatic Islet Function and Insulin Secretion

Directory of Open Access Journals (Sweden)

Fabienne Billiard

2018-01-01

Full Text Available Although Notch signaling has been proposed as a therapeutic target for type-2 diabetes, liver steatosis, and atherosclerosis, its direct effect on pancreatic islets remains unknown. Here, we demonstrated a function of Dll4-Notch signaling inhibition on the biology of insulin-producing cells. We confirmed enhanced expression of key Notch signaling genes in purified pancreatic islets from diabetic NOD mice and showed that treatment with anti-Dll4 antibody specifically abolished Notch signaling pathway activation. Furthermore, we showed that Notch inhibition could drive proliferation of β-islet cells and confer protection from the development of STZ-induced diabetes. Importantly, inhibition of the Dll4 pathway in WT mice increased insulin secretion by inducing the differentiation of pancreatic β-islet cell progenitors, as well as the proliferation of insulin-secreting cells. These findings reveal a direct effect of Dll4-blockade on pancreatic islets that, in conjunction with its immunomodulatory effects, could be used for unmet medical needs hallmarked by inefficient insulin action.

Identifying Effective Enzyme Activity Targets for Recombinant Class I and Class II Collagenase for Successful Human Islet Isolation

OpenAIRE

Balamurugan, Appakalai N.; Green, Michael L.; Breite, Andrew G.; Loganathan, Gopalakrishnan; Wilhelm, Joshua J.; Tweed, Benjamin; Vargova, Lenka; Lockridge, Amber; Kuriti, Manikya; Hughes, Michael G.; Williams, Stuart K.; Hering, Bernhard J.; Dwulet, Francis E.; McCarthy, Robert C.

2015-01-01

Isolation following a good manufacturing practice-compliant, human islet product requires development of a robust islet isolation procedure where effective limits of key reagents are known. The enzymes used for islet isolation are critical but little is known about the doses of class I and class II collagenase required for successful islet isolation.

Engraftment Site and Effectiveness of the Pan-Caspase Inhibitor F573 to Improve Engraftment in Mouse and Human Islet Transplantation in Mice.

Science.gov (United States)

Pepper, Andrew R; Bruni, Antonio; Pawlick, Rena; Wink, John; Rafiei, Yasmin; Gala-Lopez, Boris; Bral, Mariusz; Abualhassan, Nasser; Kin, Tatsuya; Shapiro, A M James

2017-10-01

Islet transplantation is an effective therapy in type 1 diabetes and recalcitrant hypoglycemia. However, there is an ongoing need to circumvent islet loss posttransplant. We explore herein the potential of the pan-caspase inhibitor F573 to mitigate early apoptosis-mediated islet death within portal and extrahepatic portal sites in mice. Mouse or human islets were cultured in standard media ±100 μM F573 and subsequently assessed for viability and apoptosis via terminal deoxynucleotidyl transferase dUTP nick end labeling staining and caspase-3 activation. Diabetic mice were transplanted with syngeneic islets placed under the kidney capsule (KC) or into the subcutaneous deviceless (DL) site at a marginal islet dose (150 islets), or into the portal vein (PV) at a full dose (500 islets). Human islets were transplanted under the KC of diabetic immunodeficient mice at a marginal dose (500 islet equivalents). Islets were cultured in the presence of F573, and F573 was administered subcutaneously on days 0 to 5 posttransplant. Control mice were transplanted with nontreated islets and were injected with saline. Graft function was measured by nonfasting blood glucose and glucose tolerance testing. F573 markedly reduced human and mouse islet apoptosis after in vitro culture (P islet function when transplanted under the KC (P islet marginal KC transplants. Conversely, F573 significantly improved mouse islet engraftment in the PV and DL site (P islet apoptosis and improves engraftment most effectively in the portal and DL subcutaneous sites.

Insulin-Like growth factor-II (IGF-II) prevents proinflammatory cytokine-induced apoptosis and significantly improves islet survival after transplantation.

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Hughes, Amy; Mohanasundaram, Daisy; Kireta, Svjetlana; Jessup, Claire F; Drogemuller, Chris J; Coates, P Toby H

2013-03-15

The early loss of functional islet mass (50-70%) due to apoptosis after clinical transplantation contributes to islet allograft failure. Insulin-like growth factor (IGF)-II is an antiapoptotic protein that is highly expressed in β-cells during development but rapidly decreases in postnatal life. We used an adenoviral (Ad) vector to overexpress IGF-II in isolated rat islets and investigated its antiapoptotic action against exogenous cytokines interleukin-1β- and interferon-γ-induced islet cell death in vitro. Using an immunocompromised marginal mass islet transplant model, the ability of Ad-IGF-II-transduced rat islets to restore euglycemia in nonobese diabetic/severe combined immunodeficient diabetic recipients was assessed. Ad-IGF-II transduction did not affect islet viability or function. Ad-IGF-II cytokine-treated islets exhibited decreased cell death (40% ± 2.8%) versus Ad-GFP and untransduced control islets (63.2% ± 2.5% and 53.6% ± 2.3%, respectively). Ad-IGF-II overexpression during cytokine treatment resulted in a marked reduction in terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive apoptotic cells (8.3% ± 1.4%) versus Ad-GFP control (41% ± 4.2%) and untransduced control islets (46.5% ± 6.2%). Western blot analysis confirmed that IGF-II inhibits apoptosis via activation of the phosphatidylinositol 3-kinase/Akt signaling pathway. Transplantation of IGF-II overexpressing islets under the kidney capsule of diabetic mice restored euglycemia in 77.8% of recipients compared with 18.2% and 47.5% of Ad-GFP and untransduced control islet recipients, respectively (Pislet transplant outcomes in vivo. Antiapoptotic gene transfer is a potentially powerful tool to improve islet survival after transplantation.

Increased androgen levels in rats impair glucose-stimulated insulin secretion through disruption of pancreatic beta cell mitochondrial function.

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Wang, Hongdong; Wang, Xiaping; Zhu, Yunxia; Chen, Fang; Sun, Yujie; Han, Xiao

2015-11-01

Although insulin resistance is recognized to contribute to the reproductive and metabolic phenotypes of polycystic ovary syndrome (PCOS), pancreatic beta cell dysfunction plays an essential role in the progression from PCOS to the development of type 2 diabetes. However, the role of insulin secretory abnormalities in PCOS has received little attention. In addition, the precise changes in beta cells and the underlying mechanisms remain unclear. In this study, we therefore attempted to elucidate potential mechanisms involved in beta cell alterations in a rat model of PCOS. Glucose-induced insulin secretion was measured in islets isolated from DHT-treated and control rats. Oxygen consumption rate (OCR), ATP production, and mitochondrial copy number were assayed to evaluate mitochondrial function. Glucose-stimulated insulin secretion is significantly decreased in islets from DHT-treated rats. On the other hand, significant reductions are observed in the expression levels of several key genes involved in mitochondrial biogenesis and in mitochondrial OCR and ATP production in DHT-treated rat islets. Meanwhile, we found that androgens can directly impair beta cell function by inducing mitochondrial dysfunction in vitro in an androgen receptor dependent manner. For the first time, our study demonstrates that increased androgens in female rats can impair glucose-stimulated insulin secretion partly through disruption of pancreatic beta cell mitochondrial function. This work has significance for hyperandrogenic women with PCOS: excess activation of the androgen receptor by androgens may provoke beta cell dysfunction via mitochondrial dysfunction. Copyright © 2015 Elsevier Ltd. All rights reserved.

Decrease of glucose-induced insulin secretion of pancreatic rat islets after irradiation in vitro

Energy Technology Data Exchange (ETDEWEB)

Heinzmann, D; Nadrowitz, R; Besch, W; Schmidt, W; Hahn, H J

1983-01-01

Irradiation of pancreatic rat islets up to a dose of 2.5 Gy did neither alter glucose-nor IBMX-induced insulin secretion studied in vitro. The insulin as well as glucagon content of irradiated islets were similar as in the control tissue. This was also true in islets irradiated with 25 Gy which were characterized by a decreased insulin secretion in the presence of glucose and IBMX, respectively. Since we did not find indications of an enhanced hormone output in the radiation medium, we want to suggest that higher irradiation doses affect insulin release of pancreatic islets in vitro. This observation has to be taken into account for application of radioimmunosuppression for transplantation.

Ex vivo expanded human regulatory T cells delay islet allograft rejection via inhibiting islet-derived monocyte chemoattractant protein-1 production in CD34+ stem cells-reconstituted NOD-scid IL2rγnull mice.

Science.gov (United States)

Xiao, Fang; Ma, Liang; Zhao, Min; Huang, Guocai; Mirenda, Vincenzo; Dorling, Anthony; Lechler, Robert; Lombardi, Giovanna

2014-01-01

Type 1 diabetes mellitus (T1DM) is an autoimmune disease caused by immune-mediated destruction of insulin-secreting β cells of the pancreas. Near complete dependence on exogenous insulin makes T1DM very difficult to control, with the result that patients are exposed to high blood glucose and risk of diabetic complications and/or intermittent low blood glucose that can cause unconsciousness, fits and even death. Allograft transplantation of pancreatic islets restores normoglycemia with a low risk of surgical complications. However, although successful immediately after transplantation, islets are progressively lost, with most of the patients requiring exogenous insulin within 2 years post-transplant. Therefore, there is an urgent requirement for the development of new strategies to prevent islet rejection. In this study, we explored the importance of human regulatory T cells in the control of islets allograft rejection. We developed a pre-clinical model of human islet transplantation by reconstituting NOD-scid IL2rγnull mice with cord blood-derived human CD34+ stem cells and demonstrated that although the engrafted human immune system mediated the rejection of human islets, their survival was significantly prolonged following adoptive transfer of ex vivo expanded human Tregs. Mechanistically, Tregs inhibited the infiltration of innate immune cells and CD4+ T cells into the graft by down-regulating the islet graft-derived monocyte chemoattractant protein-1. Our findings might contribute to the development of clinical strategies for Treg therapy to control human islet rejection. We also show for the first time that CD34+ cells-reconstituted NOD-scid IL2rγnull mouse model could be beneficial for investigating human innate immunity in vivo.

Ex Vivo Expanded Human Regulatory T Cells Delay Islet Allograft Rejection via Inhibiting Islet-Derived Monocyte Chemoattractant Protein-1 Production in CD34+ Stem Cells-Reconstituted NOD-scid IL2rγnull Mice

Science.gov (United States)

Xiao, Fang; Ma, Liang; Zhao, Min; Huang, Guocai; Mirenda, Vincenzo; Dorling, Anthony

2014-01-01

Type 1 diabetes mellitus (T1DM) is an autoimmune disease caused by immune-mediated destruction of insulin-secreting β cells of the pancreas. Near complete dependence on exogenous insulin makes T1DM very difficult to control, with the result that patients are exposed to high blood glucose and risk of diabetic complications and/or intermittent low blood glucose that can cause unconsciousness, fits and even death. Allograft transplantation of pancreatic islets restores normoglycemia with a low risk of surgical complications. However, although successful immediately after transplantation, islets are progressively lost, with most of the patients requiring exogenous insulin within 2 years post-transplant. Therefore, there is an urgent requirement for the development of new strategies to prevent islet rejection. In this study, we explored the importance of human regulatory T cells in the control of islets allograft rejection. We developed a pre-clinical model of human islet transplantation by reconstituting NOD-scid IL2rγnull mice with cord blood-derived human CD34+ stem cells and demonstrated that although the engrafted human immune system mediated the rejection of human islets, their survival was significantly prolonged following adoptive transfer of ex vivo expanded human Tregs. Mechanistically, Tregs inhibited the infiltration of innate immune cells and CD4+ T cells into the graft by down-regulating the islet graft-derived monocyte chemoattractant protein-1. Our findings might contribute to the development of clinical strategies for Treg therapy to control human islet rejection. We also show for the first time that CD34+ cells-reconstituted NOD-scid IL2rγnull mouse model could be beneficial for investigating human innate immunity in vivo. PMID:24594640

Ex vivo expanded human regulatory T cells delay islet allograft rejection via inhibiting islet-derived monocyte chemoattractant protein-1 production in CD34+ stem cells-reconstituted NOD-scid IL2rγnull mice.

Directory of Open Access Journals (Sweden)

Fang Xiao

Full Text Available Type 1 diabetes mellitus (T1DM is an autoimmune disease caused by immune-mediated destruction of insulin-secreting β cells of the pancreas. Near complete dependence on exogenous insulin makes T1DM very difficult to control, with the result that patients are exposed to high blood glucose and risk of diabetic complications and/or intermittent low blood glucose that can cause unconsciousness, fits and even death. Allograft transplantation of pancreatic islets restores normoglycemia with a low risk of surgical complications. However, although successful immediately after transplantation, islets are progressively lost, with most of the patients requiring exogenous insulin within 2 years post-transplant. Therefore, there is an urgent requirement for the development of new strategies to prevent islet rejection. In this study, we explored the importance of human regulatory T cells in the control of islets allograft rejection. We developed a pre-clinical model of human islet transplantation by reconstituting NOD-scid IL2rγnull mice with cord blood-derived human CD34+ stem cells and demonstrated that although the engrafted human immune system mediated the rejection of human islets, their survival was significantly prolonged following adoptive transfer of ex vivo expanded human Tregs. Mechanistically, Tregs inhibited the infiltration of innate immune cells and CD4+ T cells into the graft by down-regulating the islet graft-derived monocyte chemoattractant protein-1. Our findings might contribute to the development of clinical strategies for Treg therapy to control human islet rejection. We also show for the first time that CD34+ cells-reconstituted NOD-scid IL2rγnull mouse model could be beneficial for investigating human innate immunity in vivo.

Glucose and carbachol activate phospholipase C in digitonin-permeabilized islets

International Nuclear Information System (INIS)

Wolf, B.A.; Florholmen, J.; Turk, J.; McDaniel, M.L.

1987-01-01

Stimulation of intact islets with D-glucose, the major insulin secretagogue, or with carbachol, a muscarinic agonist, results in the accumulation of inositoltrisphosphate (IP 3 ) suggesting that activation of phospholipase C (PLC) has a major role in stimulus-secretion coupling. Carbachol activation of PLC is an example of receptor-mediated activation in islets, whereas, the mechanism of glucose activation of PLC is controversial since a glucose receptor has not been identified. They have measured PLC activity in digitonin-permeabilized islets. Islets were labeled with 3 H-inositol, permeabilized and IP 3 accumulation measured by HPLC. Carbachol, in the presence of ATP, GTP and 1 μM free Ca 2+ released two-fold more Ins 1,3,4-P 3 than control in a time-dependent manner. Glucose, under the same conditions also significantly released more Ins 1,3,4-P 3 than control. This effect was not due to metabolism of glucose nor to an effect on the IP 3 -phosphomonoesterase. Preliminary Ca 2+ -dependency studies indicate that PLC is not activated by Ca 2+ in the submicromolar range. In conclusion, these studies show that Ca 2+ does not activate PLC, and furthermore, that D-glucose may be recognized directly by PLC

UCP2 mRNA expression is dependent on glucose metabolism in pancreatic islets

International Nuclear Information System (INIS)

Dalgaard, Louise T.

2012-01-01

Highlights: ► UCP2 mRNA levels are decreased in islets of Langerhans from glucokinase deficient mice. ► UCP2 mRNA up-regulation by glucose is dependent on glucokinase. ► Absence of UCP2 increases GSIS of glucokinase heterozygous pancreatic islets. ► This may protect glucokinase deficient mice from hyperglycemic damages. -- Abstract: Uncoupling Protein 2 (UCP2) is expressed in the pancreatic β-cell, where it partially uncouples the mitochondrial proton gradient, decreasing both ATP-production and glucose-stimulated insulin secretion (GSIS). Increased glucose levels up-regulate UCP2 mRNA and protein levels, but the mechanism for UCP2 up-regulation in response to increased glucose is unknown. The aim was to examine the effects of glucokinase (GK) deficiency on UCP2 mRNA levels and to characterize the interaction between UCP2 and GK with regard to glucose-stimulated insulin secretion in pancreatic islets. UCP2 mRNA expression was reduced in GK+/− islets and GK heterozygosity prevented glucose-induced up-regulation of islet UCP2 mRNA. In contrast to UCP2 protein function UCP2 mRNA regulation was not dependent on superoxide generation, but rather on products of glucose metabolism, because MnTBAP, a superoxide dismutase mimetic, did not prevent the glucose-induced up-regulation of UCP2. Glucose-stimulated insulin secretion was increased in UCP2−/− and GK+/− islets compared with GK+/− islets and UCP2 deficiency improved glucose tolerance of GK+/− mice. Accordingly, UCP2 deficiency increased ATP-levels of GK+/− mice. Thus, the compensatory down-regulation of UCP2 is involved in preserving the insulin secretory capacity of GK mutant mice and might also be implicated in limiting disease progression in MODY2 patients.

Comparison of the Ovary and Kidney as Sites for Islet Transplantation in Diabetic Rats.

Science.gov (United States)

Karakose, M; Pinarli, F A; Arslan, M S; Boyuk, G; Boztok, B; Albayrak, A; Ulus, A T; Cakal, E; Delibasi, T

2016-01-01

Currently, the most commonly used site for clinical islet transplantation is the liver although it is far from being an ideal site. Low oxygen tension and the induction of an inflammatory response impair islet implantation and lead to significant early loss of islet. The present study aimed to investigate and compare the efficacy of islet transplantation to the ovary and kidney subcapsule in diabetic rats. The study was performed with 3 groups of rats (control, ovary, and kidney subcapsule) including 6 Sprague female rats each. Diabetes model was created with the use of streptozotocin, and blood glucose levels of the rats were measured after 72 hours. Thirty days after the transplantation, blood samples were obtained from the rats, and then pancreas, kidney, and ovary specimens were fixed in 10% formaldehyde and the experiment completed. After staining with hematoxylin and eosin, the tissue samples were morphologically evaluated by a specialist histopathologist. Changes in mean blood glucose and C-peptide levels were statistically significant in the ovary and kidney subcapsule groups. Histologic examination revealed that granulosus insulin-bearing cells were detected in the islet grafts of both ovary and kidney subcapsule groups. The renal subcapsule group had inflammation signs on histologic examination. The islet cells of both ovary and renal subcapsule groups had no vacuolization. We showed that the ovary might be a new site for islet transplantation. Further research should be done on whether the initial results of this study can be reproduced in larger numbers of animal models and eventually in humans. Copyright © 2016 Elsevier Inc. All rights reserved.

Single-donor islet transplantation and long-term insulin independence in select patients with type 1 diabetes mellitus.

Science.gov (United States)

Al-Adra, David P; Gill, Richdeep S; Imes, Sharleen; O'Gorman, Doug; Kin, Tatsuya; Axford, Sara J; Shi, Xinzhe; Senior, Peter A; Shapiro, A M James

2014-11-15

Islet transplantation is a recognized treatment option for select patients with type I diabetes mellitus. However, islet infusions from multiple donors are often required to achieve insulin independence. Ideally, insulin independence would be achieved routinely with only a single donor. Identification of factors associated with insulin independence after single-donor islet transplantation may help to select recipient-donor combinations with the highest probability of success. Subjects undergoing islet transplantation at a single center (Edmonton, Canada) between March 1999 and August 2013 were included. Recipient, donor, and transplant characteristics were collected and compared between recipients who became insulin independent after one islet transplantation and those who did not. Thirty-one patients achieved insulin independence after a single-donor islet transplantation, and 149 did not. Long-term insulin-free survival was not different between the groups. Factors significantly associated with single-donor success included recipient age, insulin requirement at baseline, donor weight, donor body mass index, islet transplant mass, and peritransplant heparin and insulin administration. On multivariate analysis, pretransplantation daily insulin requirements, the use of peritransplantation heparin and insulin infusions, and islet transplant mass remained significant. We have identified clinically relevant differences defining the achievement of insulin independence after single-donor transplantation. Based on these differences, a preoperative insulin requirement of less than 0.6 U/kg per day and receiving more than 5,646 islet equivalents (IEQ)/kg have a sensitivity of 84% and 71% and specificity of 50% and 50%, respectively, for insulin independence after single-donor islet transplantation. With ideal patient selection, this finding could potentially increase single-donor transplantation success and may be especially relevant for presensitized subjects or those who

Factors influencing the properties and performance of microcapsules for immunoprotection of pancreatic islets

NARCIS (Netherlands)

van Schilfgaarde, R; de Vos, P

There are several approaches of immunoprotection of pancreatic islets for the purpose of successful allo- or xenotransplantation in the absence of immunosuppressive medication. Extravasculair approaches are either mac roencapsulation (large numbers of islets together in one device) or