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Sample records for human islet allografts

  1. 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.

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    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.

  2. Effects of Acute Cytomegalovirus Infection on Rat Islet Allograft Survival

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    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 receive

  3. Human Islet Amyloid Polypeptide

    DEFF Research Database (Denmark)

    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....

  4. Study of the immunoisolating effects of barium-alginate microencapsulation on rat islets allograft survival

    Institute of Scientific and Technical Information of China (English)

    Mei Zhang; Chao Liu; Cuiping Liu; Youwen Qin; Zhaosun Zhen

    2005-01-01

    Objective: To evaluate the immunoisolating effects of barium-alginate microencapsulation on islets allograft survival. Methods: The nonmicroencapsulated and microencapsulated islets were transplanted under the kidney capsule or intraperitoneally into Wistar rat with STZ-induced diabetes. The blood glucose and insulin secretion of grafts were observed. Graft function was tested by oral rats was associated with normal glucose and insulin profiles in response to OGTT. Conclusion: Microencapsulation with barium-alginate membrane can prolong islet survival and protect islets against allorejection.

  5. IL-12p40 is not required for islet allograft rejection

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    En-guang BI; Wei SHI; Jia ZOU; Zhen-hua HAO; Zhen-hu LI; Duan CAI; Hua-qun ZHANG; Bing SUN

    2006-01-01

    Aim: To investigate whether IL-12p40 plays a crucial role in regulating islet allograft rejection in a streptozotocin (STZ)-induced diabetes mouse model. Methods: C57BL/6 and IL-12p40 gene knockout mice were selected as recipient mice, to which the diabetes was induced with a treatment of STZ (150-200 mg/kg) by a single ip injection. BALB/c mice were selected as donor mice and islet cells were isolated from the mice. The 500 islets were transplanted into recipient mice beneath the capsule of the left kidney. Following the islet transplantation the glucose from the mice sera was monitored and the rejection rate of islets was analyzed. Results: STZ could induce diabetes in the recipient mice within 1 week. After transplantation of allograft islets, the increased glucose in wild-type (WT) mice returned to normal level and was maintained for 10 d. Unexpectedly, the rejection rate of islet allograft between IL-12p40-deficient mice and WT mice was similar. Conclusion: The results suggested that, although islet allograft rejection is believed to be Th1-cell predominant, the Th1 response inducer, IL-12 and IL-23 are not essential to induce islet allograft rejection.

  6. 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.

  7. Hair follicle dermal sheath derived cells improve islet allograft survival without systemic immunosuppression.

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    Wang, Xiaojie; Hao, Jianqiang; Leung, Gigi; Breitkopf, Trisia; Wang, Eddy; Kwong, Nicole; Akhoundsadegh, Noushin; Warnock, Garth L; Shapiro, Jerry; McElwee, Kevin J

    2015-01-01

    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.

  8. Reversal of Diabetes by Islet Transplantation: Vulnerability of the Established Allograft

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    Bowen, K. M.; Prowse, S. J.; Lafferty, K. J.

    1981-09-01

    Nonspecific stimulation of the immune system of CBA mice carrying a functional BALB/c islet allograft failed to trigger graft rejection. Only three of six animals rejected their graft when injected intravenously with 105, 106, and 107 peritoneal cells of BALB/c origin over a 3-month period commencing 100 days after transplantation.

  9. Application of anti-CD103 immunotoxin for saving islet allograft in context of transplantation

    Institute of Scientific and Technical Information of China (English)

    ZHANG Lei; Gregg A. Hadley

    2010-01-01

    Background Previous studies using knockout mice document a key role for the integrin CD103 in promoting organ allograft rejection and graft-versus-host disease. However, a determination of whether blockade of the CD103 pathway represents a viable therapeutic strategy for intervention in these processes has proven problematic due to the lack of reagents that efficiently deplete CD103+ cells from wild type hosts. To circumvent this problem, in the present study, we invented an anti-CD103 immunotoxin (M290-SAP). We investigated whether M290-SAP has capacity to eliminate CD103-expressing cells in vivo and protect transplanted islets from destroying by host immune cells.Methods Flow cytometry was used to analyze the efficacy of M290-SAP in depleting CD103-expressing cells in vivo.Then using allogenic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, the therapeutic efficacy of CD103-expressing cell depletion was addressed.Results M290-SAP dramatically reduces the frequency and absolute numbers of CD103-expressing leukocytes in peripheral lymphatic tissues of treated mice. Balb/c islets transplanted into streptozotocin-induced diabetic C57BL/6 mice under single M290-SAP treatment showed an indefinite survival time compared with untreated mice, M290-treated mice and IgG-SAP treated mice (mean survival time, >100 days vs. <20 days). C57BL/6 islets transplanted into hyperglycemic NOD mice under single M290-SAP treatment showed a pronounced delay in allograft rejection compared with untreated mice (mean survival time 12-13 days vs. <7 days). Immunological analysis of mice with long-term islet allograft survival revealed an obvious atrophy thymus and severe downregulation of alloimmunity of CD8 subpopulation response to allogenic stimulation.Conclusion Regardless of the underlying mechanisms, these data document that depletion of CD103-expressing cells represents a viable strategy for therapeutic intervention in islet allograft

  10. A TLR9 agonist promotes IL-22-dependent pancreatic islet allograft survival in type 1 diabetic mice

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    Tripathi, Deepak; Venkatasubramanian, Sambasivan; Cheekatla, Satyanarayana S.; Paidipally, Padmaja; Welch, Elwyn; Tvinnereim, Amy R.; Vankayalapati, Ramakrishna

    2016-01-01

    Pancreatic islet transplantation is a promising potential cure for type 1 diabetes (T1D). Islet allografts can survive long term in the liver parenchyma. Here we show that liver NK1.1+ cells induce allograft tolerance in a T1D mouse model. The tolerogenic effects of NK1.1+ cells are mediated through IL-22 production, which enhances allograft survival and increases insulin secretion. Increased expression of NKG2A by liver NK1.1+ cells in islet allograft-transplanted mice is involved in the production of IL-22 and in the reduced inflammatory response to allografts. Vaccination of T1D mice with a CpG oligonucleotide TLR9 agonist (ODN 1585) enhances expansion of IL-22-producing CD3-NK1.1+ cells in the liver and prolongs allograft survival. Our study identifies a role for liver NK1.1+ cells, IL-22 and CpG oligonucleotides in the induction of tolerance to islet allografts in the liver parenchyma. PMID:27982034

  11. Demand for human allograft tissue in Canada.

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    Lakey, Jonathan R T; Mirbolooki, Mohammadreza; Rogers, Christina; Mohr, Jim

    2007-01-01

    There is relatively little known about the demand for allograft tissues in Canada. The Canadian Council for Donation and Transplantation (CCDT) is a national advisory body that undertook a comprehensive "market survey" to estimate surgical demand for human allograft tissues in Canada. The report "Demand for Human Allograft Tissue in Canada" reflects survey results sent to 5 prominent User Groups. User Groups were identified as orthopaedic surgeons; neurosurgeons; corneal transplant surgeons; plastic surgeons, specifically those at Canadian Burn Units; and cardiac surgeons (adult and paediatric surgery). The demand for allograft grafts was determined and then extrapolated across the total User Group and then increases in allograft tissue use over the next 1-2 years across User Groups were predicted. The overall response rate for the survey was 21.4%. It varied from a low of 19.6% for the orthopaedic survey to a high of 40.5% for the corneal survey. The estimated current demand for allograft tissue in Canada ranges from a low of 34,442 grafts per year to a high of 62,098 grafts per year. The predicted increase in use of allograft tissue over the next 1-2 year period would suggest that annual demand could rise to somewhere in the range of 42,589-72,210 grafts. The highest rated preferences (98% and 94%) were for accredited and Canadian tissue banks, respectively. This study represents a key step in addressing the paucity of information concerning the demand for allograft tissue in Canada.

  12. 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 BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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.

  13. Prolongation of islet allograft survival in mice by combined treatment with pravastatin and low-dose cyclosporine.

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    Arita, S; Kasraie, A; Une, S; Ohtsuka, S; Smith, C V; Mullen, Y

    2001-01-01

    Pravastatin, a 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitor, is known to have suppressive effects on immune and inflammatory cells. We have previously shown in mice and dogs that this agent prevents primary nonfunction of islet iso- and autografts by reducing inflammation at the graft site. The present study was designed to further investigate whether pravastatin has a synergistic effect with cyclosporine (Cs) to prolong islet allograft survival in mice. Unpurified 3000 BALB/c newborn islets were transplanted under the renal capsule of a streptozotocin-diabetic C57BL/6 mouse. Pravastatin and Cs were administered for 10 days starting on the day of grafting (day 0). Five groups were set up based on the treatment protocol: group 1, treatment with 40 mg/kg pravastatin; group 2, 30 mg/kg Cs; group 3, 50 mg/kg Cs; group 4, 40 mg/kg pravastatin and 30 mg/kg Cs; group 5, vehicle alone. Graft survival was indicated by blood glucose levels sustained at 250 mg/dl for 2 consecutive days. Hyperglycemia persisted in six of the eight (75%) mice and grafts were rejected in 3.6 +/- 0.5 days (mean +/- SD) in group 5. In group 1, grafts were also rejected in 3.8 +/- 0.8 days, but blood glucose was transiently 60 days, the other rejected the graft on day 15, and the remaining four died with functioning grafts between 9 and 13 days due to Cs toxicity. A combination of a low dose of Cs and pravastatin (group 4) prolonged graft survival for >19 days in five of the eight mice, and for 7-13 days in the remaining three mice. Histological examination of the grafts in this group showed significantly reduced local inflammation. Results indicate a synergistic effect of pravastatin and Cs on prevention of islet allograft rejection.

  14. Technique of endoscopic biopsy of islet allografts transplanted into the gastric submucosal space in pigs

    NARCIS (Netherlands)

    T. Fujita (Tetsuji); K.M. McGrath (Kevin); R. Bottino (Rita); E.M. Dons (Eefje); C. Long (Cassandra); G. Kumar (Goutham); B. Ekser; G.J. Echeverri (Gabriel); A. Hata (Akira); K. Haruma (Ken); D.K.C. Cooper (David); H. Hara (Hidetaka)

    2013-01-01

    textabstractCurrently, islet cells are transplanted into the liver via portal vein infusion. One disadvantage of this approach is that it is not possible to adequately biopsy the islets in the liver to assess for rejection. Islet transplantation (Tx) into the gastric submucosal space (GSMS) can be p

  15. Early metabolic markers that anticipate loss of insulin independence in type 1 diabetic islet allograft recipients.

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    Hirsch, D; Odorico, J; Danobeitia, J S; Alejandro, R; Rickels, M R; Hanson, M; Radke, N; Baidal, D; Hullett, D; Naji, A; Ricordi, C; Kaufman, D; Fernandez, L

    2012-05-01

    The objective of this study was to identify predictors of insulin independence and to establish the best clinical tools to follow patients after pancreatic islet transplantation (PIT). Sequential metabolic responses to intravenous (I.V.) glucose (I.V. glucose tolerance test [IVGTT]), arginine and glucose-potentiated arginine (glucose-potentiated arginine-induced insulin secretion [GPAIS]) were obtained from 30 patients. We determined the correlation between transplanted islet mass and islet engraftment and tested the ability of each assay to predict return to exogenous insulin therapy. We found transplanted islet mass within an average of 16 709 islet equivalents per kg body weight (IEQ/kg BW; range between 6602 and 29 614 IEQ/kg BW) to be a poor predictor of insulin independence at 1 year, having a poor correlation between transplanted islet mass and islet engraftment. Acute insulin response to IVGTT (AIR(GLU) ) and GPAIS (AIR(max) ) were the most accurate methods to determine suboptimal islet mass engraftment. AIR(GLU) performed 3 months after transplant also proved to be a robust early metabolic marker to predict return to insulin therapy and its value was positively correlated with duration of insulin independence. In conclusion, AIR(GLU) is an early metabolic assay capable of anticipating loss of insulin independence at 1 year in T1D patients undergoing PIT and constitutes a valuable, simple and reliable method to follow patients after transplant.

  16. 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.

  17. [The clinical use of cryopreserved human skin allografts for transplantation].

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    Martínez-Flores, Francisco; Chacón-Gómez, María; Madinaveitia-Villanueva, Juan Antonio; Barrera-Lopez, Araceli; Aguirre-Cruz, Lucinda; Querevalu-Murillo, Walter

    2015-01-01

    The biological recovery of human skin allografts is the gold standard for preservation in Skin Banks. However, there is no worldwide consensus about specific allocation criteria for preserved human skin allografts with living cells. A report is presented on the results of 5 years of experience of using human skin allografts in burned patient in the Skin and Tissue Bank at the "Instituto Nacional de Rehabilitacion" The human skin allografts were obtained from multi-organ donors. processed and preserved at -80 °C for 12 months. Allocation criteria were performed according to blood type match, clinical history, and burned body surface. Up to now, the Skin and Tissue Bank at 'Instituto Nacional de Rehabilitacion" has processed and recovered 125,000 cm(2) of human skin allografts. It has performed 34 surgical implants on 21 burned patients. The average of burn body surface was 59.2%. More than two-thirds (67.7%) of recipients of skin allografts were matched of the same to type blood of the donor, and 66.6% survived after 126 days hospital stay. It is proposed to consider recipient's blood group as allocation criteria to assign tissue; and use human skin allografts on patiens affected with burns over 30% of body surface (according the "rule of the 9"). Copyright © 2015 Academia Mexicana de Cirugía A.C. Published by Masson Doyma México S.A. All rights reserved.

  18. Expression and regulation of nampt in human islets.

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    Karen Kover

    Full Text Available Nicotinamide phosphoribosyltransferase (Nampt is a rate-limiting enzyme in the mammalian NAD+ biosynthesis of a salvage pathway and exists in 2 known forms, intracellular Nampt (iNampt and a secreted form, extracellular Nampt (eNampt. eNampt can generate an intermediate product, nicotinamide mononucleotide (NMN, which has been reported to support insulin secretion in pancreatic islets. Nampt has been reported to be expressed in the pancreas but islet specific expression has not been adequately defined. The aim of this study was to characterize Nampt expression, secretion and regulation by glucose in human islets. Gene and protein expression of Nampt was assessed in human pancreatic tissue and isolated islets by qRT-PCR and immunofluorescence/confocal imaging respectively. Variable amounts of Nampt mRNA were detected in pancreatic tissue and isolated islets. Immunofluorescence staining for Nampt was found in the exocrine and endocrine tissue of fetal pancreas. However, in adulthood, Nampt expression was localized predominantly in beta cells. Isolated human islets secreted increasing amounts of eNampt in response to high glucose (20 mM in a static glucose-stimulated insulin secretion assay (GSIS. In addition to an increase in eNampt secretion, exposure to 20 mM glucose also increased Nampt mRNA levels but not protein content. The secretion of eNampt was attenuated by the addition of membrane depolarization inhibitors, diazoxide and nifedipine. Islet-secreted eNampt showed enzymatic activity in a reaction with increasing production of NAD+/NADH over time. In summary, we show that Nampt is expressed in both exocrine and endocrine tissue early in life but in adulthood expression is localized to endocrine tissue. Enzymatically active eNampt is secreted by human islets, is regulated by glucose and requires membrane depolarization.

  19. Ex Situ Perfusion of Human Limb Allografts for 24 Hours.

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    Werner, Nicole L; Alghanem, Fares; Rakestraw, Stephanie L; Sarver, Dylan C; Nicely, Bruce; Pietroski, Richard E; Lange, Paul; Rudich, Steven M; Mendias, Christopher L; Rojas-Pena, Alvaro; Magee, John C; Bartlett, Robert H; Ozer, Kagan

    2017-03-01

    Vascularized composite allografts, particularly hand and forearm, have limited ischemic tolerance after procurement. In bilateral hand transplantations, this demands a 2 team approach and expedited transfer of the allograft, limiting the recovery to a small geographic area. Ex situ perfusion may be an alternative allograft preservation method to extend allograft survival time. This is a short report of 5 human limbs maintained for 24 hours with ex situ perfusion. Upper limbs were procured from brain-dead organ donors. Following recovery, the brachial artery was cannulated and flushed with 10 000 U of heparin. The limb was then attached to a custom-made, near-normothermic (30-33°C) ex situ perfusion system composed of a pump, reservoir, and oxygenator. Perfusate was plasma-based with a hemoglobin concentration of 4 to 6 g/dL. Average warm ischemia time was 76 minutes. Perfusion was maintained at an average systolic pressure of 93 ± 2 mm Hg, flow 310 ± 20 mL/min, and vascular resistance 153 ± 16 mm Hg/L per minute. Average oxygen consumption was 1.1 ± 0.2 mL/kg per minute. Neuromuscular electrical stimulation continually displayed contraction until the end of perfusion, and histology showed no myocyte injury. Human limb allografts appeared viable after 24 hours of near-normothermic ex situ perfusion. Although these results are early and need validation with transplantation, this technology has promise for extending allograft storage times.

  20. Blockade of the OX40/OX40L pathway and induction of PD-L1 synergistically protects mouse islet allografts from rejection

    Institute of Scientific and Technical Information of China (English)

    Li Tao; Ma Rui; Zhu Jiye; Wang Fushun; Huang Lei; Leng Xisheng

    2014-01-01

    Background OX40/OX40 ligand (OX40/OX40L) and programmed death-1/programmed death ligand-1 (PD-1/PD-L1) costimulatory signals play important roles in T cell-induced immune responses.The aim of this study was to investigate the roles of OX40/OX40L and PD-1/PD-L1 costimulatory pathways in mouse islet allograft rejection.Methods Lentiviral vectors containing OX40L siRNA sequences and an adenovirus vector containing the PD-L1 gene were constructed.The streptozotocin-induced model of diabetes was established in C57BL/6 (H-2b) mice.Diabetic C57BL/6 mice were randomly allocated into five groups:group 1,untreated control; group 2,Ad-EGFP treatment; group 3,Ad-PD-L1 treatment; group 4,OX40L-RNAi-LV treatment; group 5,OX40L-RNAi-LV combined with Ad-PD-L1 treatment.Lentiviral vector and the adenovirus vector were injected,singly or combined,into the caudal vein one day before islet transplantation.The islets of DBA/2 (H-2d) mice were transplanted into the renal subcapsular space of the diabetic recipients.Recipient blood glucose and the survival time of the allografts were monitored.Antigen-specific mixed lymphocyte reaction was also evaluated.Results The recombinant lentiviral RNA interference vector OX40L-RNAi-LV reduced OX40L protein expression by 70%.The recombinant adenovirus vector Ad-PD-L1 increased PD-L1 protein expression in vivo in C57BL/6 recipient mice.Combined OX40L-RNAi-LV/Ad-PD-L1 treatment induced a synergistic protective effect in pancreatic islet allografts.Allograft survival time in the combined treatment group was (92.27±9.65) days,not only longer than that of the control ((6.51±0.27) days) and Ad-EGFP groups ((7.09±0.13) days) (P <0.01),but also significantly longer than that of Ad-PD-L1 and OX40L-RNAi-LV single treatment groups ((40.64±3.95) days and (55.14±5.48) days respectively,P <0.01).The blood glucose concentration of recipient mice in the combined treatment group was also stable and kept within the normal range.Flow cytometry analysis

  1. Anakinra and tocilizumab enhance survival and function of human islets during culture: implications for clinical islet transplantation.

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    Sahraoui, Afaf; Kloster-Jensen, Kristine; Ueland, Thor; Korsgren, Olle; Foss, Aksel; Scholz, Hanne

    2014-01-01

    Pretreatment culture before islet transplantation represents a window of opportunity to ameliorate the proinflammatory profile expressed by human β-cells in duress. Anakinra (IL-1 receptor antagonist) and tocilizumab (monoclonal IL-6 receptor antibody) are two known anti-inflammatory agents successfully used in the treatment of inflammatory states like rheumatoid arthritis. Both compounds have also been shown to reduce blood glucose and glycosylated hemoglobin in diabetic patients. We therefore sought to evaluate the impact of anakinra and tocilizumab on human β-cells. The islets were precultured with or without anakinra or tocilizumab and then transplanted in a marginal mass model using human islets in immunodeficient mice. Islet viability was evaluated in an in vitro model. The pretreatment culture led to a significantly improved engraftment in treated islets compared to the vehicle. Anakinra and tocilizumab are not toxic to human islets and significantly reduce markers of inflammation and cell death. These results strongly support a pretreatment culture with anakinra and tocilizumab prior to human islet transplantation.

  2. 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.......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...... no effect on insulin release in tissue culture, on acute responses to stimulatory glucose concentrations or on the insulin content of cells. These results suggest that dithizone staining can assist in the identification of islets from the human pancreas and may prove to be a useful tool in developing...

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

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    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

  4. Neurotransmitters act as paracrine signals to regulate insulin secretion from the human pancreatic islet.

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    Rodriguez-Diaz, Rayner; Menegaz, Danusa; Caicedo, Alejandro

    2014-08-15

    In this symposium review we discuss the role of neurotransmitters as paracrine signals that regulate pancreatic islet function. A large number of neurotransmitters and their receptors has been identified in the islet, but relatively little is known about their involvement in islet biology. Interestingly, neurotransmitters initially thought to be present in autonomic axons innervating the islet are also present in endocrine cells of the human islet. These neurotransmitters can thus be released as paracrine signals to help control hormone release. Here we propose that the role of neurotransmitters may extend beyond controlling endocrine cell function to work as signals modulating vascular flow and immune responses within the islet.

  5. Generation of pancreatic islet cells from human embryonic stem cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG DongHui; JIANG Wei; SHI Yan; DENG HongKui

    2009-01-01

    Efficiently obtaining functional pancreaUc islet cells derived from human embryonic stem (hES) cells not only provides great potential to solve the shortage of islets sources for type I diabetes cell therapy,but also benefits the study of the development of the human pancreas and diabetes pathology. In 2001,hES cells were reported to have the capacity to generate insulin-producing cells by spontaneous differentiation in vitro. Since then, many strategies (such as overexpression of key transcription factors,delivery of key proteins for pancreatic development, co-transplantation of differentiated hES cells along with fetal pancreas, stepwise differentiation by mimicking in vivo pancreatic development) have been employed in order to induce the differentiation of pancreatic islet cells from hES cells. Moreover, patient-specific induced pluripotent stem (iPS) cells can be generated by reprogramming somatic cells.iPS cells have characteristics similar to those of ES cells and offer a new cell source for type I diabetes cell therapy that reduces the risk of immunologic rejection. In this review, we summarize the recent progress made in the differentiation of hES and iPS cells into functional pancreatic islet cells and discuss the challenges for their future study.

  6. Generation of pancreatic islet cells from human embryonic stem cells

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Efficiently obtaining functional pancreatic islet cells derived from human embryonic stem(hES) cells not only provides great potential to solve the shortage of islets sources for type I diabetes cell therapy,but also benefits the study of the development of the human pancreas and diabetes pathology.In 2001,hES cells were reported to have the capacity to generate insulin-producing cells by spontaneous differentiation in vitro.Since then,many strategies(such as overexpression of key transcription factors,delivery of key proteins for pancreatic development,co-transplantation of differentiated hES cells along with fetal pancreas,stepwise differentiation by mimicking in vivo pancreatic development) have been employed in order to induce the differentiation of pancreatic islet cells from hES cells.Moreover,patient-specific induced pluripotent stem(iPS) cells can be generated by reprogramming somatic cells.iPS cells have characteristics similar to those of ES cells and offer a new cell source for type I diabetes cell therapy that reduces the risk of immunologic rejection.In this review,we summarize the recent progress made in the differentiation of hES and iPS cells into functional pancreatic islet cells and discuss the challenges for their future study.

  7. Transmission of infection with human allografts: essential considerations in donor screening.

    Science.gov (United States)

    Fishman, Jay A; Greenwald, Melissa A; Grossi, Paolo A

    2012-09-01

    Transmission of infection via transplantation of allografts including solid organs, eyes, and tissues are uncommon but potentially life-threatening events. Donor-derived infections have been documented following organ, tissue, and ocular transplants. Each year, more than 70 000 organs, 100 000 corneas, and 2 million human tissue allografts are implanted worldwide. Single donors may provide allografts for >100 organ and tissue recipients; each allograft carries some, largely unquantifiable, risk of disease transmission. Protocols for screening of organ or tissue donors for infectious risk are nonuniform, varying with the type of allograft, national standards, and availability of screening assays. In the absence of routine, active surveillance, coupled with the common failure to recognize or report transmission events, few data are available on the incidence of allograft-associated disease transmission. Research is needed to define the optimal screening assays and the transmissibility of infection with allografts. Approaches are reviewed that may contribute to safety in allograft transplantation.

  8. Human pancreatic islet progenitor cells demonstrate phenotypic plasticity in vitro

    Indian Academy of Sciences (India)

    Maithili P Dalvi; Malati R Umrani; Mugdha V Joglekar; Anandwardhan A Hardikar

    2009-10-01

    Phenotypic plasticity is a phenomenon that describes the occurrence of 2 or more distinct phenotypes under diverse conditions. This article discusses the work carried out over the past few years in understanding the potential of human pancreatic islet-derived progenitors for cell replacement therapy in diabetes. The phenotypic plasticity exhibited by pancreatic progenitors during reversible epithelial-to-mesenchymal transition (EMT) and possible role of microRNAs in regulation of this process is also presented herein.

  9. Research-Focused Isolation of Human Islets From Donors With and Without Diabetes at the Alberta Diabetes Institute IsletCore.

    Science.gov (United States)

    Lyon, James; Manning Fox, Jocelyn E; Spigelman, Aliya F; Kim, Ryekjang; Smith, Nancy; O'Gorman, Doug; Kin, Tatsuya; Shapiro, A M James; Rajotte, Raymond V; MacDonald, Patrick E

    2016-02-01

    Recent years have seen an increased focus on human islet biology, and exciting findings in the stem cell and genomic arenas highlight the need to define the key features of mature human islets and β-cells. Donor and organ procurement parameters impact human islet yield, although for research purposes islet yield may be secondary in importance to islet function. We examined the feasibility of a research-only human islet isolation, distribution, and biobanking program and whether key criteria such as cold ischemia time (CIT) and metabolic status may be relaxed and still allow successful research-focused isolations, including from donors with type 1 diabetes and type 2 diabetes. Through 142 isolations over approximately 5 years, we confirm that CIT and glycated hemoglobin each have a weak negative impacts on isolation purity and yield, and extending CIT beyond the typical clinical isolation cutoff of 12 hours (to ≥ 18 h) had only a modest impact on islet function. Age and glycated hemoglobin/type 2 diabetes status negatively impacted secretory function; however, these and other biological (sex, body mass index) and procurement/isolation variables (CIT, time in culture) appear to make only a small contribution to the heterogeneity of human islet function. This work demonstrates the feasibility of extending acceptable CIT for research-focused human islet isolation and highlights the biological variation in function of human islets from donors with and without diabetes.

  10. IMPROVEMENT OF HUMAN ISLET FUNCTION BY ADENOVIRUS MEDIATED HO-1 GENE TRANSFER

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective To investigate in vitro heme oxygenase-1 gene (HO-1) delivery to human pancreatic islets by adenovirus vectors. Methods Recombinant adenovirus containing HO-1 or enhanced green fluorescent protein gene(EGFP) was generated by using the AdEasy System. The purified human pancreatic islets were infected with recombinant adenovirus vectors at various multiplicity of infection (MOI). Transduction was confirmed by fluorescence photographs and Western blot. Glucose-stimulated insulin secretion was detected by using Human insulin radioimmunoassay kits and was used to assess the function of human islets infected by recombinant adenovirus.Results Viral titers of Ad-hHO-1 and Ad-EGFP were 1.96×109 and 1.99×109 pfu/mL, respectively. Human pancreatic islets were efficiently infected by recombinant adenovirus vectors in vitro. Transfection of human islets at an MOI of 20 did not inhibit islet function. Recombinant adenovirus mediated HO-1gene transfer significantly improved the islet function of insulin release when simulated by high level glucose. Conclusion Recombinant adenovirus is efficient to deliver exogenous gene into human pancreatic islets in vitro. HO-1 gene transfection can improve human islet function.

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

    Directory of Open Access Journals (Sweden)

    Nasser Abualhassan

    Full Text Available 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  13. Transplanted human pancreatic islets after long-term insulin independence

    DEFF Research Database (Denmark)

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

    2013-01-01

    independence. Islets were pooled from two donors with respectively one and five HLA mismatches. Insulin-positive islets were found throughout the right and left liver, and absent in the pancreas. Two- and three-dimensional analysis showed that islets lost their initial rounded and compact morphology, had...

  14. Human Islet Oxygen Consumption Rate and DNA Measurements Predict Diabetes Reversal in Nude Mice

    OpenAIRE

    Papas, K.K.; Colton, C. K.; Nelson, R. A.; Rozak, P.R.; Avgoustiniatos, E.S.; Scott, W. E.; Wildey, G. M.; Pisania, A.; Weir, G. C.; Hering, B. J.

    2007-01-01

    There is a need for simple, quantitative and prospective assays for islet quality assessment that are predictive of islet transplantation outcome. The current state-of-the-art athymic nude mouse bioassay is costly, technically challenging and retrospective. In this study, we report on the ability of 2 parameters characterizing human islet quality: (1) oxygen consumption rate (OCR), a measure of viable volume; and (2) OCR/DNA, a measure of fractional viability, to predict diabetes reversal in ...

  15. Urine Metabolite Profiles Predictive of Human Kidney Allograft Status.

    Science.gov (United States)

    Suhre, Karsten; Schwartz, Joseph E; Sharma, Vijay K; Chen, Qiuying; Lee, John R; Muthukumar, Thangamani; Dadhania, Darshana M; Ding, Ruchuang; Ikle, David N; Bridges, Nancy D; Williams, Nikki M; Kastenmüller, Gabi; Karoly, Edward D; Mohney, Robert P; Abecassis, Michael; Friedewald, John; Knechtle, Stuart J; Becker, Yolanda T; Samstein, Benjamin; Shaked, Abraham; Gross, Steven S; Suthanthiran, Manikkam

    2016-02-01

    Noninvasive diagnosis and prognostication of acute cellular rejection in the kidney allograft may help realize the full benefits of kidney transplantation. To investigate whether urine metabolites predict kidney allograft status, we determined levels of 749 metabolites in 1516 urine samples from 241 kidney graft recipients enrolled in the prospective multicenter Clinical Trials in Organ Transplantation-04 study. A metabolite signature of the ratio of 3-sialyllactose to xanthosine in biopsy specimen-matched urine supernatants best discriminated acute cellular rejection biopsy specimens from specimens without rejection. For clinical application, we developed a high-throughput mass spectrometry-based assay that enabled absolute and rapid quantification of the 3-sialyllactose-to-xanthosine ratio in urine samples. A composite signature of ratios of 3-sialyllactose to xanthosine and quinolinate to X-16397 and our previously reported urinary cell mRNA signature of 18S ribosomal RNA, CD3ε mRNA, and interferon-inducible protein-10 mRNA outperformed the metabolite signatures and the mRNA signature. The area under the receiver operating characteristics curve for the composite metabolite-mRNA signature was 0.93, and the signature was diagnostic of acute cellular rejection with a specificity of 84% and a sensitivity of 90%. The composite signature, developed using solely biopsy specimen-matched urine samples, predicted future acute cellular rejection when applied to pristine samples taken days to weeks before biopsy. We conclude that metabolite profiling of urine offers a noninvasive means of diagnosing and prognosticating acute cellular rejection in the human kidney allograft, and that the combined metabolite and mRNA signature is diagnostic and prognostic of acute cellular rejection with very high accuracy.

  16. Selection of polymers for application in scaffolds applicable for human pancreatic islet transplantation.

    Science.gov (United States)

    Smink, Alexandra M; de Haan, Bart J; Paredes-Juarez, Genaro A; Wolters, Anouk H G; Kuipers, Jeroen; Giepmans, Ben N G; Schwab, Leendert; Engelse, Marten A; van Apeldoorn, Aart A; de Koning, Eelco; Faas, Marijke M; de Vos, Paul

    2016-05-13

    The liver is currently the site for transplantation of islets in humans. This is not optimal for islets, but alternative sites in humans are not available. Polymeric scaffolds in surgically accessible areas are a solution. As human donors are rare, the polymers should not interfere with functional survival of human-islets. We applied a novel platform to test the adequacy of polymers for application in scaffolds for human-islet transplantation. Viability, functionality, and immune parameters were included to test poly(D,L-lactide-co-ε-caprolactone) (PDLLCL), poly(ethylene oxide terephthalate)/polybutylene terephthalate (PEOT/PBT) block copolymer, and polysulfone. The type of polymer influenced the functional survival of human islets. In islets cultured on PDLLCL the glucagon-producing α-cells and insulin-producing β-cells contained more hormone granules than in islets in contact with PEOT/PBT or polysulfone. This was studied with ultrastructural analysis by electron microscopy (nanotomy) during 7 d of culture. PDLLCL was also associated with statistically significant lower release of double-stranded DNA (dsDNA, a so called danger-associate molecular pattern (DAMP)) from islets on PDLLCL when compared to the other polymers. DAMPs support undesired immune responses. Hydrophilicity of the polymers did not influence dsDNA release. Islets on PDLLCL also showed less cellular outgrowth. These outgrowing cells were mainly fibroblast and some β-cells undergoing epithelial to mesenchymal cell transition. None of the polymers influenced the glucose-stimulated insulin secretion. As PDLLCL was associated with less release of DAMPs, it is a promising candidate for creating a scaffold for human islets. Our study demonstrates that for sensitive, rare cadaveric donor tissue such as pancreatic islets it might be necessary to first select materials that do not influence functionality before proposing the biomaterial for in vivo application. Our presented platform may facilitate

  17. Activation of GPR119 Stimulates Human β-Cell Replication and Neogenesis in Humanized Mice with Functional Human Islets

    Science.gov (United States)

    Ansarullah; Free, Colette; Christopherson, Jenica; Chen, Quanhai; Gao, Jie; Liu, Chengyang; Naji, Ali; Rabinovitch, Alex; Guo, Zhiguang

    2016-01-01

    Using humanized mice with functional human islets, we investigated whether activating GPR119 by PSN632408, a small molecular agonist, can stimulate human β-cell regeneration in vivo. Human islets were transplanted under the left kidney capsule of immunodeficient mice with streptozotocin- (STZ-) induced diabetes. The recipient mice were treated with PSN632408 or vehicle and BrdU daily. Human islet graft function in the mice was evaluated by nonfasting glucose levels, oral glucose tolerance, and removal of the grafts. Immunostaining for insulin, glucagon, and BrdU or Ki67 was performed in islet grafts to evaluate α- and β-cell replication. Insulin and CK19 immunostaining was performed to evaluate β-cell neogenesis. Four weeks after human islet transplantation, 71% of PSN632408-treated mice achieved normoglycaemia compared with 24% of vehicle-treated mice. Also, oral glucose tolerance was significantly improved in the PSN632408-treated mice. PSN632408 treatment significantly increased both human α- and β-cell areas in islet grafts and stimulated α- and β-cell replication. In addition, β-cell neogenesis was induced from pancreatic duct cells in the islet grafts. Our results demonstrated that activation of GPR119 increases β-cell mass by stimulating human β-cell replication and neogenesis. Therefore, GPR119 activators may qualify as therapeutic agents to increase human β-cell mass in patients with diabetes. PMID:27413754

  18. Characterization of the Human Pancreatic Islet Proteome by Two-Dimensional LC/MS/MS

    Energy Technology Data Exchange (ETDEWEB)

    Metz, Thomas O.; Jacobs, Jon M.; Gritsenko, Marina A.; Fontes, Ghislaine; Qian, Weijun; Camp, David G.; Poitout, Vincent J.; Smith, Richard D.

    2006-12-01

    Research to elucidate the pathogenesis of type 1 diabetes mellitus has traditionally focused on the genetic and immunological factors associated with the disease, and, until recently, has not considered the target cell. While there have been reports detailing proteomic analyses of established islet cell lines or isolated rodent islets, the information gained is not always easily extrapolated to humans. Therefore, extensive characterization of the human islet proteome could result in better understanding of islet biology and lead to more effective treatment strategies. We have applied a two-dimensional LC-MS/MS-based analysis to the characterization of the human islet proteome, resulting in the detection of 29,021 unique peptides corresponding to 4,925 proteins. As expected, major islet hormones (insulin, glucagon, somatostatin), beta-cell enriched secretory products (IAPP), ion channels (K-ATP channel), and transcription factors (PDX-1, Nkx 6.1, HNF-1 beta) were detected. In addition, significant proteome coverage of metabolic enzymes and cellular pathways was obtained, including the insulin signaling cascade and the MAP kinase, NF-κβ, and JAK/STAT pathways. This work represents the most extensive characterization of the human islet proteome to date and provides a peptide reference library that may be utilized in future studies of islet biology and type 1 diabetes.

  19. Single-Cell Sequencing of Human Pancreatic Islets-New Kids on the Block.

    Science.gov (United States)

    Prasad, Rashmi B; Groop, Leif

    2016-10-11

    RNA sequencing of human pancreatic islets has provided important insights into the islet transcriptome but little information on the specific cells. In this issue, Segerstolpe et al. (2016) and Xin et al. (2016b) report on the transcriptome of single pancreatic cells from non-diabetic and type 2 diabetic donors. Copyright © 2016. Published by Elsevier Inc.

  20. 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.

  1. 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.

  2. Human islets and dendritic cells generate post-translationally modified islet auto-antigens

    NARCIS (Netherlands)

    McLaughlin, Rene J; de Haan, Anne; Zaldumbide, Arnaud; de Koning, Eelco J; de Ru, Arnoud H; van Veelen, Peter A; van Lummel, Menno; Roep, Bart O

    2016-01-01

    Initiation of type 1 diabetes (T1D) requires a break in peripheral tolerance. New insights into neo-epitope formation indicate that post-translational modification of islet auto-antigens, for example via deamidation, may be an important component of disease initiation or exacerbation. Indeed, deamid

  3. Proteins differentially expressed in human beta-cells-enriched pancreatic islet cultures and human insulinomas

    DEFF Research Database (Denmark)

    Terra, Letícia F; Teixeira, Priscila C; Wailemann, Rosangela A M

    2013-01-01

    In view of the great demand for human beta-cells for physiological and medical studies, we generated cell lines derived from human insulinomas which secrete insulin, C-peptide and express neuroendocrine and islet markers. In this study, we set out to characterize their proteomes, comparing them t...

  4. Expression of innate immunity genes and damage of primary human pancreatic islets by epidemic strains of Echovirus: implication for post-virus islet autoimmunity.

    Directory of Open Access Journals (Sweden)

    Luis Sarmiento

    Full Text Available Three large-scale Echovirus (E epidemics (E4,E16,E30, each differently associated to the acute development of diabetes related autoantibodies, have been documented in Cuba. The prevalence of islet cell autoantibodies was moderate during the E4 epidemic but high in the E16 and E30 epidemic. The aim of this study was to evaluate the effect of epidemic strains of echovirus on beta-cell lysis, beta-cell function and innate immunity gene expression in primary human pancreatic islets. Human islets from non-diabetic donors (n = 7 were infected with the virus strains E4, E16 and E30, all isolated from patients with aseptic meningitis who seroconverted to islet cell antibody positivity. Viral replication, degree of cytolysis, insulin release in response to high glucose as well as mRNA expression of innate immunity genes (IFN-b, RANTES, RIG-I, MDA5, TLR3 and OAS were measured. The strains of E16 and E30 did replicate well in all islets examined, resulting in marked cytotoxic effects. E4 did not cause any effects on cell lysis, however it was able to replicate in 2 out of 7 islet donors. Beta-cell function was hampered in all infected islets (P<0.05; however the effect of E16 and E30 on insulin secretion appeared to be higher than the strain of E4. TLR3 and IFN-beta mRNA expression increased significantly following infection with E16 and E30 (P<0.033 and P<0.039 respectively. In contrast, the expression of none of the innate immunity genes studied was altered in E4-infected islets. These findings suggest that the extent of the epidemic-associated islet autoimmunity may depend on the ability of the viral strains to damage islet cells and induce pro-inflammatory innate immune responses within the infected islets.

  5. Monitoring of human liver and kidney allograft tolerance: a tissue/histopathology perspective.

    Science.gov (United States)

    Demetris, Anthony J; Lunz, John G; Randhawa, Parmjeet; Wu, Tong; Nalesnik, Michael; Thomson, Angus W

    2009-01-01

    Several factors acting together have recently enabled clinicians to seriously consider whether chronic immunosuppression is needed in all solid organ allograft recipients. This has prompted a dozen or so centers throughout the world to prospectively wean immunosuppression from conventionally treated liver allograft recipients. The goal is to lessen the impact of chronic immunosuppression and empirically identify occasional recipients who show operational tolerance, defined as gross phenotype of tolerance in the presence of an immune response and/or immune deficit that has little or no significant clinical impact. Rare operationally tolerant kidney allograft recipients have also been identified, usually by single case reports, but only a couple of prospective weaning trials in conventionally treated kidney allograft recipients have been attempted and reported. Pre- and postweaning allograft biopsy monitoring of recipients adds a critical dimension to these trials, not only for patient safety but also for determining whether events in the allografts can contribute to a mechanistic understanding of allograft acceptance. The following is based on a literature review and personal experience regarding the practical and scientific aspects of biopsy monitoring of potential or actual operationally tolerant human liver and kidney allograft recipients where the goal, intended or attained, was complete withdrawal of immunosuppression.

  6. 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 than...... 9 months, with preservation of the ability to release insulin in response to glucose stimulation. Replacement of calf serum with serum from normal human subjects did not affect B-cell survival, but resulted in elevated insulin values partly due to lower insulin degrading activity. Thus the described...

  7. Human pancreatic islet preparations release HMGB1: (ir)relevance for graft engraftment.

    Science.gov (United States)

    Nano, Rita; Racanicchi, Leda; Melzi, Raffaella; Mercalli, Alessia; Maffi, Paola; Sordi, Valeria; Ling, Zhidong; Scavini, Marina; Korsgren, Olle; Celona, Barbara; Secchi, Antonio; Piemonti, Lorenzo

    2013-01-01

    High levels of donor-derived high-mobility group box 1 (HMGB1) protein have been associated with poor islet graft outcome in mouse models. The aim of our work was to determine whether HMGB1 released by human islets had independent proinflammatory effects that influence engraftment in humans. Human islet preparations contained and released HMGB1 in different amounts, as determined by Western blot and ELISA (median 17 pg/ml/IEQ/24 h; min-max 0-211, n = 74). HMGB1 release directly correlated with brain death, donor hyperamilasemia, and factors related to the pancreas digestion procedure (collagenase and digestion time). HMGB1 release was significantly positively associated with the release of other cytokines/chemokines, particularly with the highly released "proinflammatory" CXCL8/IL-8, CXCL1/GRO-α, and the IFN-γ-inducible chemokines CXCL10/IP-10 and CXCL9/MIG. HMGB1 release was not modulated by Toll-like receptor 2, 3, 4, 5, and 9 agonists or by exposure to IL-1β. When evaluated after islet transplantation, pretransplant HMGB1 release was weakly associated with the activation of the coagulation cascade (evaluated as serum cross-linked fibrin products), but not with the immediate posttransplant inflammatory response. Concordantly, HMGB1 did not affect short-term human islet function. Our data show that human islet HMGB1 release is a sign of "damaged" islets, although without any independent direct role in graft failure.

  8. 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

    . These effects were dose-dependent and reproducible when using three different Interleukin-1 preparations. Highly purified human Interleukin-2, Lymphotoxin, Leucocyte Migration Inhibitory Factor and Macrophage Migration Inhibitory Factor were ineffective. These findings suggest that Interleukin-1 may play......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...

  9. Characterization of the human pancreatic islet proteome by two-dimensional LC/MS/MS.

    Science.gov (United States)

    Metz, Thomas O; Jacobs, Jon M; Gritsenko, Marina A; Fontès, Ghislaine; Qian, Wei-Jun; Camp, David G; Poitout, Vincent; Smith, Richard D

    2006-12-01

    The pancreatic beta-cell plays a central role in the maintenance of glucose homeostasis and in the pathogenesis of both type 1 and type 2 diabetes mellitus. Elucidation of the insulin secretory defects observed in diabetes first requires a better understanding of the complex mechanisms regulating insulin secretion, which are only partly understood. While there have been reports detailing proteomic analyses of islet cell lines or isolated rodent islets, the information gained is not always applicable to humans. Therefore, definition of the human islet proteome could contribute to a better understanding of islet biology and lead to more effective treatment strategies. We have applied a two-dimensional LC-MS/MS-based analysis to the characterization of the human islet proteome, resulting in the confident identification of 29,021 different tryptic peptides covering 3365 proteins (> or =2 unique peptide identifications per protein). As expected, the three major islet hormones (insulin, glucagon, and somatostatin) were detected, as well as various beta-cell enriched secretory products, ion channels, and transcription factors. In addition, significant proteome coverage of metabolic enzymes and cellular pathways was observed, including the integrin signaling cascade and the MAP kinase, NF-kappa beta, and JAK/STAT pathways. The resulting peptide reference library provides a resource for future higher throughput and quantitative studies of islet biology.

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

    Directory of Open Access Journals (Sweden)

    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.

  11. Enzymes for Pancreatic Islet Isolation Impact Chemokine-Production and Polarization of Insulin-Producing β-Cells with Reduced Functional Survival of Immunoisolated Rat Islet-Allografts as a Consequence.

    Science.gov (United States)

    de Vos, Paul; Smink, Alexandra M; Paredes, Genaro; Lakey, Jonathan R T; Kuipers, Jeroen; Giepmans, Ben N G; de Haan, Bart J; Faas, Marijke M

    2016-01-01

    The primary aim of this study was to determine whether normal variations in enzyme-activities of collagenases applied for rat-islet isolation impact longevity of encapsulated islet grafts. Also we studied the functional and immunological properties of rat islets isolated with different enzyme preparations to determine whether this impacts these parameters. Rat-islets were isolated from the pancreas with two different collagenases with commonly accepted collagenase, neutral protease, and clostripain activities. Islets had a similar and acceptable glucose-induced insulin-release profile but a profound statistical significant difference in production of the chemokines IP-10 and Gro-α. The islets were studied with nanotomy which is an EM-based technology for unbiased study of ultrastructural features of islets such as cell-cell contacts, endocrine-cell condition, ER stress, mitochondrial conditions, and cell polarization. The islet-batch with higher chemokine-production had a lower amount of polarized insulin-producing β-cells. All islets had more intercellular spaces and less interconnected areas with tight cell-cell junctions when compared to islets in the pancreas. Islet-graft function was studied by implanting encapsulated and free islet grafts in rat recipients. Alginate-based encapsulated grafts isolated with the enzyme-lot inducing higher chemokine production and lower polarization survived for a two-fold shorter period of time. The lower survival-time of the encapsulated grafts was correlated with a higher influx of inflammatory cells at 7 days after implantation. Islets from the same two batches transplanted as free unencapsulated-graft, did not show any difference in survival or function in vivo. Lack of insight in factors contributing to the current lab-to-lab variation in longevity of encapsulated islet-grafts is considered to be a threat for clinical application. Our data suggest that seemingly minor variations in activity of enzymes applied for islet

  12. Amyloid Deposition in Transplanted Human Pancreatic Islets: A Conceivable Cause of Their Long-Term Failure

    Directory of Open Access Journals (Sweden)

    Arne Andersson

    2008-01-01

    Full Text Available Following the encouraging report of the Edmonton group, there was a rejuvenation of the islet transplantation field. After that, more pessimistic views spread when long-term results of the clinical outcome were published. A progressive loss of the β-cell function meant that almost all patients were back on insulin therapy after 5 years. More than 10 years ago, we demonstrated that amyloid deposits rapidly formed in human islets and in mouse islets transgenic for human IAPP when grafted into nude mice. It is, therefore, conceivable to consider amyloid formation as one potential candidate for the long-term failure. The present paper reviews attempts in our laboratories to elucidate the dynamics of and mechanisms behind the formation of amyloid in transplanted islets with special emphasis on the impact of long-term hyperglycemia.

  13. 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...... activity at sites of leukocyte penetration of the peri-islet BM in association with a macrophage subpopulation in NOD mice and human type 1 diabetic samples and, hence, potentially a novel therapeutic target specifically acting at the islet penetration stage. Interestingly, the peri-islet BM and underlying...... 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....

  14. Volumetric properties of human islet amyloid polypeptide in liquid water.

    Science.gov (United States)

    Brovchenko, I; Andrews, M N; Oleinikova, A

    2010-04-28

    The volumetric properties of human islet amyloid polypeptide (hIAPP) in water were studied in a wide temperature range by computer simulations. The intrinsic density rho(p) and the intrinsic thermal expansion coefficient alpha(p) of hIAPP were evaluated by taking into account the difference between the volumetric properties of hydration and bulk water. The density of hydration water rho(h) was found to decrease almost linearly with temperature upon heating and its thermal expansion coefficient was found to be notably higher than that of bulk water. The peptide surface exposed to water is more hydrophobic and its rho(h) is smaller in conformation with a larger number of intrapeptide hydrogen bonds. The two hIAPP peptides studied (with and without disulfide bridge) show negative alpha(p), which is close to zero at 250 K and decreases to approximately -1.5 x 10(-3) K(-1) upon heating to 450 K. The analysis of various structural properties of peptides shows a correlation between the intrinsic peptide volumes and the number of intrapeptide hydrogen bonds. The obtained negative values of alpha(p) can be attributed to the shrinkage of the inner voids of the peptides upon heating.

  15. Aspects of structural landscape of human islet amyloid polypeptide

    Science.gov (United States)

    He, Jianfeng; Dai, Jin; Li, Jing; Peng, Xubiao; Niemi, Antti J.

    2015-01-01

    The human islet amyloid polypeptide (hIAPP) co-operates with insulin to maintain glycemic balance. It also constitutes the amyloid plaques that aggregate in the pancreas of type-II diabetic patients. We have performed extensive in silico investigations to analyse the structural landscape of monomeric hIAPP, which is presumed to be intrinsically disordered. For this, we construct from first principles a highly predictive energy function that describes a monomeric hIAPP observed in a nuclear magnetic resonance experiment, as a local energy minimum. We subject our theoretical model of hIAPP to repeated heating and cooling simulations, back and forth between a high temperature regime where the conformation resembles a random walker and a low temperature limit where no thermal motions prevail. We find that the final low temperature conformations display a high level of degeneracy, in a manner which is fully in line with the presumed intrinsically disordered character of hIAPP. In particular, we identify an isolated family of α-helical conformations that might cause the transition to amyloidosis, by nucleation.

  16. Aspects of structural landscape of human islet amyloid polypeptide

    Energy Technology Data Exchange (ETDEWEB)

    He, Jianfeng, E-mail: hjf@bit.edu.cn; Dai, Jin, E-mail: daijing491@gmail.com [School of Physics, Beijing Institute of Technology, Beijing 100081 (China); Li, Jing, E-mail: jinglichina@139.com [Institute of Biopharmaceutical Research, Yangtze River Pharmaceutical Group Beijing Haiyan Pharmaceutical Co., Ltd, Beijing 102206 (China); Peng, Xubiao, E-mail: xubiaopeng@gmail.com [Department of Physics and Astronomy, Uppsala University, P.O. Box 803, S-75108 Uppsala (Sweden); Niemi, Antti J., E-mail: Antti.Niemi@physics.uu.se [School of Physics, Beijing Institute of Technology, Beijing 100081 (China); Department of Physics and Astronomy, Uppsala University, P.O. Box 803, S-75108 Uppsala (Sweden); Laboratoire de Mathematiques et Physique Theorique CNRS UMR 6083, Fédération Denis Poisson, Université de Tours, Parc de Grandmont, F37200 Tours (France)

    2015-01-28

    The human islet amyloid polypeptide (hIAPP) co-operates with insulin to maintain glycemic balance. It also constitutes the amyloid plaques that aggregate in the pancreas of type-II diabetic patients. We have performed extensive in silico investigations to analyse the structural landscape of monomeric hIAPP, which is presumed to be intrinsically disordered. For this, we construct from first principles a highly predictive energy function that describes a monomeric hIAPP observed in a nuclear magnetic resonance experiment, as a local energy minimum. We subject our theoretical model of hIAPP to repeated heating and cooling simulations, back and forth between a high temperature regime where the conformation resembles a random walker and a low temperature limit where no thermal motions prevail. We find that the final low temperature conformations display a high level of degeneracy, in a manner which is fully in line with the presumed intrinsically disordered character of hIAPP. In particular, we identify an isolated family of α-helical conformations that might cause the transition to amyloidosis, by nucleation.

  17. Adaptive changes of human islets to an obesogenic environment in the mouse

    OpenAIRE

    Gargani, Sofia

    2013-01-01

    Introduction: Under normal healthy conditions, organisms maintain a dynamic endocrinecell mass throughout life. Pancreatic beta cell mass are able to maintain plasma glucose levels increasing insulin secretion in conditions as obesity. Beta cell inability to compensate in insulin demand provokes hyperglycemia and Type 2 Diabetes. Clinically, most obese individuals do not develop diabetes because islets compensate for insulin resistance. Direct evidence that human islet mass adapts longitudina...

  18. The human insulin gene is part of a large open chromatin domain specific for human islets

    OpenAIRE

    Mutskov, Vesco; Felsenfeld, Gary

    2009-01-01

    Knowledge of how insulin (INS) gene expression is regulated will lead to better understanding of normal and abnormal pancreatic β cell function. We have mapped histone modifications over the INS region, coupled with an expression profile, in freshly isolated islets from multiple human donors. Unlike many other human genes, in which active modifications tend to be concentrated within 1 kb around the transcription start site, these marks are distributed over the entire coding region of INS as w...

  19. Pathogen inactivation of human serum facilitates its clinical use for islet cell culture and subsequent transplantation.

    Science.gov (United States)

    Ståhle, Magnus U; Brandhorst, Daniel; Korsgren, Olle; Knutson, Folke

    2011-01-01

    Serum is regarded as an essential supplement to promote survival and growth of cells during culture. However, the potential risk of transmitting diseases disqualifies the use of serum for clinical cell therapy in most countries. Hence, most clinical cell therapy programs have replaced human serum with human serum albumin, which can result in inferior quality of released cell products. Photochemical treatment of different blood products utilizing Intercept® technology has been shown to inactivate a broad variety of pathogens of RNA and DNA origin. The present study assesses the feasibility of using pathogen-inactivated, blood group-compatible serum for use in human pancreatic islet culture. Isolated human islets were cultured at 37°C for 3-4 days in CMRL 1066 supplemented with 10% of either pathogen-inactivated or nontreated human serum. Islet quality assessment included glucose-stimulated insulin release (perifusion), ADP/ATP ratio, cytokine expression, and posttransplant function in diabetic nude mice. No differences were found between islets cultured in pathogen-inactivated or control serum regarding stimulated insulin release, intracellular insulin content, and ADP/ATP ratio. Whether media was supplemented with treated or nontreated serum, islet expression of IL-6, IL-8, MCP-1, or tissue factor was not affected. The final diabetes-reversal rate of mice receiving islets cultured in pathogen-inactivated or nontreated serum was 78% and 87%, respectively (NS). As reported here, pathogen-inactivated human serum does not affect viability or functional integrity of cultured human islets. The implementation of this technology for RNA- and DNA-based pathogen inactivation should enable reintroduction of human serum for clinical cell therapy.

  20. Efficient gene delivery and silencing of mouse and human pancreatic islets

    Directory of Open Access Journals (Sweden)

    Moerman Ericka

    2010-03-01

    Full Text Available Abstract Background In view of the importance of beta cells in glucose homeostasis and the profound repercussions of beta cell pathology on human health, the acquisition of tools to study pancreatic islet function is essential for the design of alternative novel therapies for diabetes. One promising approach toward this goal involves the modification of gene expression profile of beta cells. Results This study describes a new method of gene and siRNA delivery into human pancreatic islets by microporation technology. We demonstrated that mild islet distention with accutase greatly enhanced the transfection efficiency without compromising in vitro function (secretion, apoptosis and viability. As an example, the recently identified gene involved in type 2 diabetes, ZnT8, can be over-expressed or silenced by RNA interference using this technology. Microporation can also be used on rodent islets. Conclusions Taken together, our results demonstrate that microporation technology can be used to modify gene expression in whole rodent and human islets without altering their in vitro function and will be key to the elucidation of the factors responsible for proper islet function.

  1. Controlled aggregation of primary human pancreatic islet cells leads to glucose-responsive pseudoislets comparable to native islets

    NARCIS (Netherlands)

    Hilderink, Janneke; Spijker, Siebe; Carlotti, Françoise; Lange, Lydia; Engelse, Marten; van Blitterswijk, Clemens; de Koning, Eelco; Karperien, Marcel; van Apeldoorn, Aart

    2015-01-01

    Clinical islet transplantation is a promising treatment for patients with type 1 diabetes. However, pancreatic islets vary in size and shape affecting their survival and function after transplantation because of mass transport limitations. To reduce diffusion restrictions and improve islet cell surv

  2. Cytotoxic T-lymphocyte-mediated killing of human pancreatic islet cells in vitro.

    Science.gov (United States)

    Campbell, Peter D; Estella, Eugene; Dudek, Nadine L; Jhala, Gaurang; Thomas, Helen E; Kay, Thomas W H; Mannering, Stuart I

    2008-09-01

    Cytotoxic T lymphocytes (CTL) are believed to play an essential role in beta-cell destruction leading to development of type 1 diabetes and allogeneic islet graft failure. We aimed to identify the mechanisms used by CTL to kill human beta cells. CTL clones that recognize epitopes from influenza virus and Epstein-Barr virus restricted by human leukocyte antigen (HLA)-A0201 and -B0801, respectively, were used to investigate the susceptibility of human beta cells to CTL. In a short-term (5-hour) assay, CTL killed human islet cells of the appropriate major histocompatibility complex (MHC) class I type that had been pulsed with viral peptides. Killing was increased by pretreating islets with interferon gamma that increases MHC class I on target cells. Killing was abolished by incubation of CTL with the perforin inhibitor concanamycin A. The Fas pathway did not contribute to killing because blocking with neutralizing anti-Fas ligand antibody did not significantly reduce beta-cell killing. In conclusion, we report a novel way of investigating the interaction between CTL and human islets. Human islets were rapidly killed in vitro by MHC class I-restricted CTL predominantly by the granule exocytosis pathway.

  3. St. John's wort extract and hyperforin protect rat and human pancreatic islets against cytokine toxicity.

    Science.gov (United States)

    Novelli, Michela; Beffy, Pascale; Menegazzi, Marta; De Tata, Vincenzo; Martino, Luisa; Sgarbossa, Anna; Porozov, Svetlana; Pippa, Anna; Masini, Matilde; Marchetti, Piero; Masiello, Pellegrino

    2014-02-01

    The extract of Hypericum perforatum (St. John's wort, SJW) and its component hyperforin (HPF) were previously shown to inhibit cytokine-induced activation of signal transducer and activator of transcription-1 and nuclear factor κB and prevent apoptosis in a cultured β-cell line. Objective of this study was to assess the protection exerted by SJW and HPF on isolated rat and human islets exposed to cytokines in vitro. Functional, ultrastructural, biomolecular and cell death evaluation studies were performed. In both rat and human islets, SJW and HPF counteracted cytokine-induced functional impairment and down-regulated mRNA expression of pro-inflammatory target genes, such as iNOS, CXCL9, CXCL10, COX2. Cytokine-induced NO production from cultured islets, evaluated by nitrites measurement in the medium, was significantly reduced in the presence of the vegetal compounds. Noteworthy, the increase in apoptosis and necrosis following 48-h exposure to cytokines was fully prevented by SJW and partially by HPF. Ultrastructural morphometric analysis in human islets exposed to cytokines for 20 h showed that SJW or HPF avoided early β-cell damage (e.g., mitochondrial alterations and loss of insulin granules). In conclusion, SJW compounds protect rat and human islets against cytokine effects by counteracting key mechanisms of cytokine-mediated β-cell injury and represent promising pharmacological tools for prevention or limitation of β-cell dysfunction and loss in type 1 diabetes.

  4. The human insulin gene is part of a large open chromatin domain specific for human islets.

    Science.gov (United States)

    Mutskov, Vesco; Felsenfeld, Gary

    2009-10-13

    Knowledge of how insulin (INS) gene expression is regulated will lead to better understanding of normal and abnormal pancreatic beta cell function. We have mapped histone modifications over the INS region, coupled with an expression profile, in freshly isolated islets from multiple human donors. Unlike many other human genes, in which active modifications tend to be concentrated within 1 kb around the transcription start site, these marks are distributed over the entire coding region of INS as well. Moreover, a region of approximately 80 kb around the INS gene, which contains the {tyrosine hydroxylase (TH)-(INS)-insulin-like growth factor 2 antisense (IGF2AS)-insulin-like growth factor 2 (IGF2)} gene cluster, unusually is marked by almost uniformly elevated levels of histone acetylation and H3K4 dimethylation, extending both downstream into IGF2 and upstream beyond the TH gene. This is accompanied by islet specific coordinate expression with INS of the neighboring TH and IGF2 genes. The presence of islet specific intergenic transcripts suggests their possible function in the maintenance of this unusual large open chromatin domain.

  5. Candidate genes expressed in human islets and their role in the pathogenesis of type 1 diabetes

    DEFF Research Database (Denmark)

    Storling, Joachim; Brorsson, Caroline Anna

    2013-01-01

    In type 1 diabetes (T1D), the insulin-producing β cells are destroyed by an immune-mediated process leading to complete insulin deficiency. There is a strong genetic component in T1D. Genes located in the human leukocyte antigen (HLA) region are the most important genetic determinants of disease...... exposure to proinflammatory cytokines highlighting that these genes may be involved in the response of β cells to immune attack. In this review, the compiling evidence that many of the candidate genes are expressed in islets and β cells will be presented. Further, we perform the first systematic human...... islet expression analysis of all genes located in 50 T1D-associated GWAS loci using a published RNA sequencing dataset. We find that 336 out of 857 genes are expressed in human islets and that many of these interact in protein networks. Finally, the potential pathogenetic roles of some candidate genes...

  6. Pancreatic islet allograft in spleen with immunossuppression with cyclosporine. Experimental model in dogs Alotransplante de ilhotas pancreáticas no baço com imunossupressão com ciclosporina. Modelo experimental em cães

    Directory of Open Access Journals (Sweden)

    Jaques Waisberg

    2011-01-01

    Full Text Available PURPOSE: To study the functional behavior of the allograft with immunosuppression of pancreatic islets in the spleen. METHODS: 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. RESULTS: The animals with autotransplant and those with allografts with immunosuppression that became normoglycemic showed altered results of intravenous tolerance glucose (p OBJETIVO: Avaliar o comportamento funcional do alotransplante com imunossupressão de ilhotas pancreáticas no baço. MÉTODOS: Foram utilizados cinco grupos de 10 cães mestiços: grupo A (controle submetido aos exames bioquímicos; grupo B, submetido à pancreatectomia total; grupo C (autotransplante submetido à pancreatectomia total e autotransplantação de ilhotas pancreáticas no baço; grupo D, submetido à alotransplantação de ilhotas pancreáticas no baço sem terapia imunossupressiva; grupo E, submetido à alotransplantação de ilhotas no baço e imunossupressão com ciclosporina. Todos os animais transplantados receberam ilhotas pancreáticas isoladas pelo método mecânico-enzimático, digestão estacionária com colagenase e purificação com gradiente de densidade descontínua de dextran e foram implantadas no baço. RESULTADOS: Animais autotransplantados e alotransplantados com imunossupressão que se tornaram normoglicêmicos apresentaram testes de tolerância à glicose intravenosa alterados (p<0,001 e o

  7. Differential expression of glutamic acid decarboxylase in rat and human islets

    DEFF Research Database (Denmark)

    Petersen, J S; Russel, S; Marshall, M O;

    1993-01-01

    The GABA synthesizing enzyme GAD is a prominent islet cell autoantigen in type I diabetes. The two forms of GAD (GAD64 and GAD67) are encoded by different genes in both rats and humans. By in situ hybridization analysis of rat and human pancreases, expression of both genes was detected in rat isl...

  8. Insulin resistance alters islet morphology in nondiabetic humans

    DEFF Research Database (Denmark)

    Mezza, Teresa; Muscogiuri, Giovanna; Sorice, Gian Pio

    2014-01-01

    Type 2 diabetes is characterized by poor glucose uptake in metabolic tissues and manifests when insulin secretion fails to cope with worsening insulin resistance. In addition to its effects on skeletal muscle, liver, and adipose tissue metabolism, it is evident that insulin resistance also affects...... pancreatic β-cells. To directly examine the alterations that occur in islet morphology as part of an adaptive mechanism to insulin resistance, we evaluated pancreas samples obtained during pancreatoduodenectomy from nondiabetic subjects who were insulin-resistant or insulin-sensitive. We also compared...... insulin sensitivity, insulin secretion, and incretin levels between the two groups. We report an increased islet size and an elevated number of β- and α-cells that resulted in an altered β-cell-to-α-cell area in the insulin- resistant group. Our data in this series of studies suggest that neogenesis from...

  9. An Apparent Deficiency of Lymphatic Capillaries in the Islets of Langerhans in the Human Pancreas.

    Science.gov (United States)

    Korsgren, Erik; Korsgren, Olle

    2016-04-01

    The lymphatic system is crucial for efficient immune surveillance and for the maintenance of a physiological pressure in the interstitial space. Even so, almost no information is available concerning the lymph drainage of the islets of Langerhans in the human pancreas. Immunohistochemical staining allowed us to distinguish lymphatic capillaries from blood capillaries. Almost no lymphatic capillaries were found within the islets in pancreatic biopsy specimens from subjects without diabetes or from subjects with type 1 or type 2 diabetes. Lymphatic capillaries were, however, found at the islet-exocrine interface, frequently located along blood capillaries and other fibrotic structures within or close to the islet capsule. Lymphatic capillaries were regularly found in the exocrine pancreas, with small lymphatic vessels located close to and around acini. Larger collecting lymphatic vessels were located in fibrotic septa between the exocrine lobules and adjacent to the ductal system of the pancreas. In summary, we report a pronounced deficiency of lymphatic capillaries in human islets, a finding with implications for immune surveillance and the regulation of interstitial fluid transport in the endocrine pancreas as well as for the pathophysiology of both type 1 and type 2 diabetes. © 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

  10. Direct long-term effects of L-asparaginase on rat and human pancreatic islets

    DEFF Research Database (Denmark)

    Clausen, Niels; Nielsen, Jens Høiriis

    1989-01-01

    L-Asparaginase, an effective agent in the treatment of acute lymphoblastic leukemia, may induce a diabetic state. The pathogenesis of the diabetogenic effect was studied in cultured pancreatic islets. Mean serum concentrations in three children with acute lymphoblastic leukemia were 2.4 U/mL (range...... 1.4-4.5) before and 31.5 U/mL (range 18.6-51.8) immediately after an intravenous injection of 1000 U/kg L-asparaginase. Glucose-induced insulin release from pancreatic islets of rat and man was measured after 3 and 7 days of culture in media with or without clinically relevant concentrations...... of Escherichia coli L-asparaginase (0.01-100 U/mL). After culture, the remaining insulin, glucagon, and DNA in the islets were determined. After 7 days of culture of adult rat or human islets, both the accumulation of insulin in the medium and the content of insulin and glucagon in the islets were significantly...

  11. Comparison of Modified Celsior Solution and M-Kyoto Solution for Pancreas Preservation in Human Islet Isolation.

    Science.gov (United States)

    Noguchi, Hirofumi; Naziruddin, Bashoo; Onaca, Nicholas; Jackson, Andrew; Shimoda, Masayuki; Ikemoto, Tetsuya; Fujita, Yasutaka; Kobayashi, Naoya; Levy, Marlon F; Matsumoto, Shinichi

    2010-06-01

    Since the successful demonstration of the Edmonton protocol, islet transplantation has advanced significantly on several fronts, including improved pancreas preservation systems. In this study, we evaluated two different types of organ preservation solutions for human islet isolation. Modified Celsior (Celsior solution with hydroxyethyl starch and nafamostat mesilate; HNC) solution and modified Kyoto (MK) solution were compared for pancreas preservation prior to islet isolation. Islet yield after purification was significantly higher in the MK group than in the HNC group (MK = 6186 ± 985 IE/g; HNC = 3091 ± 344 IE/g). The HNC group had a longer phase I period (digestion time), a higher volume of undigested tissue, and a higher percentage of embedded islets, suggesting that the solution may inhibit collagenase. However, there was no significant difference in ATP content in the pancreata or in the attainability of posttransplant normoglycemia in diabetic nude mice between the two groups, suggesting that the quality of islets was similar among the two groups. In conclusion, MK solution is better for pancreas preservation before islet isolation than HNC solution due to the higher percentage of islets that can be isolated from the donor pancreas. MK solution should be the solution of choice among the commercially available solutions for pancreatic islet isolation leading to transplantation.

  12. Comparison of modified Celsior solution and M-kyoto solution for pancreas preservation in human islet isolation.

    Science.gov (United States)

    Noguchi, Hirofumi; Naziruddin, Bashoo; Onaca, Nicholas; Jackson, Andrew; Shimoda, Masayuki; Ikemoto, Tetsuya; Fujita, Yasutaka; Kobayashi, Naoya; Levy, Marlon F; Matsumoto, Shinichi

    2010-01-01

    Since the successful demonstration of the Edmonton protocol, islet transplantation has advanced significantly on several fronts, including improved pancreas preservation systems. In this study, we evaluated two different types of organ preservation solutions for human islet isolation. Modified Celsior (Celsior solution with hydroxyethyl starch and nafamostat mesilate; HNC) solution and modified Kyoto (MK) solution were compared for pancreas preservation prior to islet isolation. Islet yield after purification was significantly higher in the MK group than in the HNC group (MK = 6186 ± 985 IE/g; HNC = 3091 ± 344 IE/g). The HNC group had a longer phase I period (digestion time), a higher volume of undigested tissue, and a higher percentage of embedded islets, suggesting that the solution may inhibit collagenase. However, there was no significant difference in ATP content in the pancreata or in the attainability of posttransplant normoglycemia in diabetic nude mice between the two groups, suggesting that the quality of islets was similar among the two groups. In conclusion, MK solution is better for pancreas preservation before islet isolation than HNC solution due to the higher percentage of islets that can be isolated from the donor pancreas. MK solution should be the solution of choice among the commercially available solutions for pancreatic islet isolation leading to transplantation.

  13. Leptin modulates β cell expression of IL-1 receptor antagonist and release of IL-1β in human islets

    Science.gov (United States)

    Maedler, Kathrin; Sergeev, Pavel; Ehses, Jan A.; Mathe, Zoltan; Bosco, Domenico; Berney, Thierry; Dayer, Jean-Michel; Reinecke, Manfred; Halban, Philippe A.; Donath, Marc Y.

    2004-01-01

    High concentrations of glucose induce β cell production of IL-1β, leading to impaired β cell function and apoptosis in human pancreatic islets. IL-1 receptor antagonist (IL-1Ra) is a naturally occurring antagonist of IL-1β and protects cultured human islets from glucotoxicity. Therefore, the balance of IL-1β and IL-1Ra may play a crucial role in the pathogenesis of diabetes. In the present study, we observed expression of IL-1Ra in human pancreatic β cells of nondiabetic individuals, which was decreased in tissue sections of type 2 diabetic patients. In vitro, chronic exposure of human islets to leptin, a hormone secreted by adipocytes, decreased β cell production of IL-1Ra and induced IL-1β release from the islet preparation, leading to impaired β cell function, caspase-3 activation, and apoptosis. Exogenous addition of IL-1Ra protected cultured human islets from the deleterious effects of leptin. Antagonizing IL-1Ra by introduction of small interfering RNA to IL-1Ra into human islets led to caspase-3 activation, DNA fragmentation, and impaired β cell function. Moreover, siIL-1Ra enhanced glucose-induced β cell apoptosis. These findings demonstrate expression of IL-1Ra in the human β cell, providing localized protection against leptin- and glucose-induced islet IL-1β. PMID:15141093

  14. Human Monoclonal Islet Cell Antibodies From a Patient with Insulin- Dependent Diabetes Mellitus Reveal Glutamate Decarboxylase as the Target Antigen

    Science.gov (United States)

    Richter, Wiltrud; Endl, Josef; Eiermann, Thomas H.; Brandt, Michael; Kientsch-Engel, Rosemarie; Thivolet, Charles; Jungfer, Herbert; Scherbaum, Werner A.

    1992-09-01

    The autoimmune phenomena associated with destruction of the β cell in pancreatic islets and development of type 1 (insulin-dependent) diabetes mellitus (IDDM) include circulating islet cell antibodies. We have immortalized peripheral blood lymphocytes from prediabetic individuals and patients with newly diagnosed IDDM by Epstein-Barr virus transformation. IgG-positive cells were selected by anti-human IgG-coupled magnetic beads and expanded in cell culture. Supernatants were screened for cytoplasmic islet cell antibodies using the conventional indirect immunofluorescence test on cryostat sections of human pancreas. Six islet cell-specific B-cell lines, originating from a patient with newly diagnosed IDDM, could be stabilized on a monoclonal level. All six monoclonal islet cell antibodies (MICA 1-6) were of the IgG class. None of the MICA reacted with human thyroid, adrenal gland, anterior pituitary, liver, lung, stomach, and intestine tissues but all six reacted with pancreatic islets of different mammalian species and, in addition, with neurons of rat cerebellar cortex. MICA 1-6 were shown to recognize four distinct antigenic epitopes in islets. Islet cell antibody-positive diabetic sera but not normal human sera blocked the binding of the monoclonal antibodies to their target epitopes. Immunoprecipitation of 35S-labeled human islet cell extracts revealed that a protein of identical size to the enzyme glutamate decarboxylase (EC 4.1.1.15) was a target of all MICA. Furthermore, antigen immunotrapped by the MICA from brain homogenates showed glutamate decarboxylase enzyme activity. MICA 1-6 therefore reveal glutamate decarboxylase as the predominant target antigen of cytoplasmic islet cell autoantibodies in a patient with newly diagnosed IDDM.

  15. CTLA4Ig基因对大鼠胰岛移植后排斥反应的治疗作用%Treatment of CTLA4Ig gene for the rejection of pancreatic islet allograft in diabetic rats

    Institute of Scientific and Technical Information of China (English)

    金世龙; 刘萍; 顾红光; 王亚旭; 刘宝华; 王代科; 文亚渊; 王仁云; 王东; 周月庆

    2000-01-01

    Objectives To study the expression of CTLA4Ig gene in diabetic rat and the action of CTLA4Ig on prolonging survival time of the pancreatic islet.Methods The rat pancreatic islet cells and muscle cells were transfected with the cDNA for CTLA4Ig packaged with lipofectin vector.The expression levels of CTLA4Ig gene and T lymphocyte reaction(MLR)were determined and the action of CTLA4IgcDNA on prolonging survival time of the pancreatic islet allograft in diabetic rats observed.Results T lymphocyte transforming test from peripheral intravenous blood at 7 th day after pancreatic islet transplantationrevealed that cpm in group A and group B were 175.7±98.2,254.4±116.3,respectively,with the difference between the two groups being significant(P<0.05).At 7th day after pancreatic islet allograft in group A,only 2 out of 10 recipients were serum CTLA4Ig positive with their concentrations being 14 ng/ml and 31 ng/ml respectively. The average time of maintaining blood glucose normal levels in the group A after pancreatic islet allograft,(14.8±12.3)days,was significantly longer than (3.6±5.1)days of the roup B(P<0.05),The average survival time in the group A.(24.0±10.8)days (the longest time was 45 days), was significantly longer than(11.8±4.8)days(the longest time was 21 days)of the group B (P<0.01). Conclusions The muscle cells and pancreatic islets of the recipient rats were transfected with CTLA4Ig cDNA packaged with lipofectin.CTLA4Ig cDNA could be expressed in the recipient tissues. The expressed product could obviously prolong the survival time of the pancreatic islet allograft and the recipient rats and partly inhibit the cellular immunity to prevent rejection.%目的 研究CTLA4Ig基因在糖尿病大鼠体内表达及其产物对胰岛移植物存活的作用.方法 利用Lipofectin载体包裹CTLA41g cDNA 质粒后转染鼠胰岛和肌肉细胞,检测移植后CTLA4Ig表达和T淋巴细胞转化率.结果 胰岛移植术后7dT淋巴细胞转化试验,实验

  16. Whole-Genome Bisulfite Sequencing of Human Pancreatic Islets Reveals Novel Differentially Methylated Regions in Type 2 Diabetes Pathogenesis.

    Science.gov (United States)

    Volkov, Petr; Bacos, Karl; Ofori, Jones K; Esguerra, Jonathan Lou S; Eliasson, Lena; Rönn, Tina; Ling, Charlotte

    2017-04-01

    Current knowledge about the role of epigenetics in type 2 diabetes (T2D) remains limited. Only a few studies have investigated DNA methylation of selected candidate genes or a very small fraction of genomic CpG sites in human pancreatic islets, the tissue of primary pathogenic importance for diabetes. Our aim was to characterize the whole-genome DNA methylation landscape in human pancreatic islets, to identify differentially methylated regions (DMRs) in diabetic islets, and to investigate the function of DMRs in islet biology. Here, we performed whole-genome bisulfite sequencing, which is a comprehensive and unbiased method to study DNA methylation throughout the genome at a single nucleotide resolution, in pancreatic islets from donors with T2D and control subjects without diabetes. We identified 25,820 DMRs in islets from individuals with T2D. These DMRs cover loci with known islet function, e.g., PDX1, TCF7L2, and ADCY5 Importantly, binding sites previously identified by ChIP-seq for islet-specific transcription factors, enhancer regions, and different histone marks were enriched in the T2D-associated DMRs. We also identified 457 genes, including NR4A3, PARK2, PID1, SLC2A2, and SOCS2, that had both DMRs and significant expression changes in T2D islets. To mimic the situation in T2D islets, candidate genes were overexpressed or silenced in cultured β-cells. This resulted in impaired insulin secretion, thereby connecting differential methylation to islet dysfunction. We further explored the islet methylome and found a strong link between methylation levels and histone marks. Additionally, DNA methylation in different genomic regions and of different transcript types (i.e., protein coding, noncoding, and pseudogenes) was associated with islet expression levels. Our study provides a comprehensive picture of the islet DNA methylome in individuals with and without diabetes and highlights the importance of epigenetic dysregulation in pancreatic islets and T2D

  17. Cyclosporine-induced renal dysfunction in human renal allograft recipients.

    Science.gov (United States)

    Kiberd, B A

    1989-12-01

    Cyclosporine-treated renal allograft recipients frequently suffer CsA-related nephrotoxicity and hypertension. This study demonstrates that glomerular filtration rate is reduced acutely by 13% (P less than 0.02) and renal vascular resistance increased by 30% (P less than 0.05), immediately after patients take their CsA dose. The reduction in GFR is directly related to their trough CsA level (r = 0.82; P less than 0.01). The lower the trough CsA level the greater the fall in GFR after the CsA dose. Plasma renin activity does not increase after the CsA dose (pre-CsA 0.6 +/- 0.2 ng/L/sec vs. post-CsA 0.4 +/- 0.1 ng/L/sec; P = NS), and therefore cannot be responsible for the reduction in renal function. Short-term nifedipine treatment is effective in preventing the acute reduction in GFR (P less than 0.05). This occurred despite no apparent effect of nifedipine in altering trough or post-dose CsA levels. Furthermore nifedipine was effective in lowering both the mean arterial blood pressure (109 mmHg to 94 mmHg; P less than 0.01) and the elevated renal vascular resistance (25% reduction; P less than 0.02) observed in these patients. These results suggest that nifedipine may be a suitable agent for limiting acute CsA nephrotoxicity and for treating CsA-associated hypertension in renal allograft recipients.

  18. B7-H4 as a protective shield for pancreatic islet beta cells

    Institute of Scientific and Technical Information of China (English)

    Annika; C; Sun; Dawei; Ou; Dan; S; Luciani; Garth; L; Warnock

    2014-01-01

    Auto- and alloreactive T cells are major culprits that damage β-cells in type 1 diabetes(T1D) and islet transplantation. Current immunosuppressive drugs can alleviate immune-mediated attacks on islets. T cell co-stimulation blockade has shown great promise in autoimmunity and transplantation as it solely targets activated T cells, and therefore avoids toxicity of current immunosuppressive drugs. An attractive approach is offered by the newly-identified negative T cell cosignaling molecule B7-H4 which is expressed in normal human islets, and its expression co-localizes with insulin. A concomitant decrease in B7-H4/insulin colocalization is observed in human type 1 diabetic islets. B7-H4 may play protective roles in the pancreatic islets, preserving their function and survival. In this review we outline the protective effect of B7-H4 in the contexts of T1 D, islet cell transplantation, and potentially type 2 diabetes. Current evidence offers encouraging data regarding the role of B7-H4 in reversal of autoimmune diabetes and donor-specific islet allograft tolerance. Additionally, unique expression of B7-H4 may serve as a potential biomarker for the development of T1 D. Futurestudies should continue to focus on the islet-specific effects of B7-H4 with emphasis on mechanistic pathways in order to promote B7-H4 as a potential therapy and cure for T1 D.

  19. Diabetes mellitus is associated with an increased expression of resistin in human pancreatic islet cells.

    Science.gov (United States)

    Al-Salam, Suhail; Rashed, Hameed; Adeghate, Ernest

    2011-01-01

    The pattern of distribution of resistin in the pancreas of diabetic patients was investigated to determine whether diabetes mellitus influences the expression of resistin. Pancreatic tissue samples retrieved, during pancreatectomy for pancreatic cancer, from cancer patients with and without type 2 diabetes were processed for immunohistochemistry. The pancreatic tissue samples were retrieved from non-cancerous and clear margins. An immunofluorescence technique was used to examine the expression of resistin and its co-localization with insulin and glucagon in pancreatic islet cells. Resistin was observed in many cells located in the central region of pancreatic islet. The expression of resistin increased significantly (p diabetic patients compared to control. Resistin co-localized with insulin but not glucagon in pancreatic islet cells of both normal and diabetic patients. However, the degree of co-localization was higher in pancreata of diabetic patients compared to normal. The number of human pancreatic islet cells expressing resistin increased significantly after the onset of type 2 diabetes. In conclusion, resistin may play a role in the regulation of pancreatic β-cell function.

  20. Clinical Experience: Using Dehydrated Human Amnion/Chorion Membrane Allografts for Acute and Reconstructive Burn Care.

    Science.gov (United States)

    Reilly, Debra Ann; Hickey, Sean; Glat, Paul; Lineaweaver, William C; Goverman, Jeremy

    2017-02-01

    Amniotic membrane is immunologically privileged and is a reservoir of growth factors and cytokines known to modulate inflammation and enhance the healing process, while also possessing antimicrobial, antifibrosis, and antiscarring properties. These properties establish a strong argument for using amniotic membrane derived products as a treatment for burns. The purpose of this article is to describe the use of commercially available dehydrated human amnion/chorion membrane allografts in patients with partial-thickness and full-thickness burns.

  1. Assessment of intracellular insulin content during all steps of human islet isolation procedure.

    Science.gov (United States)

    Brandhorst, H; Brandhorst, D; Brendel, M D; Hering, B J; Bretzel, R G

    1998-01-01

    This study investigated the recovery of pancreatic insulin content during human islet isolation prior to and after digestion-filtration, continuous Hanks-Ficoll gradient purification (n = 20), and 3-4 day culture at 22 degrees C (n = 6). The native insulin content varied in a wide range from 28.4 U to 360.8 U/pancreas. After digestion the initially measured average insulin content of 115.8 +/- 20.8 U/pancreas (mean +/- SEM) increased to 264.6 +/- 22.8% (p asymetrical distribution of insulin within the pancreas. Sampling of insulin within the pancreatic caput seemed not to be representative for the insulin content of the complete native organ, because the ratio of insulin per gram tissue within the pancreatic cauda compared to the caput (n = 5) was 2.4 +/- 0.4 (p < 0.05). After purification total insulin recovery was 55.3 +/- 4.8% (p < 0.001). Because recovery of islet equivalent number (IEQ) (83.7 +/- 4.4%) exceeded insulin recovery, insulin/IEQ ratio decreased from 656.8 +/- 70.6 microU/IEQ before purification to 436.4 +/- 58.1 microU/IEQ (p < 0.001) after purification. After 22 degrees C culture (n = 6) recovery of insulin and IEQ was 80.1 +/- 8.1% (p < 0.05) and 92.8 +/- 3.5% (p = NS), respectively. Insulin content per IEQ decreased to 85.8 +/- 6.5% (p < 0.05). This study clearly shows that most of islet insulin is lost during purification. This seems to be caused rather by an amplified insulin release than by the loss of islets itself. This release may facilitate the separation of endocrine and exocrine tissue by gradient centrifugation, but may also accelerate islet exhaustion detrimental for long-term insulin independence.

  2. Islet transplantation in type 1 diabetes: ongoing challenges, refined procedures, and long-term outcome.

    Science.gov (United States)

    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.

  3. Blood-based biomarkers of age-associated epigenetic changes in human islets associate with insulin secretion and diabetes

    DEFF Research Database (Denmark)

    Bacos, Karl; Gillberg, Linn; Volkov, Petr

    2016-01-01

    Aging associates with impaired pancreatic islet function and increased type 2 diabetes (T2D) risk. Here we examine whether age-related epigenetic changes affect human islet function and if blood-based epigenetic biomarkers reflect these changes and associate with future T2D. We analyse DNA...... methylation genome-wide in islets from 87 non-diabetic donors, aged 26-74 years. Aging associates with increased DNA methylation of 241 sites. These sites cover loci previously associated with T2D, for example, KLF14. Blood-based epigenetic biomarkers reflect age-related methylation changes in 83 genes...... demonstrate that blood-based epigenetic biomarkers reflect age-related DNA methylation changes in human islets, and associate with insulin secretion in vivo and T2D....

  4. A systems genetics approach identifies genes and pathways for type 2 diabetes in human islets

    DEFF Research Database (Denmark)

    Taneera, Jalal; Lang, Stefan; Sharma, Amitabh;

    2012-01-01

    Close to 50 genetic loci have been associated with type 2 diabetes (T2D), but they explain only 15% of the heritability. In an attempt to identify additional T2D genes, we analyzed global gene expression in human islets from 63 donors. Using 48 genes located near T2D risk variants, we identified...... gene coexpression and protein-protein interaction networks that were strongly associated with islet insulin secretion and HbA(1c). We integrated our data to form a rank list of putative T2D genes, of which CHL1, LRFN2, RASGRP1, and PPM1K were validated in INS-1 cells to influence insulin secretion...... of genes potentially involved in T2D....

  5. Cross-correlative 3D micro-structural investigation of human bone processed into bone allografts

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Atul Kumar [Centre for Research in Nanotechnology & Science, Indian Institute of Technology Bombay, Mumbai 400076 (India); Gajiwala, Astrid Lobo [Tissue Bank, Tata Memorial Hospital, Parel, Mumbai 400012 (India); Rai, Ratan Kumar [Centre of Biomedical Research, SGPGIMS Campus, Lucknow 226014 (India); Khan, Mohd Parvez [Division of Endocrinology, Center for Research in Anabolic Skeletal Targets in Health and Illness (ASTHI) CSIR-Central Drug Research Institute, Lucknow 226031 (India); Singh, Chandan [Centre of Biomedical Research, SGPGIMS Campus, Lucknow 226014 (India); Barbhuyan, Tarun [Division of Endocrinology, Center for Research in Anabolic Skeletal Targets in Health and Illness (ASTHI) CSIR-Central Drug Research Institute, Lucknow 226031 (India); Vijayalakshmi, S. [Centre for Research in Nanotechnology & Science, Indian Institute of Technology Bombay, Mumbai 400076 (India); Chattopadhyay, Naibedya [Division of Endocrinology, Center for Research in Anabolic Skeletal Targets in Health and Illness (ASTHI) CSIR-Central Drug Research Institute, Lucknow 226031 (India); Sinha, Neeraj, E-mail: neerajcbmr@gmail.com [Centre of Biomedical Research, SGPGIMS Campus, Lucknow 226014 (India); Kumar, Ashutosh, E-mail: ashutoshk@iitb.ac.in [Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Mumbai 400076 (India); Bellare, Jayesh R., E-mail: jb@iitb.ac.in [Centre for Research in Nanotechnology & Science, Indian Institute of Technology Bombay, Mumbai 400076 (India); Department of Chemical Engineering, Indian Institute of Technology Bombay, Mumbai 400076 (India)

    2016-05-01

    Bone allografts (BA) are a cost-effective and sustainable alternative in orthopedic practice as they provide a permanent solution for preserving skeletal architecture and function. Such BA however, must be processed to be disease free and immunologically safe as well as biologically and clinically useful. Here, we have demonstrated a processing protocol for bone allografts and investigated the micro-structural properties of bone collected from osteoporotic and normal human donor samples. In order to characterize BA at different microscopic levels, a combination of techniques such as Solid State Nuclear Magnetic Resonance (ssNMR), Scanning Electron Microscope (SEM), micro-computed tomography (μCT) and Thermal Gravimetric Analysis (TGA) were used for delineating the ultra-structural property of bone. ssNMR revealed the extent of water, collagen fine structure and crystalline order in the bone. These were greatly perturbed in the bone taken from osteoporotic bone donor. Among the processing methods analyzed, pasteurization at 60 °C and radiation treatment appeared to substantially alter the bone integrity. SEM study showed a reduction in Ca/P ratio and non-uniform distribution of elements in osteoporotic bones. μ-CT and MIMICS® (Materialize Interactive Medical Image Control System) demonstrated that pasteurization and radiation treatment affects the BA morphology and cause a shift in the HU unit. However, the combination of all these processes restored all-important parameters that are critical for BA integrity and sustainability. Cross-correlation between the various probes we used quantitatively demonstrated differences in morphological and micro-structural properties between BA taken from normal and osteoporotic human donor. Such details could also be instrumental in designing an appropriate bone scaffold. For the best restoration of bone microstructure and to be used as a biomaterial allograft, a step-wise processing method is recommended that preserves all

  6. Blood-based biomarkers of age-associated epigenetic changes in human islets associate with insulin secretion and diabetes.

    Science.gov (United States)

    Bacos, Karl; Gillberg, Linn; Volkov, Petr; Olsson, Anders H; Hansen, Torben; Pedersen, Oluf; Gjesing, Anette Prior; Eiberg, Hans; Tuomi, Tiinamaija; Almgren, Peter; Groop, Leif; Eliasson, Lena; Vaag, Allan; Dayeh, Tasnim; Ling, Charlotte

    2016-03-31

    Aging associates with impaired pancreatic islet function and increased type 2 diabetes (T2D) risk. Here we examine whether age-related epigenetic changes affect human islet function and if blood-based epigenetic biomarkers reflect these changes and associate with future T2D. We analyse DNA methylation genome-wide in islets from 87 non-diabetic donors, aged 26-74 years. Aging associates with increased DNA methylation of 241 sites. These sites cover loci previously associated with T2D, for example, KLF14. Blood-based epigenetic biomarkers reflect age-related methylation changes in 83 genes identified in human islets (for example, KLF14, FHL2, ZNF518B and FAM123C) and some associate with insulin secretion and T2D. DNA methylation correlates with islet expression of multiple genes, including FHL2, ZNF518B, GNPNAT1 and HLTF. Silencing these genes in β-cells alter insulin secretion. Together, we demonstrate that blood-based epigenetic biomarkers reflect age-related DNA methylation changes in human islets, and associate with insulin secretion in vivo and T2D.

  7. Islet-like cell aggregates generated from human adipose tissue derived stem cells ameliorate experimental diabetes in mice.

    Directory of Open Access Journals (Sweden)

    Vikash Chandra

    Full Text Available BACKGROUND: Type 1 Diabetes Mellitus is caused by auto immune destruction of insulin producing beta cells in the pancreas. Currently available treatments include transplantation of isolated islets from donor pancreas to the patient. However, this method is limited by inadequate means of immuno-suppression to prevent islet rejection and importantly, limited supply of islets for transplantation. Autologous adult stem cells are now considered for cell replacement therapy in diabetes as it has the potential to generate neo-islets which are genetically part of the treated individual. Adopting methods of islet encapsulation in immuno-isolatory devices would eliminate the need for immuno-suppressants. METHODOLOGY/PRINCIPAL FINDINGS: In the present study we explore the potential of human adipose tissue derived adult stem cells (h-ASCs to differentiate into functional islet like cell aggregates (ICAs. Our stage specific differentiation protocol permit the conversion of mesodermic h-ASCs to definitive endoderm (Hnf3β, TCF2 and Sox17 and to PDX1, Ngn3, NeuroD, Pax4 positive pancreatic endoderm which further matures in vitro to secrete insulin. These ICAs are shown to produce human C-peptide in a glucose dependent manner exhibiting in-vitro functionality. Transplantation of mature ICAs, packed in immuno-isolatory biocompatible capsules to STZ induced diabetic mice restored near normoglycemia within 3-4 weeks. The detection of human C-peptide, 1155±165 pM in blood serum of experimental mice demonstrate the efficacy of our differentiation approach. CONCLUSIONS: h-ASC is an ideal population of personal stem cells for cell replacement therapy, given that they are abundant, easily available and autologous in origin. Our findings present evidence that h-ASCs could be induced to differentiate into physiologically competent functional islet like cell aggregates, which may provide as a source of alternative islets for cell replacement therapy in type 1 diabetes.

  8. Autoimmunity against INS-IGF2 protein expressed in human pancreatic islets.

    Science.gov (United States)

    Kanatsuna, Norio; Taneera, Jalal; Vaziri-Sani, Fariba; Wierup, Nils; Larsson, Helena Elding; Delli, Ahmed; Skärstrand, Hanna; Balhuizen, Alexander; Bennet, Hedvig; Steiner, Donald F; Törn, Carina; Fex, Malin; Lernmark, Åke

    2013-10-04

    Insulin is a major autoantigen in islet autoimmunity and progression to type 1 diabetes. It has been suggested that the insulin B-chain may be critical to insulin autoimmunity in type 1 diabetes. INS-IGF2 consists of the preproinsulin signal peptide, the insulin B-chain, and eight amino acids of the C-peptide in addition to 138 amino acids from the IGF2 gene. We aimed to determine the expression of INS-IGF2 in human pancreatic islets and autoantibodies in newly diagnosed children with type 1 diabetes and controls. INS-IGF2, expressed primarily in beta cells, showed higher levels of expression in islets from normal compared with donors with either type 2 diabetes (p = 0.006) or high HbA1c levels (p INS-IGF2 autoantibody levels were increased in newly diagnosed patients with type 1 diabetes (n = 304) compared with healthy controls (n = 355; p INS-IGF2 revealed that more patients than controls had doubly reactive insulin-INS-IGF2 autoantibodies. These data suggest that INS-IGF2, which contains the preproinsulin signal peptide, the B-chain, and eight amino acids of the C-peptide may be an autoantigen in type 1 diabetes. INS-IGF2 and insulin may share autoantibody-binding sites, thus complicating the notion that insulin is the primary autoantigen in type 1 diabetes.

  9. Treatment with Tacrolimus and Sirolimus Reveals No Additional Adverse Effects on Human Islets In Vitro Compared to Each Drug Alone but They Are Reduced by Adding Glucocorticoids

    Directory of Open Access Journals (Sweden)

    Kristine Kloster-Jensen

    2016-01-01

    Full Text Available Tacrolimus and sirolimus are important immunosuppressive drugs used in human islet transplantation; however, they are linked to detrimental effects on islets and reduction of long-term graft function. Few studies investigate the direct effects of these drugs combined in parallel with single drug exposure. Human islets were treated with or without tacrolimus (30 μg/L, sirolimus (30 μg/L, or a combination thereof for 24 hrs. Islet function as well as apoptosis was assessed by glucose-stimulated insulin secretion (GSIS and Cell Death ELISA. Proinflammatory cytokines were analysed by qRT-PCR and Bio-Plex. Islets exposed to the combination of sirolimus and tacrolimus were treated with or without methylprednisolone (1000 μg/L and the expression of the proinflammatory cytokines was investigated. We found the following: (i No additive reduction in function and viability in islets existed when tacrolimus and sirolimus were combined compared to the single drug. (ii Increased expression of proinflammatory cytokines mRNA and protein levels in islets took place. (iii Methylprednisolone significantly decreased the proinflammatory response in islets induced by the drug combination. Although human islets are prone to direct toxic effect of tacrolimus and sirolimus, we found no additive effects of the drug combination. Short-term exposure of glucocorticoids could effectively reduce the proinflammatory response in human islets induced by the combination of tacrolimus and sirolimus.

  10. Involvement of gene methylation changes in the differentiation of human amniotic epithelial cells into islet-like cell clusters.

    Science.gov (United States)

    Peng, Lin; Wang, Jian; Lu, Guangxiu

    2014-09-01

    Insulin-dependent diabetes results from destruction of the insulin-producing β-cells of the pancreas. Islet cell transplantation is a promising cure for diabetes. Here, we induced human amniotic epithelial cells (hAECs) to differentiate into islet-like cell clusters by nicotinamide plus betacellulin in vitro, and further investigated the DNA methylation status by a Nimble MeDIP microarray before and after cell differentiation to shed light on the molecular mechanisms of this differentiation. In addition, 5-Aza-2'-deoxycytidine was used to investigate whether the differentiation of hAECs into islet-like cells occurred through demethylation. Purified hAECs (CK18(+)/E-cadherin(+)/CD29(+)/CD90(-)/CD34(-)/CD45(-)) were isolated from human amnia. After induction, hAECs were found to be insulin positive and sensitive to glucose, indicating successful induction to islet-like cells. The methylation status of cell cytoskeleton-related genes was down-regulated and that of negative regulation of cell adhesion-related genes was up-regulated. The methylation status of pancreas development-related genes such as HNF1α and DGAT1 was decreased in hAECs after induction. After brief demethylation, INS gene expression was up-regulated in islet-like cell clusters, suggesting that DNA methylation changes were associated with the differentiation of hAECs into islet-like cell clusters.

  11. Distinct differences in the responses of the human pancreatic β-cell line EndoC-βH1 and human islets to proinflammatory cytokines.

    Science.gov (United States)

    Oleson, Bryndon J; McGraw, Jennifer A; Broniowska, Katarzyna A; Annamalai, Mani; Chen, Jing; Bushkofsky, Justin R; Davis, Dawn B; Corbett, John A; Mathews, Clayton E

    2015-09-01

    While insulinoma cells have been developed and proven to be extremely useful in studies focused on mechanisms controlling β-cell function and viability, translating findings to human β-cells has proven difficult because of the limited access to human islets and the absence of suitable insulinoma cell lines of human origin. Recently, a human β-cell line, EndoC-βH1, has been derived from human fetal pancreatic buds. The purpose of this study was to determine whether human EndoC-βH1 cells respond to cytokines in a fashion comparable to human islets. Unlike most rodent-derived insulinoma cell lines that respond to cytokines in a manner consistent with rodent islets, EndoC-βH1 cells fail to respond to a combination of cytokines (IL-1, IFN-γ, and TNF) in a manner consistent with human islets. Nitric oxide, produced following inducible nitric oxide synthase (iNOS) expression, is a major mediator of cytokine-induced human islet cell damage. We show that EndoC-βH1 cells fail to express iNOS or produce nitric oxide in response to this combination of cytokines. Inhibitors of iNOS prevent cytokine-induced loss of human islet cell viability; however, they do not prevent cytokine-induced EndoC-βH1 cell death. Stressed human islets or human islets expressing heat shock protein 70 (HSP70) are resistant to cytokines, and, much like stressed human islets, EndoC-βH1 cells express HSP70 under basal conditions. Elevated basal expression of HSP70 in EndoC-βH1 cells is consistent with the lack of iNOS expression in response to cytokine treatment. While expressing HSP70, EndoC-βH1 cells fail to respond to endoplasmic reticulum stress activators, such as thapsigargin. These findings indicate that EndoC-βH1 cells do not faithfully recapitulate the response of human islets to cytokines. Therefore, caution should be exercised when making conclusions regarding the actions of cytokines on human islets when using this human-derived insulinoma cell line.

  12. Pancreatic hormones are expressed on the surfaces of human and rat islet cells through exocytotic sites

    DEFF Research Database (Denmark)

    Larsson, L I; Hutton, J C; Madsen, O D

    1989-01-01

    . Electron microscopy reveals the labeling to occur at sites of exocytotic granule release, involving the surfaces of extruded granule cores. The surfaces of islet cells were labeled both by polyclonal and monoclonal antibodies, excluding that receptor-interacting, anti-idiotypic hormone antibodies were...... responsible for the staining. Human insulin cells were surface-labeled by monoclonal antibodies recognizing the mature secretory products, insulin and C-peptide but not with monoclonal antibodies specific for proinsulin. Thus, routing of unprocessed preproinsulin to the cell surface may not account...... for these results. It is concluded that the staining reflects interactions between the appropriate antibodies and exocytotic sites of hormone release....

  13. TGFβ Pathway Inhibition Redifferentiates Human Pancreatic Islet β Cells Expanded In Vitro.

    Directory of Open Access Journals (Sweden)

    Ginat Toren-Haritan

    Full Text Available In-vitro expansion of insulin-producing cells from adult human pancreatic islets could provide an abundant cell source for diabetes therapy. However, proliferation of β-cell-derived (BCD cells is associated with loss of phenotype and epithelial-mesenchymal transition (EMT. Nevertheless, BCD cells maintain open chromatin structure at β-cell genes, suggesting that they could be readily redifferentiated. The transforming growth factor β (TGFβ pathway has been implicated in EMT in a range of cell types. Here we show that human islet cell expansion in vitro involves upregulation of the TGFβ pathway. Blocking TGFβ pathway activation using short hairpin RNA (shRNA against TGFβ Receptor 1 (TGFBR1, ALK5 transcripts inhibits BCD cell proliferation and dedifferentiation. Treatment of expanded BCD cells with ALK5 shRNA results in their redifferentiation, as judged by expression of β-cell genes and decreased cell proliferation. These effects, which are reproducible in cells from multiple human donors, are mediated, at least in part, by AKT-FOXO1 signaling. ALK5 inhibition synergizes with a soluble factor cocktail to promote BCD cell redifferentiation. The combined treatment may offer a therapeutically applicable way for generating an abundant source of functional insulin-producing cells following ex-vivo expansion.

  14. Biomechanical Evaluation of Human Allograft Compression in Anterior Cruciate Ligament Reconstruction

    Science.gov (United States)

    Lord, Breck; Yasen, Sam; Amis, Andrew; Wilson, Adrian

    2016-01-01

    Introduction: A common problem encountered during ACL reconstruction is asymmetry of proximal-distal graft diameter leading to tunnel upsizing and potential graft-tunnel mismatch. Human allografts are often oedematous, compounding this issue in the context of multi-ligament reconstructions. Tunnel upsizing reduces bone stock, increases the complexity of multi-bundle surgery and may compromise graft-osseous integration if cortical suspensory fixation is used. Graft compression provides uniform size, allowing easy passage into a smaller tunnel, potentially improving the ‘press-fit’ graft-osseous interaction whilst preserving bone stock. To our knowledge, no biomechanical evaluation of this increasing popular technique has been reported. Hypotheses: Graft compression would not cause any significant changes in the biomechanical properties of human allograft tendon that would be detrimental to the function of an ACL reconstruction. Compressed Bioclense® allograft will increase in size when soaked in Ringer’s solution at 36° improving the ‘press-fit’ within the bone socket, decreasing micro-motion at the graft-osseous interface following ACL reconstruction. Method: In-vitro laboratory study. Sixteen samples of Bioclense® treated peroneus longus allograft were quadrupled into GraftLink constructs randomly divided into control and compressed groups. Cross-sectional area (CSA) was determined using alginate moulds and specimens immersed, under tension, in Ringer’s solution at 36.5°. CSA was measured at 8 hours. A further 32 samples were randomised and evaluated under cyclic loading of 70N-220N (1020 cycles) followed by test to failure. A further 30 samples were quadrupled into GraftLink constructs and mounted within porcine femurs using suspensory fixation. High resolution videometer recorded motion at the graft-osseous interface under the same cyclic loading protocol. An independent samples t-test was used to compare changes in CSA whilst a one-way ANOVA was

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

    Science.gov (United States)

    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.

  16. Differentiation of human multipotent dermal fibroblasts into islet-like cell clusters

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    Liu Wei

    2010-06-01

    Full Text Available Abstract Background We have previously obtained a clonal population of cells from human foreskin that is able to differentiate into mesodermal, ectodermal and endodermal progenies. It is of great interest to know whether these cells could be further differentiated into functional insulin-producing cells. Results Sixty-one single-cell-derived dermal fibroblast clones were established from human foreskin by limiting dilution culture. Of these, two clones could be differentiated into neuron-, adipocyte- or hepatocyte-like cells under certain culture conditions. In addition, those two clones were able to differentiate into islet-like clusters under pancreatic induction. Insulin, glucagon and somatostatin were detectable at the mRNA and protein levels after induction. Moreover, the islet-like clusters could release insulin in response to glucose in vitro. Conclusions This is the first study to demonstrate that dermal fibroblasts can differentiate into insulin-producing cells without genetic manipulation. This may offer a safer cell source for future stem cell-based therapies.

  17. Label-Free Detection of Insulin and Glucagon within Human Islets of Langerhans Using Raman Spectroscopy

    Science.gov (United States)

    Hilderink, Janneke; Otto, Cees; Slump, Cees; Lenferink, Aufried; Engelse, Marten; van Blitterswijk, Clemens; de Koning, Eelco; Karperien, Marcel; van Apeldoorn, Aart

    2013-01-01

    Intrahepatic transplantation of donor islets of Langerhans is a promising therapy for patients with type 1 diabetes. It is of critical importance to accurately monitor islet quality before transplantation, which is currently done by standard histological methods that are performed off-line and require extensive sample preparation. As an alternative, we propose Raman spectroscopy which is a non-destructive and label-free technique that allows continuous real-time monitoring of the tissue to study biological changes as they occur. By performing Raman spectroscopic measurements on purified insulin and glucagon, we showed that the 520 cm-1 band assigned to disulfide bridges in insulin, and the 1552 cm-1 band assigned to tryptophan in glucagon are mutually exclusive and could therefore be used as indirect markers for the label-free distinction between both hormones. High-resolution hyperspectral Raman imaging for these bands showed the distribution of disulfide bridges and tryptophan at sub-micrometer scale, which correlated with the location of insulin and glucagon as revealed by conventional immunohistochemistry. As a measure for this correlation, quantitative analysis was performed comparing the Raman images with the fluorescence images, resulting in Dice coefficients (ranging between 0 and 1) of 0.36 for insulin and 0.19 for glucagon. Although the use of separate microscope systems with different spatial resolution and the use of indirect Raman markers cause some image mismatch, our findings indicate that Raman bands for disulfide bridges and tryptophan can be used as distinctive markers for the label-free detection of insulin and glucagon in human islets of Langerhans. PMID:24167603

  18. A novel comprehensive approach for human vascular allografts cryopreservation and radiation sterilization for the tissue engineering industry

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    Lauk-Dubitsky S.E.

    2015-12-01

    Full Text Available Aim: to verify new techniques for human cadaveric vascular allografts cryopreservation, thawing and sterilization for the tissue engineering purposes. We use polydimethylsiloxane (PDMS as a well-known, promising coolant. This allowed us to completely omit any cryoprotective or vitrifying solutions. Using of PDMS also makes possible an applying these allografts directly after freezing and decellularization and also it will also provide an opportunity to develop secure protocols of tissue— engineered vascular conduits cryopreservation. Matherial and methods. After mathematical modeling of cooling process and its validation the experiment for sealed (isolated freezing at low temperature conditions of 30 femoral arterial segments has been conducted. The segments were at least 10 cm in length and taken from 15 cadaveric donors in the age of 65-85 years. The freezing process was carried out using the abovementioned coolant— PDMS, and then physico-mechanical properties of these allografts were evaluated with the special Instron machine. According to the results obtained, a modeling of their sterilization conditions was conducted (the grafts were freezed. Results. By physico-mechanical properties validation and restricted histological analysis it was shown that there was an accordance between freezed/thawed allografts properties and native vessels. Conclusion. The abovementioned approach for allografts cryopreservation and thawing was efficient enough for further work in this direction.

  19. Characterization of Acute Renal Allograft Rejection by Human Serum Proteomic Analysis

    Institute of Scientific and Technical Information of China (English)

    Ying GAO; Ke WU; Yi XU; Hongmin ZHOU; Wentao HE; Weina ZHANG; Lanjun CAI; Xingguang LIN; Zemin FANG; Zhenlong LUO; Hui GUO; Zhonghua CHEN

    2009-01-01

    To identify acute renal allograft rejection biomarkers in human serum, two-dimensional differential in-gel electrophoresis (2-D DIGE) and reversed phase high-performance liquid chromatog-raphy (RP-HPLC) followed by electrospray ionization mass spectrometry (ESI-MS) were used. Serum samples from renal allograft patients and normal volunteers were divided into three groups: acute rejec-tion (AR), stable renal function (SRF) and normal volunteer (N). Serum samples were firstly processed using Multiple Affinity Removal Column to selectively remove the highest abundance proteins. Differ-entially expressed proteins were analyzed using 2-D DIGE. These differential protein spots were ex-cised, digested by trypsin, and identified by RP-HPLC-ESI/MS. Twenty-two differentially expressed proteins were identified in serum from AR group. These proteins included complement C9 precursor,apolipoprotein A-Ⅳ precursor, vitamin D-binding protein precursor, beta-2-glycoprotein 1 precursor,etc. Vitamin D-binding protein, one of these proteins, was confirmed by ELISA in the independent set of serum samples. In conclusion, the differentially expressed proteins as serum biomarker candidates may provide the basis of acute rejection noninvasive diagnosis. Confirmed vitamin D-binding protein may be one of serum biomarkers of acute rejection. Furthermore, it may provide great insights into un-derstanding the mechanisms and potential treatment strategy of acute rejection.

  20. MicroRNA expression in alpha and beta cells of human pancreatic islets.

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    Dagmar Klein

    Full Text Available microRNAs (miRNAs play an important role in pancreatic development and adult β-cell physiology. Our hypothesis is based on the assumption that each islet cell type has a specific pattern of miRNA expression. We sought to determine the profile of miRNA expression in α-and β-cells, the main components of pancreatic islets, because this analysis may lead to a better understanding of islet gene regulatory pathways. Highly enriched (>98% subsets of human α-and β-cells were obtained by flow cytometric sorting after intracellular staining with c-peptide and glucagon antibody. The method of sorting based on intracellular staining is possible because miRNAs are stable after fixation. MiRNA expression levels were determined by quantitative high throughput PCR-based miRNA array platform screening. Most of the miRNAs were preferentially expressed in β-cells. From the total of 667 miRNAs screened, the Significant Analysis of Microarray identified 141 miRNAs, of which only 7 were expressed more in α-cells (α-miRNAs and 134 were expressed more in β-cells (β-miRNAs. Bioinformatic analysis identified potential targets of β-miRNAs analyzing the Beta Cell Gene Atlas, described in the T1Dbase, the web platform, supporting the type 1 diabetes (T1D community. cMaf, a transcription factor regulating glucagon expression expressed selectively in α-cells (TFα is targeted by β-miRNAs; miR-200c, miR-125b and miR-182. Min6 cells treated with inhibitors of these miRNAs show an increased expression of cMaf RNA. Conversely, over expression of miR-200c, miR-125b or miR-182 in the mouse alpha cell line αTC6 decreases the level of cMAF mRNA and protein. MiR-200c also inhibits the expression of Zfpm2, a TFα that inhibits the PI3K signaling pathway, at both RNA and protein levels.In conclusion, we identified miRNAs differentially expressed in pancreatic α- and β-cells and their potential transcription factor targets that could add new insights into different

  1. Rat-derived processed nerve allografts support more axon regeneration in rat than human-derived processed nerve xenografts.

    Science.gov (United States)

    Wood, Matthew D; Kemp, Stephen W P; Liu, Edward H; Szynkaruk, Mark; Gordon, Tessa; Borschel, Gregory H

    2014-04-01

    Processed nerve allografts are increasingly used as "off the shelf" nerve replacements for surgically bridging nerve gaps. Benchmarking the regenerative capacity of a commercially available human-derived nerve or xenograft in a rat nerve injury model would provide a convenient platform for future studies seeking to modify the processed nerve graft. Human and rat processed nerve grafts were used to bridge a 14 mm defect in a Sprague-Dawley rat sciatic nerve. Reversed autografts served as a positive control group. Twelve weeks following surgery, the distal nerve stumps were retrograde labeled and harvested for histology and histomorphometry. The cross-sectional areas of the human- and rat-derived processed nerve grafts were similar. Neuron counts and myelinated axon counts following use of the human-derived processed xenografts were decreased compared with those obtained from both the rat-derived processed nerve allografts and the autografts; the rat-derived processed nerve allografts were statistically equivalent to autografts. Measures of nerve fiber diameter and myelination revealed inferior axon regeneration maturity in both processed nerve grafts compared with autografts. Processed xenografts showed significantly reduced regeneration compared with autografts or processed allografts indicating that cross-species immunological reactions are important considerations in this rat model. Copyright © 2013 Wiley Periodicals, Inc.

  2. Islet transplantation: immunological perspectives.

    Science.gov (United States)

    Inverardi, Luca; Kenyon, Norma S; Ricordi, Camillo

    2003-10-01

    Clinical trials of islet transplantation are showing remarkable success, but they require administration of chronic immunosuppression, and are underscoring the large gap that exists between the number of human donors available and the number of patients that could benefit from the procedure. Recent progress has been made in the definition of key immunological mechanisms that are involved in determining islet transplant outcome. Clinical and preclinical studies, and studies in small animal model systems, will all eventually contribute to the definition of efficient and safe protocols for islet transplantation. If the use of xenografts is successful, it might represent a solution to the shortage of human organs.

  3. Single-cell transcriptomes identify human islet cell signatures and reveal cell-type–specific expression changes in type 2 diabetes

    Science.gov (United States)

    Bolisetty, Mohan; Kursawe, Romy; Sun, Lili; Sivakamasundari, V.; Kycia, Ina

    2017-01-01

    Blood glucose levels are tightly controlled by the coordinated action of at least four cell types constituting pancreatic islets. Changes in the proportion and/or function of these cells are associated with genetic and molecular pathophysiology of monogenic, type 1, and type 2 (T2D) diabetes. Cellular heterogeneity impedes precise understanding of the molecular components of each islet cell type that govern islet (dys)function, particularly the less abundant delta and gamma/pancreatic polypeptide (PP) cells. Here, we report single-cell transcriptomes for 638 cells from nondiabetic (ND) and T2D human islet samples. Analyses of ND single-cell transcriptomes identified distinct alpha, beta, delta, and PP/gamma cell-type signatures. Genes linked to rare and common forms of islet dysfunction and diabetes were expressed in the delta and PP/gamma cell types. Moreover, this study revealed that delta cells specifically express receptors that receive and coordinate systemic cues from the leptin, ghrelin, and dopamine signaling pathways implicating them as integrators of central and peripheral metabolic signals into the pancreatic islet. Finally, single-cell transcriptome profiling revealed genes differentially regulated between T2D and ND alpha, beta, and delta cells that were undetectable in paired whole islet analyses. This study thus identifies fundamental cell-type–specific features of pancreatic islet (dys)function and provides a critical resource for comprehensive understanding of islet biology and diabetes pathogenesis. PMID:27864352

  4. Human Islet Amyloid Polypeptide Transgenic Mice: In Vivo and Ex Vivo Models for the Role of hIAPP in Type 2 Diabetes Mellitus

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    J. W. M. Höppener

    2008-01-01

    Full Text Available Human islet amyloid polypeptide (hIAPP, a pancreatic islet protein of 37 amino acids, is the main component of islet amyloid, seen at autopsy in patients with type 2 diabetes mellitus (DM2. To investigate the roles of hIAPP and islet amyloid in DM2, we generated transgenic mice expressing hIAPP in their islet beta cells. In this study, we found that after a long-term, high-fat diet challenge islet amyloid was observed in only 4 of 19 hIAPP transgenic mice. hIAPP transgenic females exhibited severe glucose intolerance, which was associated with a downregulation of GLUT-2 mRNA expression. In isolated islets from hIAPP males cultured for 3 weeks on high-glucose medium, the percentage of amyloid containing islets increased from 5.5% to 70%. This ex vivo system will allow a more rapid, convenient, and specific study of factors influencing islet amyloidosis as well as of therapeutic strategies to interfere with this pathological process.

  5. Induction of human umbilical cord blood-derived stem cells with embryonic stem cell phenotypes into insulin producing islet-like structure.

    Science.gov (United States)

    Sun, Bo; Roh, Kyung-Hwan; Lee, Sae-Rom; Lee, Yong-Soon; Kang, Kyung-Sun

    2007-03-23

    Success in islet-transplantation-based therapies for type I diabetes, coupled with a worldwide shortage of transplant-ready islets, has motivated efforts to develop renewable sources of islet-replacement tissue. Embryonic stem cells (ESCs) have been successfully induced into insulin producing islet-like structure in several studies. However, the source of the ESCs has presented ethical and technical concerns. Here, we isolated a population of stem cells from human cord blood (UCB), which expressed embryo stage specific maker, SSEA-4, and the multi-potential stem cell marker, Oct4. Subsequently, we successfully induced them into insulin-producing islet-like structures, which co-express insulin and C-peptide. These findings might have a significant potential to advance human UCB derived stem-cell-based therapeutics for diabetes.

  6. Ridge preservation comparing socket allograft alone to socket allograft plus facial overlay xenograft: a clinical and histologic study in humans.

    Science.gov (United States)

    Poulias, Evmenios; Greenwell, Henry; Hill, Margaret; Morton, Dean; Vidal, Ricardo; Shumway, Brian; Peterson, Thomas L

    2013-11-01

    Previous studies of ridge preservation showed a loss of ≈18% or 1.5 mm of crestal ridge width in spite of treatment. The primary aim of this randomized, controlled, masked clinical trial is to compare a socket graft to the same treatment plus a buccal overlay graft, both with a polylactide membrane, to determine if loss of ridge width can be prevented by use of an overlay graft. Twelve patients who served as positive controls received an intrasocket mineralized cancellous allograft (socket group), and 12 patients received the same socket graft procedure plus buccal overlay cancellous xenograft (overlay group). Horizontal ridge dimensions were measured with a digital caliper, and vertical ridge changes were measured from a stent. Before implant placement, at 4 months, a trephine core was obtained for histologic analysis. The mean horizontal ridge width at the crest for the socket group decreased from 8.7 ± 1.0 to 7.1 ± 1.5 mm for a mean loss of 1.6 ± 0.8 mm (P overlay group decreased from 8.4 ± 1.4 to 8.1 ± 1.4 mm for a mean loss of 0.3 ± 0.9 mm (P >0.05). The overlay group was significantly different from the socket group (P overlay group had 40% ± 16% (P >0.05). The overlay treatment significantly prevented loss of ridge width and preserved or augmented the buccal contour. The socket and overlay groups healed with a high percentage of vital bone.

  7. Identification of Donor Origin and Condition of Transplanted Islets In Situ in the Liver of a Type 1 Diabetic Recipient.

    Science.gov (United States)

    van der Torren, Cornelis R; Suwandi, Jessica S; Lee, DaHae; Van't Wout, Ernst-Jan T; Duinkerken, Gaby; Swings, Godelieve; Mulder, Arend; Claas, Frans H J; Ling, Zhidong; Gillard, Pieter; Keymeulen, Bart; In't Veld, Peter; Roep, Bart O

    2017-01-24

    Transplantation of islet allografts into type 1 diabetic recipients usually requires multiple pancreas donors to achieve insulin independence. This adds to the challenges of immunological monitoring of islet transplantation currently relying on surrogate immune markers in peripheral blood. We investigated donor origin and infiltration of islets transplanted in the liver of a T1D patient who died of hemorrhagic stroke 4 months after successful transplantation with two intraportal islet grafts combining six donors. Immunohistological staining for donor HLA using a unique panel of human monoclonal HLA-specific alloantibodies was performed on liver cryosections after validation on cryopreserved kidney, liver, and pancreas and compared with auto- and alloreactive T-cell immunity in peripheral blood. HLA-specific staining intensity and signal-to-noise ratio varied between tissues from very strong on kidney glomeruli, less in liver, kidney tubuli, and endocrine pancreas to least in exocrine pancreas, complicating the staining of inflamed islets in an HLA-disparate liver. Nonetheless, five islets from different liver lobes could be attributed to donors 1, 2, and 5 by staining patterns with multiple HLA types. All islets showed infiltration with CD8+ cytotoxic T cells that was mirrored by progressive alloreactive responses in peripheral blood mononuclear cells (PBMCs) to donors 1, 2, and 5 after transplantation. Stably low rates of peripheral islet autoreactive T-cell responses after islet infusion fit with a complete HLA mismatch between grafts and recipient and exclude the possibility that the islet-infiltrating CD8 T cells were autoreactive. HLA-specific immunohistochemistry can identify donor origin in situ and differentiate graft dysfunction and immunological destruction.

  8. Human fetal islet transplantation in type 1 diabetic patients: comparison of metabolic effects between single and multiple implantation regimens.

    Science.gov (United States)

    Djordjevic, P B; Lalic, N M; Jotic, A; Paunovic, I; Lalic, K; Raketic, N; Nikolic, D; Zamaklar, M; Rajkovic, N; Lukic, L; Dimitrijevic-Sreckovic, V; Dragasevic, M; Nikolic, D; Markovic, I

    2004-11-01

    Previous studies suggest that multiple transplantations might be equally efficient to a single regimen for human adult islets. The aim of this study was to compare metabolic parameters after each of the two regimens of human fetal islet (HFI) transplantation in type 1 diabetics. In group A (single transplant, n = 9), 180 +/- 20 x 1000 HFI equivalents (IEQs) were implanted by a single IM injection; in group B (multiple transplants, n = 8) islets were implanted as three consecutive injections (60 +/- 10 x 1000 IEQs) at 7-day intervals. We analyzed the metabolic parameters on days -1, 30, 60, 90, 120, 150, and 180 after the procedure. Among the metabolic parameters, we evaluated insulin secretion capacity-ISC (C peptide, RIA), metabolic control (HbA1c, chromatography), and insulin daily dose IDD. We found that C peptide levels increased, peaking on day 90 (A: 0.38 +/- 0.15; B: 0.34 +/- 0.19 nmol/L, P = NS) and then rapidly decreasing without differences, the HbA1c levels and IDD decreased in the same manner without differences between the groups. Our results demonstrate that multiple and single islet transplant regimens are equally efficient to temporarily restore a significant ISC with improvement of metabolic and clinical parameters. The results imply that the two regimens have an equal clinical value.

  9. Glucose stimulates human beta cell replication in vivo in islets transplanted into NOD–severe combined immunodeficiency (SCID) mice

    Science.gov (United States)

    Levitt, H. E.; Cyphert, T. J.; Pascoe, J. L.; Hollern, D. A.; Abraham, N.; Lundell, R. J.; Rosa, T.; Romano, L. C.; Zou, B.; O’Donnell, C. P.; Stewart, A. F.; Garcia-Ocaña, A.; Alonso, L. C.

    2011-01-01

    Aims/hypothesis We determined whether hyperglycaemia stimulates human beta cell replication in vivo in an islet transplant model Methods Human islets were transplanted into streptozotocin-induced diabetic NOD–severe combined immunodeficiency mice. Blood glucose was measured serially during a 2 week graft revascularisation period. Engrafted mice were then catheterised in the femoral artery and vein, and infused intravenously with BrdU for 4 days to label replicating beta cells. Mice with restored normoglycaemia were co-infused with either 0.9% (wt/vol.) saline or 50% (wt/vol.) glucose to generate glycaemic differences among grafts from the same donors. During infusions, blood glucose was measured daily. After infusion, human beta cell replication and apoptosis were measured in graft sections using immunofluorescence for insulin, and BrdU or TUNEL. Results Human islet grafts corrected diabetes in the majority of cases. Among grafts from the same donor, human beta cell proliferation doubled in those exposed to higher glucose relative to lower glucose. Across the entire cohort of grafts, higher blood glucose was strongly correlated with increased beta cell replication. Beta cell replication rates were unrelated to circulating human insulin levels or donor age, but tended to correlate with donor BMI. Beta cell TUNEL reactivity was not measurably increased in grafts exposed to elevated blood glucose. Conclusions/interpretation Glucose is a mitogenic stimulus for transplanted human beta cells in vivo. Investigating the underlying pathways may point to mechanisms capable of expanding human beta cell mass in vivo. PMID:20936253

  10. Islet expression of the DNA repair enzyme 8-oxoguanosine DNA glycosylase (Ogg1 in human type 2 diabetes

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    Yoon Kun-Ho

    2002-04-01

    Full Text Available Abstract Background It has become increasingly clear that β-cell failure plays a critical role in the pathogenesis of type 2 diabetes. Free-radical mediated β-cell damage has been intensively studied in type 1 diabetes, but not in human type 2 diabetes. Therefore, we studied the protein expression of the DNA repair enzyme Ogg1 in pancreases from type 2 diabetics. Ogg1 was studied because it is the major enzyme involved in repairing 7,8-dihydro-8-oxoguanosine DNA adducts, a lesion previously observed in a rat model of type 2 diabetes. Moreover, in a gene expression screen, Ogg1 was over-expressed in islets from a human type 2 diabetic. Methods Immunofluorescent staining of Ogg1 was performed on pancreatic specimens from healthy controls and patients with diabetes for 2–23 years. The intensity and islet area stained for Ogg1 was evaluated by semi-quantitative scoring. Results Both the intensity and the area of islet Ogg1 staining were significantly increased in islets from the type 2 diabetic subjects compared to the healthy controls. A correlation between increased Ogg1 fluorescent staining intensity and duration of diabetes was also found. Most of the staining observed was cytoplasmic, suggesting that mitochondrial Ogg1 accounts primarily for the increased Ogg1 expression. Conclusion We conclude that oxidative stress related DNA damage may be a novel important factor in the pathogenesis of human type 2 diabetes. An increase of Ogg1 in islet cell mitochondria is consistent with a model in which hyperglycemia and consequent increased β-cell oxidative metabolism lead to DNA damage and the induction of Ogg1 expression.

  11. Differential sensitivity to beta-cell secretagogues in cultured rat pancreatic islets exposed to human interleukin-1 beta.

    Science.gov (United States)

    Eizirik, D L; Sandler, S; Hallberg, A; Bendtzen, K; Sener, A; Malaisse, W J

    1989-08-01

    The early stages of insulin-dependent diabetes mellitus are characterized by a selective inability to secrete insulin in response to glucose, coupled to a better response to nonnutrient secretagogues. The deficient glucose response may be a result of the autoimmune process directed toward the beta-cells. Interleukin-1 (IL-1) has been suggested to be one possible mediator of immunological damage of the beta-cells. In the present study we characterized the sensitivity of beta-cells to different secretagogues after human recombinant IL-1 beta (rIL-1 beta) exposure. Furthermore, experiments were performed to clarify the biochemical mechanisms behind the defective insulin response observed in these islets. Rat pancreatic islets were isolated and kept in tissue culture (medium RPMI-1640 plus 10% calf serum) for 5 days. The islets were subsequently exposed to 60 pM human recombinant IL-1 beta during 48 h in the same culture conditions as above and examined immediately after IL-1 exposure. The rIL-1 beta-treated islets showed a marked reduction of glucose-stimulated insulin release. Stimulation with arginine plus different glucose concentrations, and leucine plus glutamine partially counteracted the rIL-1 beta-induced reduction of insulin release. The activities of the glycolytic enzymes hexokinase, glucokinase, and glyceraldehyde 3-phosphate dehydrogenase, were similar in control and IL-1-exposed islets. Treatment with IL-1 also did not impair the activities of NADH+- and NADPH+-dependent glutamate dehydrogenase, glutamate-aspartate transaminase, glutamate-alanine transaminase, citrate synthase, and NAD+-linked isocitrate dehydrogenase. The oxidation of D-[6-14C]glucose and L-[U-14C]leucine were decreased by 50% in IL-1-treated islets. Furthermore, there was a significant decrease in the ratios of [2-14C]pyruvate oxidation/[1-14C]pyruvate decarboxylation and L-[U-14C]leucine oxidation/L-[1-14C]leucine decarboxylation, indicating that IL-1 decreases the proportion of

  12. Residue specific effects of human islet polypeptide amyloid on self-assembly and on cell toxicity.

    Science.gov (United States)

    Khemtemourian, Lucie; Guillemain, Ghislaine; Foufelle, Fabienne; Killian, J Antoinette

    2017-08-01

    Type 2 diabetes mellitus is characterized histopathologically by the presence of fibrillary amyloid deposits in the pancreatic islets of Langerhans. Human islet amyloid polypeptide (hIAPP), the 37-residue pancreatic hormone, is the major constituent of these amyloid deposits. The propensity of IAPP to form amyloid fibrils is strongly dependent on its primary sequence. An intriguing example is His at residue 18. Although H18 is located outside the amyloidogenic region, it has been suggested that this residue and its charge state play an important role in the kinetics of conformational changes and fibril formation as well as in mediating cell toxicity. To gain more insight into the importance of this residue, we have synthesized four analogues (H18R-IAPP, H18K-IAPP, H18A-IAPP and H18E-IAPP) and we performed a full biophysical study on the properties of these peptides. Kinetic experiments as monitored by thioflavin-T fluorescence, transmission electron microscopy, circular dichroism and cell toxicity assays revealed that all variants are less fibrillogenic and less toxic than native hIAPP both at neutral pH and at low pH. This demonstrates that the effect of H18 in native IAPP is not simply determined by its charge state, but rather that residue 18 is important for specific intra- and intermolecular interactions that occur during fibril formation and that may involve charge, size and hydrophobicity. Furthermore, our results indicate that H18R-IAPP has a strong inhibiting effect on native hIAPP fibril formation. Together these results highlight the large impact of modifying a single residue outside the amyloidogenic domain on fibril formation and cell toxicity induced by IAPP, opening up new avenues for design of inhibitors or modulators of IAPP aggregation. Copyright © 2017. Published by Elsevier B.V.

  13. Identification of the ectonucleotidases expressed in mouse, rat, and human Langerhans islets: potential role of NTPDase3 in insulin secretion.

    Science.gov (United States)

    Lavoie, Elise G; Fausther, Michel; Kauffenstein, Gilles; Kukulski, Filip; Künzli, Beat M; Friess, Helmut; Sévigny, Jean

    2010-10-01

    Extracellular nucleotides and adenosine regulate endocrine pancreatic functions such as insulin secretion by Langerhans islet β-cells via the activation of specific P2 and P1 receptors. Membrane-bound ectonucleotidases regulate the local concentration of these ligands and consequently control the activation of their receptors. The objective of this study was to identify and localize the major ectonucleotidases, namely NTPDases and ecto-5'-nucleotidase, present in the endocrine pancreas. In addition, the potential implication of ecto-ATPase activity on insulin secretion was investigated in the rat β-cell line INS-1 (832/13). The localization of ectonucleotidase activity and protein was carried out in situ by enzyme histochemistry and immunolocalization in mouse, rat, and human pancreas sections. NTPDase1 was localized in all blood vessels and acini, and NTPDase2 was localized in capillaries of Langerhans islets and in peripheral conjunctive tissue, whereas NTPDase3 was detected in all Langerhans islet cell types. Interestingly, among the mammalian species tested, ecto-5'-nucleotidase was present only in rat Langerhans islet cells, where it was coexpressed with NTPDase3. Notably, the inhibition of NTPDase3 activity by BG0136 and NF279 facilitated insulin release from INS-1 (832/13) cells under conditions of low glycemia, probably by affecting P2 receptor activation. NTPDase3 activity also regulated the inhibitory effect of exogenous ATP in the presence of a high glucose concentration most likely by controlling adenosine production. In conclusion, all pancreatic endocrine cells express NTPDase3 that was shown to modulate insulin secretion in rat INS-1 (832/13) β-cells. Ecto-5'-nucleotidase is expressed in rat Langerhans islet cells but absent in human and mouse endocrine cells.

  14. A novel Gymnema sylvestre extract stimulates insulin secretion from human islets in vivo and in vitro.

    Science.gov (United States)

    Al-Romaiyan, A; Liu, B; Asare-Anane, H; Maity, C R; Chatterjee, S K; Koley, N; Biswas, T; Chatterji, A K; Huang, G-C; Amiel, S A; Persaud, S J; Jones, P M

    2010-09-01

    Many plant-based products have been suggested as potential antidiabetic agents, but few have been shown to be effective in treating the symptoms of Type 2 diabetes mellitus (T2DM) in human studies, and little is known of their mechanisms of action. Extracts of Gymnema sylvestre (GS) have been used for the treatment of T2DM in India for centuries. The effects of a novel high molecular weight GS extract, Om Santal Adivasi, (OSA(R)) on plasma insulin, C-peptide and glucose in a small cohort of patients with T2DM are reported here. Oral administration of OSA(R) (1 g/day, 60 days) induced significant increases in circulating insulin and C-peptide, which were associated with significant reductions in fasting and post-prandial blood glucose. In vitro measurements using isolated human islets of Langerhans demonstrated direct stimulatory effects of OSA(R) on insulin secretion from human ß-cells, consistent with an in vivo mode of action through enhancing insulin secretion. These in vivo and in vitro observations suggest that OSA(R) may provide a potential alternative therapy for the hyperglycemia associated with T2DM.

  15. Regional differences in islet distribution in the human pancreas--preferential beta-cell loss in the head region in patients with type 2 diabetes.

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    Xiaojun Wang

    Full Text Available While regional heterogeneity in islet distribution has been well studied in rodents, less is known about human pancreatic histology. To fill gaps in our understanding, regional differences in the adult human pancreas were quantitatively analyzed including the pathogenesis of type 2 diabetes (T2D. Cadaveric pancreas specimens were collected from the head, body and tail regions of each donor, including subjects with no history of diabetes or pancreatic diseases (n = 23 as well as patients with T2D (n = 12. The study further included individuals from whom islets were isolated (n = 7 to study islet yield and function in a clinical setting of islet transplantation. The whole pancreatic sections were examined using an innovative large-scale image capture and unbiased detailed quantitative analyses of the characteristics of islets from each individual (architecture, size, shape and distribution. Islet distribution/density is similar between the head and body regions, but is >2-fold higher in the tail region. In contrast to rodents, islet cellular composition and architecture were similar throughout the pancreas and there was no difference in glucose-stimulated insulin secretion in islets isolated from different regions of the pancreas. Further studies revealed preferential loss of large islets in the head region in patients with T2D. The present study has demonstrated distinct characteristics of the human pancreas, which should provide a baseline for the future studies integrating existing research in the field and helping to advance bi-directional research between humans and preclinical models.

  16. Human Islet Amyloid Polypeptide Fibril Binding to Catalase: A Transmission Electron Microscopy and Microplate Study

    Directory of Open Access Journals (Sweden)

    Nathaniel G. N. Milton

    2010-01-01

    Full Text Available The diabetes-associated human islet amyloid polypeptide (IAPP is a 37-amino-acid peptide that forms fibrils in vitro and in vivo. Human IAPP fibrils are toxic in a similar manner to Alzheimer's amyloid-β (Aβ and prion protein (PrP fibrils. Previous studies have shown that catalase binds to Aβ fibrils and appears to recognize a region containing the Gly-Ala-Ile-Ile sequence that is similar to the Gly-Ala-Ile-Leu sequence found in human IAPP residues 24-27. This study presents a transmission electron microscopy (TEM—based analysis of fibril formation and the binding of human erythrocyte catalase to IAPP fibrils. The results show that human IAPP 1-37, 8-37, and 20-29 peptides form fibrils with diverse and polymorphic structures. All three forms of IAPP bound catalase, and complexes of IAPP 1-37 or 8-37 with catalase were identified by immunoassay. The binding of biotinylated IAPP to catalase was high affinity with a KD of 0.77nM, and could be inhibited by either human or rat IAPP 1-37 and 8-37 forms. Fibrils formed by the PrP 118-135 peptide with a Gly-Ala-Val-Val sequence also bound catalase. These results suggest that catalase recognizes a Gly-Ala-Ile-Leu—like sequence in amyloid fibril-forming peptides. For IAPP 1-37 and 8-37, the catalase binding was primarily directed towards fibrillar rather than ribbon-like structures, suggesting differences in the accessibility of the human IAPP 24-27 Gly-Ala-Ile-Leu region. This suggests that catalase may be able to discriminate between different structural forms of IAPP fibrils. The ability of catalase to bind IAPP, Aβ, and PrP fibrils demonstrates the presence of similar accessible structural motifs that may be targets for antiamyloid therapeutic development.

  17. Trefoil factor 3 stimulates human and rodent pancreatic islet beta-cell replication with retention of function.

    Science.gov (United States)

    Fueger, Patrick T; Schisler, Jonathan C; Lu, Danhong; Babu, Daniella A; Mirmira, Raghavendra G; Newgard, Christopher B; Hohmeier, Hans E

    2008-05-01

    Both major forms of diabetes involve a decline in beta-cell mass, mediated by autoimmune destruction of insulin-producing cells in type 1 diabetes and by increased rates of apoptosis secondary to metabolic stress in type 2 diabetes. Methods for controlled expansion of beta-cell mass are currently not available but would have great potential utility for treatment of these diseases. In the current study, we demonstrate that overexpression of trefoil factor 3 (TFF3) in rat pancreatic islets results in a 4- to 5-fold increase in [(3)H]thymidine incorporation, with full retention of glucose-stimulated insulin secretion. This increase was almost exclusively due to stimulation of beta-cell replication, as demonstrated by studies of bromodeoxyuridine incorporation and co-immunofluorescence analysis with anti-bromodeoxyuridine and antiinsulin or antiglucagon antibodies. The proliferative effect of TFF3 required the presence of serum or 0.5 ng/ml epidermal growth factor. The ability of TFF3 overexpression to stimulate proliferation of rat islets in serum was abolished by the addition of epidermal growth factor receptor antagonist AG1478. Furthermore, TFF3-induced increases in [3H]thymidine incorporation in rat islets cultured in serum was blocked by overexpression of a dominant-negative Akt protein or treatment with triciribine, an Akt inhibitor. Finally, overexpression of TFF3 also caused a doubling of [3H]thymidine incorporation in human islets. In summary, our findings reveal a novel TFF3-mediated pathway for stimulation of beta-cell replication that could ultimately be exploited for expansion or preservation of islet beta-cell mass.

  18. Three-dimensional Reconstruction of the Microstructure of Human Acellular Nerve Allograft

    Science.gov (United States)

    Zhu, Shuang; Zhu, Qingtang; Liu, Xiaolin; Yang, Weihong; Jian, Yutao; Zhou, Xiang; He, Bo; Gu, Liqiang; Yan, Liwei; Lin, Tao; Xiang, Jianping; Qi, Jian

    2016-01-01

    The exact inner 3D microstructure of the human peripheral nerve has been a mystery for decades. Therefore, it has been difficult to solve several problems regarding peripheral nerve injury and repair. We used high-resolution X-ray computed microtomography (microCT) to scan a freeze-dried human acellular nerve allograft (hANA). The microCT images were then used to reconstruct a 3D digital model, which was used to print a 3D resin model of the nerve graft. The 3D digital model of the hANA allowed visualization of all planes. The magnified 3D resin model clearly showed the nerve bundles and basement membrane tubes of the hANA. Scanning electron microscopy (SEM) was used to analyse the microstructure of the hANA. Compared to the SEM images, the microCT image clearly demonstrated the microstructure of the hANA cross section at a resolution of up to 1.2 μm. The 3D digital model of the hANA facilitates a clear and easy understanding of peripheral nerve microstructure. Furthermore, the enlarged 3D resin model duplicates the unique inner structure of each individual hANA. This is a crucial step towards achieving 3D printing of a hANA or nerve that can be used as a nerve graft. PMID:27476584

  19. Pancreatic islet blood flow and its measurement.

    Science.gov (United States)

    Jansson, Leif; Barbu, Andreea; Bodin, Birgitta; Drott, Carl Johan; Espes, Daniel; Gao, Xiang; Grapensparr, Liza; Källskog, Örjan; Lau, Joey; Liljebäck, Hanna; Palm, Fredrik; Quach, My; Sandberg, Monica; Strömberg, Victoria; Ullsten, Sara; Carlsson, Per-Ola

    2016-05-01

    Pancreatic islets are richly vascularized, and islet blood vessels are uniquely adapted to maintain and support the internal milieu of the islets favoring normal endocrine function. Islet blood flow is normally very high compared with that to the exocrine pancreas and is autonomously regulated through complex interactions between the nervous system, metabolites from insulin secreting β-cells, endothelium-derived mediators, and hormones. The islet blood flow is normally coupled to the needs for insulin release and is usually disturbed during glucose intolerance and overt diabetes. The present review provides a brief background on islet vascular function and especially focuses on available techniques to measure islet blood perfusion. The gold standard for islet blood flow measurements in experimental animals is the microsphere technique, and its advantages and disadvantages will be discussed. In humans there are still no methods to measure islet blood flow selectively, but new developments in radiological techniques hold great hopes for the future.

  20. Disinfection of human skin allografts in tissue banking: a systematic review report.

    Science.gov (United States)

    Johnston, C; Callum, J; Mohr, J; Duong, A; Garibaldi, A; Simunovic, N; Ayeni, O R

    2016-12-01

    The use of skin allografts to temporarily replace lost or damaged skin is practiced worldwide. Naturally occurring contamination can be present on skin or can be introduced at recovery or during processing. This contamination can pose a threat to allograft recipients. Bacterial culture and disinfection of allografts are mandated, but the specific practices and methodologies are not dictated by standards. A systematic review of literature from three databases found 12 research articles that evaluated bioburden reduction processes of skin grafts. The use of broad spectrum antibiotics and antifungal agents was the most frequently identified disinfection method reported demonstrating reductions in contamination rates. It was determined that the greatest reduction in the skin allograft contamination rates utilized 0.1 % peracetic acid or 25 kGy of gamma irradiation at lower temperatures.

  1. Selective destruction of mouse islet beta cells by human T lymphocytes in a newly-established humanized type 1 diabetic model

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Yong, E-mail: yongzhao@uic.edu [Department of Medicine, University of Illinois at Chicago, Chicago, IL 60612 (United States); Guo, Chengshan; Hwang, David; Lin, Brian; Dingeldein, Michael; Mihailescu, Dan; Sam, Susan; Sidhwani, Seema [Department of Medicine, University of Illinois at Chicago, Chicago, IL 60612 (United States); Zhang, Yongkang [Department of Pharmacology, University of Illinois at Chicago, Chicago, IL 60612 (United States); Jain, Sumit [Department of Medicine, University of Illinois at Chicago, Chicago, IL 60612 (United States); Skidgel, Randal A. [Department of Pharmacology, University of Illinois at Chicago, Chicago, IL 60612 (United States); Prabhakar, Bellur S. [Department of Immunology and Microbiology, University of Illinois at Chicago, Chicago, IL 60612 (United States); Mazzone, Theodore [Department of Medicine, University of Illinois at Chicago, Chicago, IL 60612 (United States); Holterman, Mark J. [Department of Surgery, University of Illinois at Chicago, Chicago, IL 60612 (United States)

    2010-09-03

    Research highlights: {yields} Establish a human immune-mediated type 1 diabetic model in NOD-scid IL2r{gamma}{sup null} mice. {yields} Using the irradiated diabetic NOD mouse spleen mononuclear cells as trigger. {yields} The islet {beta} cells were selectively destroyed by infiltrated human T cells. {yields} The model can facilitate translational research to find a cure for type 1 diabetes. -- Abstract: Type 1 diabetes (T1D) is caused by a T cell-mediated autoimmune response that leads to the loss of insulin-producing {beta} cells. The optimal preclinical testing of promising therapies would be aided by a humanized immune-mediated T1D model. We develop this model in NOD-scid IL2r{gamma}{sup null} mice. The selective destruction of pancreatic islet {beta} cells was mediated by human T lymphocytes after an initial trigger was supplied by the injection of irradiated spleen mononuclear cells (SMC) from diabetic nonobese diabetic (NOD) mice. This resulted in severe insulitis, a marked loss of total {beta}-cell mass, and other related phenotypes of T1D. The migration of human T cells to pancreatic islets was controlled by the {beta} cell-produced highly conserved chemokine stromal cell-derived factor 1 (SDF-1) and its receptor C-X-C chemokine receptor (CXCR) 4, as demonstrated by in vivo blocking experiments using antibody to CXCR4. The specificity of humanized T cell-mediated immune responses against islet {beta} cells was generated by the local inflammatory microenvironment in pancreatic islets including human CD4{sup +} T cell infiltration and clonal expansion, and the mouse islet {beta}-cell-derived CD1d-mediated human iNKT activation. The selective destruction of mouse islet {beta} cells by a human T cell-mediated immune response in this humanized T1D model can mimic those observed in T1D patients. This model can provide a valuable tool for translational research into T1D.

  2. The human pancreatic islet transcriptome: expression of candidate genes for type 1 diabetes and the impact of pro-inflammatory cytokines.

    Directory of Open Access Journals (Sweden)

    Décio L Eizirik

    Full Text Available Type 1 diabetes (T1D is an autoimmune disease in which pancreatic beta cells are killed by infiltrating immune cells and by cytokines released by these cells. Signaling events occurring in the pancreatic beta cells are decisive for their survival or death in diabetes. We have used RNA sequencing (RNA-seq to identify transcripts, including splice variants, expressed in human islets of Langerhans under control conditions or following exposure to the pro-inflammatory cytokines interleukin-1β (IL-1β and interferon-γ (IFN-γ. Based on this unique dataset, we examined whether putative candidate genes for T1D, previously identified by GWAS, are expressed in human islets. A total of 29,776 transcripts were identified as expressed in human islets. Expression of around 20% of these transcripts was modified by pro-inflammatory cytokines, including apoptosis- and inflammation-related genes. Chemokines were among the transcripts most modified by cytokines, a finding confirmed at the protein level by ELISA. Interestingly, 35% of the genes expressed in human islets undergo alternative splicing as annotated in RefSeq, and cytokines caused substantial changes in spliced transcripts. Nova1, previously considered a brain-specific regulator of mRNA splicing, is expressed in islets and its knockdown modified splicing. 25/41 of the candidate genes for T1D are expressed in islets, and cytokines modified expression of several of these transcripts. The present study doubles the number of known genes expressed in human islets and shows that cytokines modify alternative splicing in human islet cells. Importantly, it indicates that more than half of the known T1D candidate genes are expressed in human islets. This, and the production of a large number of chemokines and cytokines by cytokine-exposed islets, reinforces the concept of a dialog between pancreatic islets and the immune system in T1D. This dialog is modulated by candidate genes for the disease at both the

  3. ISLET FORMATION AND REGENERATION

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Objective To explore the mechanisms of differentiation and development of pancreatic endocrine cells as well as pancreatic regeneration. Methods Human embryonic pancreatic tissue at 7-14 weeks of gestation was collected. Diabetes mellitus rat model was induced with 65 mg/kg of streptozotocin. Insulin, glucagon, somatostatin, nestin, and cytokeratin 19 (CK19)of pancreatic tissues were observed by immunohistochemistry. Results At 9 weeks of gestation, pancreatic epithelial cells began to co-express insulin, glucagon, somatostatin, and CK19 before migration. Islet cells gradually congregated along with the increase of aging, and at 14 weeks of gestation histological examination showed islet formation. At 12 weeks of gestation, nestin-positive cells could be seen in the pancreatic mesenchyme. During early embryogenesis, islet cells of pancreatic ducts co-expressed insulin, glucagon, and somatostatin. During pancreatic regeneration after damage, nestin expression of islet cells increased. Conclusion In the early stage of embryogenesis, islet cells of primary pancreatic ducts can be differentiated to multipotential endocrine cells before migration. During tissue regeneration, pancreatic stem cells may differentiate and proliferate to form pancreatic islet.

  4. Islet neogenesis potential of human adult stem cells and its applications in cell replacement therapy for diabetes

    Directory of Open Access Journals (Sweden)

    Bhonde RR

    2008-11-01

    Full Text Available In recent years regenerative biology has reached to greater heights due to its therapeutic potential in treating degenerative diseases; as they are not curable by modern medicine. With the advent of research in stem cells and developmental biology the regenerative potential of adult resident stem cells is becoming clearer. The long term objective of regenerative medicine or cell therapy is to treat patients with their own stem cells. These stem cells could be derived from the diseased organs such as skin, liver, pancreas etc. or from reservoirs of multipotent stem cells such as bone marrow or cord blood.Manipulating the ability of tissue resident stem cells as well as from multipotent reservoirs such as bone marrow, umbilical cord and cord blood to give rise to endocrine cells may open new avenues in the treatment of diabetes. A better understanding of stem cell biology would almost certainly allow for the establishment of efficient and reliable cell transplantation experimental programs in the clinic. We show here that multipotent mesenchymal stem cells can be isolated from various sources such as the bone marrow, placenta, umbilical cord. Upon stimulation with specific growth factors they differentiate into islet like clusters (ILCs. When ILCs obtained from the above mentioned sources were transplanted in experimental diabetic mice, restoration of normoglycemia was observed within three weeks of transplantation with concomitant increase in the body weight. These euglycemic mice exhibited normal glucose tolerance test indicating normal utilization of glucose. Allthough the MSCs isolated from all the sources had the same characteristics; they showed significant differences in their islet differentiation potential. ILCs isolated for the human bone marrow did not show any pancreatic hormones in vitro, but upon transplantation they matured into insulin and somatostatin producing hormones. Placental MSCs as well as ILCs showed insulin trascripts

  5. Glucose-induced β cell production of IL-1β contributes to glucotoxicity in human pancreatic islets

    Science.gov (United States)

    Maedler, Kathrin; Sergeev, Pavel; Ris, Frédéric; Oberholzer, José; Joller-Jemelka, Helen I.; Spinas, Giatgen A.; Kaiser, Nurit; Halban, Philippe A.; Donath, Marc Y.

    2002-01-01

    In type 2 diabetes, chronic hyperglycemia is suggested to be detrimental to pancreatic β cells, causing impaired insulin secretion. IL-1β is a proinflammatory cytokine acting during the autoimmune process of type 1 diabetes. IL-1β inhibits β cell function and promotes Fas-triggered apoptosis in part by activating the transcription factor NF-κB. Recently, we have shown that increased glucose concentrations also induce Fas expression and β cell apoptosis in human islets. The aim of the present study was to test the hypothesis that IL-1β may mediate the deleterious effects of high glucose on human β cells. In vitro exposure of islets from nondiabetic organ donors to high glucose levels resulted in increased production and release of IL-1β, followed by NF-κB activation, Fas upregulation, DNA fragmentation, and impaired β cell function. The IL-1 receptor antagonist protected cultured human islets from these deleterious effects. β cells themselves were identified as the islet cellular source of glucose-induced IL-1β. In vivo, IL-1β–producing β cells were observed in pancreatic sections of type 2 diabetic patients but not in nondiabetic control subjects. Similarly, IL-1β was induced in β cells of the gerbil Psammomys obesus during development of diabetes. Treatment of the animals with phlorizin normalized plasma glucose and prevented β cell expression of IL-1β. These findings implicate an inflammatory process in the pathogenesis of glucotoxicity in type 2 diabetes and identify the IL-1β/NF-κB pathway as a target to preserve β cell mass and function in this condition. PMID:12235117

  6. Allograft inflammatory factor-1 stimulates chemokine production and induces chemotaxis in human peripheral blood mononuclear cells.

    Science.gov (United States)

    Kadoya, Masatoshi; Yamamoto, Aihiro; Hamaguchi, Masahide; Obayashi, Hiroshi; Mizushima, Katsura; Ohta, Mitsuhiro; Seno, Takahiro; Oda, Ryo; Fujiwara, Hiroyoshi; Kohno, Masataka; Kawahito, Yutaka

    2014-06-06

    Allograft inflammatory factor-1 (AIF-1) is expressed by macrophages, fibroblasts, endothelial cells and smooth muscle cells in immune-inflammatory disorders such as systemic sclerosis, rheumatoid arthritis and several vasculopathies. However, its molecular function is not fully understood. In this study, we examined gene expression profiles and induction of chemokines in monocytes treated with recombinant human AIF (rhAIF-1). Using the high-density oligonucleotide microarray technique, we compared mRNA expression profiles of rhAIF-1-stimulated CD14(+) peripheral blood mononuclear cells (CD14(+) PBMCs) derived from healthy volunteers. We demonstrated upregulation of genes for several CC chemokines such as CCL1, CCL2, CCL3, CCL7, and CCL20. Next, using ELISAs, we confirmed that rhAIF-1 promoted the secretion of CCL3/MIP-1α and IL-6 by CD14(+) PBMCs, whereas only small amounts of CCL1, CCL2/MCP-1, CCL7/MCP-3 and CCL20/MIP-3α were secreted. Conditioned media from rhAIF-1stimulated CD14(+) PBMCs resulted in migration of PBMCs. These findings suggest that AIF-1, which induced chemokines and enhanced chemotaxis of monocytes, may represent a molecular target for the therapy of immune-inflammatory disorders. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Islet formation in mice and men: lessons for the generation of functional insulin-producing β-cells from human pluripotent stem cells.

    Science.gov (United States)

    Nair, Gopika; Hebrok, Matthias

    2015-06-01

    The Islets of Langerhans are crucial 'micro-organs' embedded in the glandular exocrine pancreas that regulate nutrient metabolism. They not only synthesize, but also secrete endocrine hormones in a modulated fashion in response to physiologic metabolic demand. These highly sophisticated structures with intricate organization of multiple cell types, namely endocrine, vascular, neuronal and mesenchymal cells, have evolved to perform this task to perfection over time. Not surprisingly, islet architecture and function are dissimilar between humans and typically studied model organisms, such as rodents and zebrafish. Further, recent findings also suggest noteworthy differences in human islet development from that in mouse, including delayed appearance and gradual resolution of key differentiation markers, a single-phase of endocrine differentiation, and prenatal association of developing islets with neurovascular milieu. In light of these findings, it is imperative that a systematic study is undertaken to compare islet development between human and mouse. Illuminating inter-species differences in islet development will likely be critical in furthering our pursuit to generate an unlimited supply of truly functional and fully mature β-cells from human pluripotent stem cell (hPSC) sources for therapeutic purposes.

  8. Influence of Aluminium and EGCG on Fibrillation and Aggregation of Human Islet Amyloid Polypeptide

    Science.gov (United States)

    Xu, Zhi-Xue; Zhang, Qiang; Ma, Gong-Li; Chen, Cong-Heng; He, Yan-Ming; Xu, Li-Hui; Zhang, Yuan; Zhou, Guang-Rong; Li, Zhen-Hua

    2016-01-01

    The abnormal fibrillation of human islet amyloid polypeptide (hIAPP) has been implicated in the development of type II diabetes. Aluminum is known to trigger the structural transformation of many amyloid proteins and induce the formation of toxic aggregate species. The (−)-epigallocatechin gallate (EGCG) is considered capable of binding both metal ions and amyloid proteins with inhibitory effect on the fibrillation of amyloid proteins. However, the effect of Al(III)/EGCG complex on hIAPP fibrillation is unclear. In the present work, we sought to view insight into the structures and properties of Al(III) and EGCG complex by using spectroscopic experiments and quantum chemical calculations and also investigated the influence of Al(III) and EGCG on hIAPP fibrillation and aggregation as well as their combined interference on this process. Our studies demonstrated that Al(III) could promote fibrillation and aggregation of hIAPP, while EGCG could inhibit the fibrillation of hIAPP and lead to the formation of hIAPP amorphous aggregates instead of the ordered fibrils. Furthermore, we proved that the Al(III)/EGCG complex in molar ratio of 1 : 1 as Al(EGCG)(H2O)2 could inhibit the hIAPP fibrillation more effectively than EGCG alone. The results provide the invaluable reference for the new drug development to treat type II diabetes. PMID:28074190

  9. Influence of Aluminium and EGCG on Fibrillation and Aggregation of Human Islet Amyloid Polypeptide

    Directory of Open Access Journals (Sweden)

    Zhi-Xue Xu

    2016-01-01

    Full Text Available The abnormal fibrillation of human islet amyloid polypeptide (hIAPP has been implicated in the development of type II diabetes. Aluminum is known to trigger the structural transformation of many amyloid proteins and induce the formation of toxic aggregate species. The (−-epigallocatechin gallate (EGCG is considered capable of binding both metal ions and amyloid proteins with inhibitory effect on the fibrillation of amyloid proteins. However, the effect of Al(III/EGCG complex on hIAPP fibrillation is unclear. In the present work, we sought to view insight into the structures and properties of Al(III and EGCG complex by using spectroscopic experiments and quantum chemical calculations and also investigated the influence of Al(III and EGCG on hIAPP fibrillation and aggregation as well as their combined interference on this process. Our studies demonstrated that Al(III could promote fibrillation and aggregation of hIAPP, while EGCG could inhibit the fibrillation of hIAPP and lead to the formation of hIAPP amorphous aggregates instead of the ordered fibrils. Furthermore, we proved that the Al(III/EGCG complex in molar ratio of 1 : 1 as Al(EGCG(H2O2 could inhibit the hIAPP fibrillation more effectively than EGCG alone. The results provide the invaluable reference for the new drug development to treat type II diabetes.

  10. Functional proteomics screen enables enrichment of distinct cell types from human pancreatic islets.

    Directory of Open Access Journals (Sweden)

    Revital Sharivkin

    Full Text Available The current world-wide epidemic of diabetes has prompted attempts to generate new sources of insulin-producing cells for cell replacement therapy. An inherent challenge in many of these strategies is the lack of cell-surface markers permitting isolation and characterization of specific cell types from differentiating stem cell populations. Here we introduce an iterative proteomics procedure allowing tag-free isolation of cell types based on their function. Our method detects and associates specific cell-surface markers with particular cell functionality by coupling cell capture on antibody arrays with immunofluorescent labeling. Using this approach in an iterative manner, we discovered marker combinations capable of enriching for discrete pancreatic cell subtypes from human islets of Langerhans: insulin-producing beta cells (CD9high/CD56+, glucagon-producing alpha cells (CD9-/CD56+ and trypsin-producing acinar cells (CD9-/CD56-. This strategy may assist future beta cell research and the development of diagnostic tools for diabetes. It can also be applied more generally for function-based purification of desired cell types from other limited and heterogeneous biological samples.

  11. The use of a biostatic fascia lata thigh allograft as a scaffold for autologous human culture of fibroblasts--An in vitro study.

    Science.gov (United States)

    Żurek, Jarek; Dominiak, Marzena; Botzenhart, Ute; Bednarz, Wojciech

    2015-05-01

    The method for covering gingival recession defects and augmenting keratinized gingiva involves the use of autogenuous connective tissue grafts obtained from palatal mucosa in combination with various techniques of flap repositioning or tunnel techniques. In the case of multiple gingival recession defects the amount of connective tissue available for grafting is insufficient. Therefore, the use of substitutes is necessary. The most widely used material in recent years has been the acellular dermal matrix allograft. The disadvantage of its application lies in the absence of cells and blood vessels, which increases incorporation time. Primary cultured human autologic fibroblasts are commonly used to optimize the healing process. The aim of this study was to examine the in vitro biocompatibility of human fascia lata allograft as a new scaffold for primary cultured human autologic fibroblasts. For that, a fibroblast culture obtained from a fragment of gingival tissue taken from the hard palate mucosa of a subject was used. After 14 days the colony cells were inoculated on a fragment of human fascia lata allograft. After a further 7 days of incubation the material was frozen, cut and prepared for histochemical examination. After two weeks of incubation, and 7 days after inoculation on a fragment of fascia lata allograft numerous accumulations of the cultured fibroblast were found that had a typical structure and produced collagen fibres. A human fascia lata allograft can be used as a scaffold for primary cultured human autologic fibroblasts. Further studies should confirm the clinical efficacy of this solution.

  12. Determination of mechanical properties of impacted human morsellized cancellous allografts for revision joint arthroplasty.

    Science.gov (United States)

    Tanabe, Y; Wakui, T; Kobayashi, A; Ohashi, H; Kadoya, Y; Yamano, Y

    1999-12-01

    This paper deals with the characterization of mechanical properties of impacted morsellized cancellous allograft (IMCA) produced by dynamic compaction of allograft femoral heads ground by commercially available bone mills, i.e. rotating rasp and reciprocating type bone mills. Various ranges and profiles of particle size in the graft aggregates were obtained using these bone mills, and the effect of number of compaction as well as the distribution of particle sizes on the mechanical properties of IMCA under quasistatic compression and shear loading conditions was discussed. The morsellized cancellous allograft prepared by the reciprocating type bone mill showed a broad distribution of particle sizes, and gave IMCA superior mechanical properties to the graft with a more uniform size distribution, or prepared by the rotating rasp type bone mills. The increase of number of compaction also improved the mechanical properties of IMCA in compression.

  13. Protection of Human Pancreatic Islets from Lipotoxicity by Modulation of the Translocon.

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    R Cassel

    Full Text Available Type 2 diabetes is characterized by peripheral insulin resistance and pancreatic beta cell dysfunction. Elevated free fatty acids (FFAs may impair beta cell function and mass (lipotoxicity. Altered calcium homeostasis may be involved in defective insulin release. The endoplasmic reticulum (ER is the major intracellular calcium store. Lipotoxicity induces ER stress and in parallel an ER calcium depletion through unknown ER calcium leak channels. The main purposes of this study is first to identify one of these channels and secondly, to check the opportunity to restore beta cells function (i.e., insulin secretion after pharmacological inhibition of ER calcium store depletion. We investigated the functionality of translocon, an ER calcium leak channel and its involvement on FFAs-induced alterations in MIN6B1 cells and in human pancreatic islets. We evidenced that translocon acts as a functional ER calcium leak channel in human beta cells using anisomycin and puromycin (antibiotics, respectively blocker and opener of this channel. Puromycin induced a significant ER calcium release, inhibited by anisomycin pretreatment. Palmitate treatment was used as FFA model to induce a mild lipotoxic effect: ER calcium content was reduced, ER stress but not apoptosis were induced and glucose induced insulin secretion was decreased in our beta cells. Interestingly, translocon inhibition by chronic anisomycin treatment prevented dysfunctions induced by palmitate, avoiding reticular calcium depletion, ER stress and restoring insulin secretion. Our results provide for the first time compelling evidence that translocon actively participates to the palmitate-induced ER calcium leak and insulin secretion decrease in beta cells. Its inhibition reduces these lipotoxic effects. Taken together, our data indicate that TLC may be a new potential target for the treatment of type 2 diabetes.

  14. Identification of a novel human islet amyloid polypeptide beta-sheet domain and factors influencing fibrillogenesis.

    Science.gov (United States)

    Jaikaran, E T; Higham, C E; Serpell, L C; Zurdo, J; Gross, M; Clark, A; Fraser, P E

    2001-05-04

    Human islet amyloid polypeptide (hIAPP) accumulates as pancreatic amyloid in type 2 diabetes and readily forms fibrils in vitro. Investigations into the mechanism of hIAPP fibril formation have focused largely on residues 20 to 29, which are considered to comprise a primary amyloidogenic domain. In rodents, proline substitutions within this region and the subsequent beta-sheet disruption, prevents fibril formation. An additional amyloidogenic fragment within the C-terminal sequence, residues 30 to 37, has been identified recently. We have extended these observations by examining a series of overlapping peptide fragments from the human and rodent sequences. Using protein spectroscopy (CD/FTIR), electron microscopy and X-ray diffraction, a previously unrecognised amyloidogenic domain was localised within residues 8 to 20. Synthetic peptides corresponding to this region exhibited a transition from random coil to beta-sheet conformation and assembled into fibrils having a typical amyloid-like morphology. The comparable rat 8-20 sequence, which contains a single His18Arg substitution, was also capable of assembling into amyloid-like fibrils. Examination of peptide fragments corresponding to residues 1 to 13 revealed that the immediate N-terminal region is likely to have only a modulating influence on fibril formation or conformational conversion. The contributions of charged residues as they relate to the amyloid-forming 8-20 sequence were also investigated using IAPP fragments and by assessing the effects of pH and counterions. The identification of these principal amyloidogenic sequences and the effects of associated factors provide details on the IAPP aggregation pathway and structure of the peptide in its fibrillar state. Copyright 2001 Academic Press.

  15. Downregulation of Type II Diabetes Mellitus and Maturity Onset Diabetes of Young Pathways in Human Pancreatic Islets from Hyperglycemic Donors

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    Jalal Taneera

    2014-01-01

    Full Text Available Although several molecular pathways have been linked to type 2 diabetes (T2D pathogenesis, it is uncertain which pathway has the most implication on the disease. Changes in the expression of an entire pathway might be more important for disease pathogenesis than changes in the expression of individual genes. To identify the molecular alterations in T2D, DNA microarrays of human pancreatic islets from donors with hyperglycemia n=20 and normoglycemia n=58 were subjected to Gene Set Enrichment Analysis (GSEA. About 178 KEGG pathways were investigated for gene expression changes between hyperglycemic donors compared to normoglycemic. Pathway enrichment analysis showed that type II diabetes mellitus (T2DM and maturity onset diabetes of the young (MODY pathways are downregulated in hyperglycemic donors, while proteasome and spliceosome pathways are upregulated. The mean centroid of gene expression of T2DM and MODY pathways was shown to be associated positively with insulin secretion and negatively with HbA1c level. To conclude, downregulation of T2DM and MODY pathways is involved in islet function and might be involved in T2D. Also, the study demonstrates that gene expression profiles from pancreatic islets can reveal some of the biological processes related to regulation of glucose hemostats and diabetes pathogenesis.

  16. Pancreatic islet transplantation

    Directory of Open Access Journals (Sweden)

    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

  17. Monocyte procoagulant activity and plasminogen activator. Role in human renal allograft rejection

    Energy Technology Data Exchange (ETDEWEB)

    Cole, E.H.; Cardella, C.J.; Schulman, J.; Levy, G.A.

    1985-10-01

    Currently the mechanism of renal allograft rejection is not well understood. This study was designed to determine whether induction of monocyte procoagulant activity (MCPA) is important in the pathogenesis of renal allograft rejection. The MPCA assay was performed utilizing a one stage clotting assay both in normal and in factor-VII-deficient plasma. There was no increase in spontaneous MPCA in 20 patients with endstage renal failure and in 10 patients following abdominal or orthopedic operation, as compared with 20 normal controls. MPCA was assessed daily in 18 patients who had received renal allografts. Rejection episodes (RE) were predicted on the basis of persistent elevation in MPCA as compared with pretransplant levels. Rejection was diagnosed clinically and treated on the basis of standard criteria. Treated RE were compared with those predicted by elevated MPCA, and 3 patients were assessed as having no RE by MPCA and by standard criteria. In 8 RE, MPCA correlated temporally with RE (same day) when compared with standard criteria. In 12 RE, MPCA was predictive of rejection preceding standard criteria by at least 24 hr. There were 7 false-positive predictions on the basis of MPCA; however, there was only 1 false negative. MPCA was shown to be a prothrombinase by its dependence only on prothrombin and fibrinogen for full activity. MPCA may be important in the pathogenesis of allograft rejection, and additionally it may be a useful adjunct in the clinical management of this disease.

  18. Islet-like clusters derived from mesenchymal stem cells in Wharton's Jelly of the human umbilical cord for transplantation to control type 1 diabetes.

    Directory of Open Access Journals (Sweden)

    Kuo Ching Chao

    Full Text Available BACKGROUND: There is a widespread interest in developing renewable sources of islet-replacement tissue for type I diabetes mellitus. Human mesenchymal cells isolated from the Wharton's jelly of the umbilical cord (HUMSCs, which can be easily obtained and processed compared with embryonic and bone marrow stem cells, possess stem cell properties. HUMSCs may be a valuable source for the generation of islets. METHODOLOGY AND PRINCIPAL FINDINGS: HUMSCs were induced to transform into islet-like cell clusters in vitro through stepwise culturing in neuron-conditioned medium. To assess the functional stability of the islet-like cell clusters in vivo, these cell clusters were transplanted into the liver of streptozotocin-induced diabetic rats via laparotomy. Glucose tolerance was measured on week 12 after transplantation accompanied with immunohistochemistry and electron microscopy analysis. These islet-like cell clusters were shown to contain human C-peptide and release human insulin in response to physiological glucose levels. Real-time RT-PCR detected the expressions of insulin and other pancreatic beta-cell-related genes (Pdx1, Hlxb9, Nkx2.2, Nkx6.1, and Glut-2 in these islet-like cell clusters. The hyperglycemia and glucose intolerance in streptozotocin-induced diabetic rats was significantly alleviated after xenotransplantation of islet-like cell clusters, without the use of immunosuppressants. In addition to the existence of islet-like cell clusters in the liver, some special fused liver cells were also found, which characterized by human insulin and nuclei-positive staining and possessing secretory granules. CONCLUSIONS AND SIGNIFICANCE: In this study, we successfully differentiate HUMSCs into mature islet-like cell clusters, and these islet-like cell clusters possess insulin-producing ability in vitro and in vivo. HUMSCs in Wharton's Jelly of the umbilical cord seem to be the preferential source of stem cells to convert into insulin

  19. Differential transcriptome analysis of diabetes-resistant and -sensitive mouse islets reveals significant overlap with human diabetes susceptibility genes.

    Science.gov (United States)

    Kluth, Oliver; Matzke, Daniela; Schulze, Gunnar; Schwenk, Robert W; Joost, Hans-Georg; Schürmann, Annette

    2014-12-01

    Type 2 diabetes in humans and in obese mice is polygenic. In recent genome-wide association studies, genetic markers explaining a small portion of the genetic contribution to the disease were discovered. However, functional evidence linking these genes with the pathogenesis of diabetes is scarce. We performed RNA sequencing-based transcriptomics of islets from two obese mouse strains, a diabetes-susceptible (NZO) and a diabetes-resistant (B6-ob/ob) mouse, after a short glucose challenge and compared these results with human data. Alignment of 2,328 differentially expressed genes to 106 human diabetes candidate genes revealed an overlap of 20 genes, including TCF7L2, IGFBP2, CDKN2A, CDKN2B, GRB10, and PRC1. The data provide a functional validation of human diabetes candidate genes, including those involved in regulating islet cell recovery and proliferation, and identify additional candidates that could be involved in human β-cell failure. © 2014 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

  20. Site-specific rectocele repair with dermal graft augmentation: comparison of porcine dermal xenograft (Pelvicol) and human dermal allograft.

    Science.gov (United States)

    Biehl, Roger C; Moore, Robert D; Miklos, John R; Kohli, Neeraj; Anand, Indu S; Mattox, T Fleming

    2008-01-01

    This study is a retrospective chart review comparing 195 women who underwent rectocele repair with either a porcine dermal xenograft or human allogenic cadaveric dermal graft augmentation over a two year period. A site-specific defect repair was completed prior to augmentation with the graft. Examinations were performed preoperatively and postoperatively using the pelvic organ prolapse quantification system. Questionnaires were used to assess constipation and dyspareunia. De novo dyspareunia and cure rates for constipation and dyspareunia were not statistically different between the two groups. Site-specific fascial rectocele repairs with xenograft or allograft augmentation were found to have similar complication rates as well as objective and subjective cure rates.

  1. Cytotoxicity of human pI 7 interleukin-1 for pancreatic islets of Langerhans

    DEFF Research Database (Denmark)

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

    1986-01-01

    . This cytotoxic activity was eliminated from crude cytokine preparations by adsorption with immobilized, purified antibody to interleukin-1 (IL-1). The islet-inhibitory activity and the IL-1 activity (determined by its comitogenic effect on thymocytes) were recovered by acid wash. Purified natural IL-1...

  2. KU-32, a Novel Drug for Diabetic Neuropathy, Is Safe for Human Islets and Improves In Vitro Insulin Secretion and Viability

    Directory of Open Access Journals (Sweden)

    Kevin Farmer

    2012-01-01

    Full Text Available KU-32 is a novel, novobiocin-based Hsp90 inhibitor that protects against neuronal glucotoxicity and reverses multiple clinical indices of diabetic peripheral neuropathy in a rodent model. However, any drug with potential for treating diabetic complications must also have no adverse effects on the function of pancreatic islets. Thus, the goal of the current study was to assess the effect of KU-32 on the in vitro viability and function of human islets. Treating human islets with KU-32 for 24 hours showed no toxicity as assessed using the alamarBlue assay. Confocal microscopy confirmed that with a minimum of 2-day exposure, KU-32 improved cellular viability by blocking apoptosis. Functionally, isolated human islets released more glucose-stimulated insulin when preincubated in KU-32. However, diabetic BKS-db/db mice, a model for type 2 diabetes, administered KU-32 for 10 weeks did not show any significant changes in blood glucose and insulin levels, despite having greater insulin staining/beta cell in the pancreas compared to untreated BKS db/db mice. In summary, KU-32 did not harm isolated human islets and may even be protective. However, the effect does not appear significant enough to alter the in vivo metabolic parameters of diabetic mice.

  3. KU-32, a novel drug for diabetic neuropathy, is safe for human islets and improves in vitro insulin secretion and viability.

    Science.gov (United States)

    Farmer, Kevin; Williams, S Janette; Novikova, Lesya; Ramachandran, Karthik; Rawal, Sonia; Blagg, Brian S J; Dobrowsky, Rick; Stehno-Bittel, Lisa

    2012-01-01

    KU-32 is a novel, novobiocin-based Hsp90 inhibitor that protects against neuronal glucotoxicity and reverses multiple clinical indices of diabetic peripheral neuropathy in a rodent model. However, any drug with potential for treating diabetic complications must also have no adverse effects on the function of pancreatic islets. Thus, the goal of the current study was to assess the effect of KU-32 on the in vitro viability and function of human islets. Treating human islets with KU-32 for 24 hours showed no toxicity as assessed using the alamarBlue assay. Confocal microscopy confirmed that with a minimum of 2-day exposure, KU-32 improved cellular viability by blocking apoptosis. Functionally, isolated human islets released more glucose-stimulated insulin when preincubated in KU-32. However, diabetic BKS-db/db mice, a model for type 2 diabetes, administered KU-32 for 10 weeks did not show any significant changes in blood glucose and insulin levels, despite having greater insulin staining/beta cell in the pancreas compared to untreated BKS db/db mice. In summary, KU-32 did not harm isolated human islets and may even be protective. However, the effect does not appear significant enough to alter the in vivo metabolic parameters of diabetic mice.

  4. Novel Small Molecule Glucagon-Like Peptide-1 Receptor Agonist Stimulates Insulin Secretion in Rodents and From Human Islets

    Science.gov (United States)

    Sloop, Kyle W.; Willard, Francis S.; Brenner, Martin B.; Ficorilli, James; Valasek, Kathleen; Showalter, Aaron D.; Farb, Thomas B.; Cao, Julia X.C.; Cox, Amy L.; Michael, M. Dodson; Gutierrez Sanfeliciano, Sonia Maria; Tebbe, Mark J.; Coghlan, Michael J.

    2010-01-01

    OBJECTIVE The clinical effectiveness of parenterally-administered glucagon-like peptide-1 (GLP-1) mimetics to improve glucose control in patients suffering from type 2 diabetes strongly supports discovery pursuits aimed at identifying and developing orally active, small molecule GLP-1 receptor agonists. The purpose of these studies was to identify and characterize novel nonpeptide agonists of the GLP-1 receptor. RESEARCH DESIGN AND METHODS Screening using cells expressing the GLP-1 receptor and insulin secretion assays with rodent and human islets were used to identify novel molecules. The intravenous glucose tolerance test (IVGTT) and hyperglycemic clamp characterized the insulinotropic effects of compounds in vivo. RESULTS Novel low molecular weight pyrimidine-based compounds that activate the GLP-1 receptor and stimulate glucose-dependent insulin secretion are described. These molecules induce GLP-1 receptor-mediated cAMP signaling in HEK293 cells expressing the GLP-1 receptor and increase insulin secretion from rodent islets in a dose-dependent manner. The compounds activate GLP-1 receptor signaling, both alone or in an additive fashion when combined with the endogenous GLP-1 peptide; however, these agonists do not compete with radiolabeled GLP-1 in receptor-binding assays. In vivo studies using the IVGTT and the hyperglycemic clamp in Sprague Dawley rats demonstrate increased insulin secretion in compound-treated animals. Further, perifusion assays with human islets isolated from a donor with type 2 diabetes show near-normalization of insulin secretion upon compound treatment. CONCLUSIONS These studies characterize the insulinotropic effects of an early-stage, small molecule GLP-1 receptor agonist and provide compelling evidence to support pharmaceutical optimization. PMID:20823098

  5. Islet Transplantation

    Science.gov (United States)

    ... transplanted islet cells failed. But in recent years, scientists have begun to make rapid advances in transplant technology, and some of the most exciting new research comes to us from researchers at the University of ... Canada. These scientists have used a new procedure called the Edmonton ...

  6. Palmitate activates autophagy in INS-1E β-cells and in isolated rat and human pancreatic islets.

    Directory of Open Access Journals (Sweden)

    Luisa Martino

    Full Text Available We have investigated the in vitro effects of increased levels of glucose and free fatty acids on autophagy activation in pancreatic beta cells. INS-1E cells and isolated rat and human pancreatic islets were incubated for various times (from 2 to 24 h at different concentrations of glucose and/or palmitic acid. Then, cell survival was evaluated and autophagy activation was explored by using various biochemical and morphological techniques. In INS-1E cells as well as in rat and human islets, 0.5 and 1.0 mM palmitate markedly increased autophagic vacuole formation, whereas high glucose was ineffective alone and caused little additional change when combined with palmitate. Furthermore, LC3-II immunofluorescence co-localized with that of cathepsin D, a lysosomal marker, showing that the autophagic flux was not hampered in PA-treated cells. These effects were maintained up to 18-24 h incubation and were associated with a significant decline of cell survival correlated with both palmitate concentration and incubation time. Ultrastructural analysis showed that autophagy activation, as evidenced by the occurrence of many autophagic vacuoles in the cytoplasm of beta cells, was associated with a diffuse and remarkable swelling of the endoplasmic reticulum. Our results indicate that among the metabolic alterations typically associated with type 2 diabetes, high free fatty acids levels could play a role in the activation of autophagy in beta cells, through a mechanism that might involve the induction of endoplasmic reticulum stress.

  7. Perioperative release of pro-regenerative biochemical signals from human renal allografts subjected to ischemia-reperfusion injury.

    Science.gov (United States)

    Błogowski, Wojciech; Dolegowska, Barbara; Budkowska, Marta; Sałata, Daria; Domański, Leszek; Starzynska, Teresa

    2014-02-01

    Complement-derived molecules modulate the intensity of renal ischemia-reperfusion injury and may lead to the generation of biochemical signals [such as stromal-derived factor-1 (SDF-1) or sphingosine-1-phosphate (S1P)], which stimulate tissue/organ regeneration after injury. We tested the association between perioperative C5b-9/membrane attack complex (MAC) levels and intensified erythrocyte lysis, and asked whether significant changes in the levels of pro-regenerative substances occur during the early phase of renal allograft reperfusion. Seventy-five recipients were enrolled and divided into the early, slow, and delayed graft function (DGF) groups. Perioperative blood samples were collected from the renal vein during consecutive minutes of reperfusion. Extracellular hemoglobin (eHb), albumin (plasma S1P transporter), 8-iPF2α-III isoprostane, SDF-1 and S1P concentrations were measured. Throughout the reperfusion period, erythrocyte lysis intensified and was most pronounced in the DGF group. However, perioperative eHb levels did not correlate significantly with C5b-9/MAC values, but rather with the intensity of oxidative stress. No significant changes were observed in S1P, its plasma transporter (albumin) or SDF-1 levels, which were relatively low in all groups throughout the reperfusion period. Our study therefore demonstrates that no known biochemical signal for bone marrow-derived stem cell mobilization is released from human renal allografts to the periphery during the early phase of reperfusion.

  8. Establishing a human pancreatic stem cell line and transplanting induced pancreatic islets to reverse experimental diabetes in rats

    Institute of Scientific and Technical Information of China (English)

    XIAO Mei; DOU ZhongYing; AN LiLong; YANG XueYi; GE Xin; QIAO Hai; ZHAO Ting; MA XiaoFei; FAN JingZhua; ZHU MengYang

    2008-01-01

    The major obstacle in using pancreatic islet transplantation to cure type Ⅰ and some type Ⅱ diabetes is the shortage of the donors. One of ways to overcome such obstacle is to isolate and clone pancreatic stem cells as "seed cells" and induce their differentiation into functional islets as an abundant trans-plantation source. In this study, a monoclonal human pancreatic stem cell (mhPSC) line was obtained from abortive fetal pancreatic tissues. Pancreatic tissues were taken from abortive fetus by sterile procedures, and digested into single cells and cell clusters with 0.1% type Ⅳ collagenase. Cultured in modified glucose-low DMEM with 10% fetal bovine serum (FBS), these single cells and cell clusters adhered to culture dishes, and then primary epidermal-like pancreatic stem ceils started to clone. After digesting with 0.25% trypsin and 0.04% EDTA, fibroblasts and other cells were gradually eliminated and epithelioid pancreatic stem cells were gradually purified during generations. Using clone-ring selection, the mhPSCs were obtained. After addition of 10 ng/mL epidermal growth factor (EGF) in cell culture medium, the mhPSCs quickly grew and formed a gravelstone-like monolayer. Continuously proliferated, a mhPSC line, which was derived from a male abortive fetus of 4 months old, has been passed through 50 generations. More than 1×109 mhPSCs were cryo-preserved in liquid nitrogen. Karyotype analysis showed that the chromosome set of the mhPSC line was normal diploid. Immunocytochemistry results demonstrated that the mhPSC line was positive for the pdxl, glucagon, nestin and CK19, and negative for the insulin, CD34, CD44 and CD45 protein expression. RT-PCR revealed further that the mhPSCs expressed transcription factors of the pdx1, glucagon, nestin and CK19. Also, in vitro induced with β-mercaptoethanol, the mhPSCs differentiated into nerve cells that expressed the NF protein. Induced with nicotinamide, the mhPSCs differentiated into functional islet

  9. Establishing a human pancreatic stem cell line and transplanting induced pancreatic islets to reverse experimental diabetes in rats

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The major obstacle in using pancreatic islet transplantation to cure type I and some type II diabetes is the shortage of the donors. One of ways to overcome such obstacle is to isolate and clone pancreatic stem cells as "seed cells" and induce their differentiation into functional islets as an abundant trans-plantation source. In this study, a monoclonal human pancreatic stem cell (mhPSC) line was obtained from abortive fetal pancreatic tissues. Pancreatic tissues were taken from abortive fetus by sterile procedures, and digested into single cells and cell clusters with 0.1% type IV collagenase. Cultured in modified glucose-low DMEM with 10% fetal bovine serum (FBS), these single cells and cell clusters adhered to culture dishes, and then primary epidermal-like pancreatic stem cells started to clone. After digesting with 0.25% trypsin and 0.04% EDTA, fibroblasts and other cells were gradually eliminated and epithelioid pancreatic stem cells were gradually purified during generations. Using clone-ring selection, the mhPSCs were obtained. After addition of 10 ng/mL epidermal growth factor (EGF) in cell culture medium, the mhPSCs quickly grew and formed a gravelstone-like monolayer. Continuously proliferated, a mhPSC line, which was derived from a male abortive fetus of 4 months old, has been passed through 50 generations. More than 1×109 mhPSCs were cryo-preserved in liquid nitrogen. Karyotype analysis showed that the chromosome set of the mhPSC line was normal diploid. Immunocytochemistry results demonstrated that the mhPSC line was positive for the pdx1, glucagon, nestin and CK19, and negative for the insulin, CD34, CD44 and CD45 protein expression. RT-PCR revealed further that the mhPSCs expressed transcription factors of the pdx1, glucagon, nestin and CK19. Also, in vitro induced with β-mercaptoethanol, the mhPSCs differentiated into nerve cells that expressed the NF protein. Induced with nicotinamide, the mhPSCs differentiated into functional islet

  10. Susceptibility of Human Pancreatic beta Cells for Cytomegalovirus Infection and the Effects on Cellular Immunogenicity

    NARCIS (Netherlands)

    Smelt, M.J.; Faas, M.M.; de Haan, Bart; Draijer, C.; Hugenholtz, G.C.G.; de Haan, A.; Engelse, M.A.; de Koning, E.J.P.; de Vos, P.

    2012-01-01

    Objectives: Human cytomegalovirus (HCMV) infection has been suggested to be a causal factor in the development of type 1 diabetes, posttransplantation diabetes, and the failure of islet allografts. This effect of CMV has been interpreted as an indirect effect on the immune system rather than direct

  11. The effects of exendin-4 treatment on graft failure: an animal study using a novel re-vascularized minimal human islet transplant model.

    Directory of Open Access Journals (Sweden)

    Afaf Sahraoui

    Full Text Available Islet transplantation has become a viable clinical treatment, but is still compromised by long-term graft failure. Exendin-4, a glucagon-like peptide 1 receptor agonist, has in clinical studies been shown to improve insulin secretion in islet transplanted patients. However, little is known about the effect of exendin-4 on other metabolic parameters. We therefore aimed to determine what influence exendin-4 would have on revascularized minimal human islet grafts in a state of graft failure in terms of glucose metabolism, body weight, lipid levels and graft survival. Introducing the bilateral, subcapsular islet transplantation model, we first transplanted diabetic mice with a murine graft under the left kidney capsule sufficient to restore normoglycemia. After a convalescent period, we performed a second transplantation under the right kidney capsule with a minimal human islet graft and allowed for a second recovery. We then performed a left-sided nephrectomy, and immediately started treatment with exendin-4 with a low (20μg/kg/day or high (200μg/kg/day dose, or saline subcutaneously twice daily for 15 days. Blood was sampled, blood glucose and body weight monitored. The transplanted human islet grafts were collected at study end point and analyzed. We found that exendin-4 exerts its effect on failing human islet grafts in a bell-shaped dose-response curve. Both doses of exendin-4 equally and significantly reduced blood glucose. Glucagon-like peptide 1 (GLP-1, C-peptide and pro-insulin were conversely increased. In the course of the treatment, body weight and cholesterol levels were not affected. However, immunohistochemistry revealed an increase in beta cell nuclei count and reduced TUNEL staining only in the group treated with a low dose of exendin-4 compared to the high dose and control. Collectively, these results suggest that exendin-4 has a potential rescue effect on failing, revascularized human islets in terms of lowering blood glucose

  12. The effects of exendin-4 treatment on graft failure: an animal study using a novel re-vascularized minimal human islet transplant model.

    Science.gov (United States)

    Sahraoui, Afaf; Winzell, Maria Sörhede; Gorman, Tracy; Smith, Dave M; Skrtic, Stanko; Hoeyem, Merete; Abadpour, Shadab; Johansson, Lars; Korsgren, Olle; Foss, Aksel; Scholz, Hanne

    2015-01-01

    Islet transplantation has become a viable clinical treatment, but is still compromised by long-term graft failure. Exendin-4, a glucagon-like peptide 1 receptor agonist, has in clinical studies been shown to improve insulin secretion in islet transplanted patients. However, little is known about the effect of exendin-4 on other metabolic parameters. We therefore aimed to determine what influence exendin-4 would have on revascularized minimal human islet grafts in a state of graft failure in terms of glucose metabolism, body weight, lipid levels and graft survival. Introducing the bilateral, subcapsular islet transplantation model, we first transplanted diabetic mice with a murine graft under the left kidney capsule sufficient to restore normoglycemia. After a convalescent period, we performed a second transplantation under the right kidney capsule with a minimal human islet graft and allowed for a second recovery. We then performed a left-sided nephrectomy, and immediately started treatment with exendin-4 with a low (20μg/kg/day) or high (200μg/kg/day) dose, or saline subcutaneously twice daily for 15 days. Blood was sampled, blood glucose and body weight monitored. The transplanted human islet grafts were collected at study end point and analyzed. We found that exendin-4 exerts its effect on failing human islet grafts in a bell-shaped dose-response curve. Both doses of exendin-4 equally and significantly reduced blood glucose. Glucagon-like peptide 1 (GLP-1), C-peptide and pro-insulin were conversely increased. In the course of the treatment, body weight and cholesterol levels were not affected. However, immunohistochemistry revealed an increase in beta cell nuclei count and reduced TUNEL staining only in the group treated with a low dose of exendin-4 compared to the high dose and control. Collectively, these results suggest that exendin-4 has a potential rescue effect on failing, revascularized human islets in terms of lowering blood glucose, maintaining beta

  13. Lixisenatide accelerates restoration of normoglycemia and improves human beta-cell function and survival in diabetic immunodeficient NOD–scid IL-2rgnull RIP-DTR mice engrafted with human islets

    Directory of Open Access Journals (Sweden)

    Yang C

    2015-08-01

    Full Text Available Chaoxing Yang,1 Matthias Loehn,2 Agata Jurczyk,1 Natalia Przewozniak,1 Linda Leehy,1 Pedro L Herrera,3 Leonard D Shultz,4 Dale L Greiner,1 David M Harlan,5 Rita Bortell1 1Program in Molecular Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, Worcester, MA, USA; 2Sanofi-Aventis, Diabetes Division, Frankfurt, Germany; 3University of Geneva, Geneva, Switzerland; 4The Jackson Laboratory, Bar Harbor, ME, USA; 5Department of Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, Worcester, MA, USA Objective: Glucagon-like peptide-1 induces glucose-dependent insulin secretion and, in rodents, increases proliferation and survival of pancreatic beta cells. To investigate the effects on human beta cells, we used immunodeficient mice transplanted with human islets. The goal was to determine whether lixisenatide, a glucagon-like peptide-1 receptor agonist, improves human islet function and survival in vivo. Methods: Five independent transplant studies were conducted with human islets from five individual donors. Diabetic human islet-engrafted immunodeficient mice were treated with lixisenatide (50, 150, and 500 µg/kg or vehicle. Islet function was determined by blood glucose, plasma human insulin/C-peptide, and glucose tolerance tests. Grafts were analyzed for total beta- and alpha-cell number, percent proliferation, and levels of apoptosis. Results: Diabetic mice transplanted with marginal human islet mass and treated with lixisenatide were restored to euglycemia more rapidly than vehicle-treated mice. Glucose tolerance tests, human plasma insulin, and glucose-stimulation indices of lixisenatide-treated mice were significantly improved compared to vehicle-treated mice. The percentages of proliferating or apoptotic beta cells at graft recovery were not different between lixisenatide-treated and vehicle-treated mice. Nevertheless, in one experiment we found a significant twofold to threefold

  14. Human Islet Amyloid Polypeptide N-Terminus Fragment Self-Assembly: Effect of Conserved Disulfide Bond on Aggregation Propensity

    Science.gov (United States)

    Ilitchev, Alexandre I.; Giammona, Maxwell J.; Do, Thanh D.; Wong, Amy G.; Buratto, Steven K.; Shea, Joan-Emma; Raleigh, Daniel P.; Bowers, Michael T.

    2016-06-01

    Amyloid formation by human islet amyloid polypeptide (hIAPP) has long been implicated in the pathogeny of type 2 diabetes mellitus (T2DM) and failure of islet transplants, but the mechanism of IAPP self-assembly is still unclear. Numerous fragments of hIAPP are capable of self-association into oligomeric aggregates, both amyloid and non-amyloid in structure. The N-terminal region of IAPP contains a conserved disulfide bond between cysteines at position 2 and 7, which is important to hIAPP's in vivo function and may play a role in in vitro aggregation. The importance of the disulfide bond in this region was probed using a combination of ion mobility-based mass spectrometry experiments, molecular dynamics simulations, and high-resolution atomic force microscopy imaging on the wildtype 1-8 hIAPP fragment, a reduced fragment with no disulfide bond, and a fragment with both cysteines at positions 2 and 7 mutated to serine. The results indicate the wildtype fragment aggregates by a different pathway than either comparison peptide and that the intact disulfide bond may be protective against aggregation due to a reduction of inter-peptide hydrogen bonding.

  15. The Pancreatic Islet Regulome Browser

    Science.gov (United States)

    Mularoni, Loris; Ramos-Rodríguez, Mireia; Pasquali, Lorenzo

    2017-01-01

    The pancreatic islet is a highly specialized tissue embedded in the exocrine pancreas whose primary function is that of controlling glucose homeostasis. Thus, understanding the transcriptional control of islet-cell may help to puzzle out the pathogenesis of glucose metabolism disorders. Integrative computational analyses of transcriptomic and epigenomic data allows predicting genomic coordinates of putative regulatory elements across the genome and, decipher tissue-specific functions of the non-coding genome. We herein present the Islet Regulome Browser, a tool that allows fast access and exploration of pancreatic islet epigenomic and transcriptomic data produced by different labs worldwide. The Islet Regulome Browser is now accessible on the internet or may be installed locally. It allows uploading custom tracks as well as providing interactive access to a wealth of information including Genome-Wide Association Studies (GWAS) variants, different classes of regulatory elements, together with enhancer clusters, stretch-enhancers and transcription factor binding sites in pancreatic progenitors and adult human pancreatic islets. Integration and visualization of such data may allow a deeper understanding of the regulatory networks driving tissue-specific transcription and guide the identification of regulatory variants. We believe that such tool will facilitate the access to pancreatic islet public genomic datasets providing a major boost to functional genomics studies in glucose metabolism related traits including diabetes. PMID:28261261

  16. Munc18b Increases Insulin Granule Fusion, Restoring Deficient Insulin Secretion in Type-2 Diabetes Human and Goto-Kakizaki Rat Islets with Improvement in Glucose Homeostasis

    Directory of Open Access Journals (Sweden)

    Tairan Qin

    2017-02-01

    Infusion of Ad-Munc18b into GK rat pancreas led to sustained improvement in glucose homeostasis. However, Munc18b overexpression in normal islets increased only newcomer SG fusion. Therefore, Munc18b could potentially be deployed in human T2D to rescue the deficient GSIS.

  17. 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.

  18. Adult Human Biliary Tree Stem Cells Differentiate to β-Pancreatic Islet Cells by Treatment with a Recombinant Human Pdx1 Peptide.

    Directory of Open Access Journals (Sweden)

    Vincenzo Cardinale

    Full Text Available Generation of β-pancreatic cells represents a major goal in research. The aim of this study was to explore a protein-based strategy to induce differentiation of human biliary tree stem cells (hBTSCs towards β-pancreatic cells. A plasmid containing the sequence of the human pancreatic and duodenal homeobox 1 (PDX1 has been expressed in E. coli. Epithelial-Cell-Adhesion-Molecule positive hBTSCs or mature human hepatocyte cell line, HepG2, were grown in medium to which Pdx1 peptide was added. Differentiation toward pancreatic islet cells were evaluated by the expression of the β-cell transcription factors, Pdx1 and musculoapo-neurotic fibrosarcoma oncogene homolog A, and of the pancreatic hormones, insulin, glucagon, and somatostatin, investigated by real time polymerase chain reaction, western blot, light microscopy and immunofluorescence. C-peptide secretion in response to high glucose was also measured. Results indicated how purified Pdx1 protein corresponding to the primary structure of the human Pdx1 by mass spectroscopy was efficiently produced in bacteria, and transduced into hBTSCs. Pdx1 exposure triggered the expression of both intermediate and mature stage β-cell differentiation markers only in hBTSCs but not in HepG2 cell line. Furthermore, hBTSCs exposed to Pdx1 showed up-regulation of insulin, glucagon and somatostatin genes and formation of 3-dimensional islet-like structures intensely positive for insulin and glucagon. Finally, Pdx1-induced islet-like structures exhibited glucose-regulated C-peptide secretion. In conclusion, the human Pdx1 is highly effective in triggering hBTSC differentiation toward functional β-pancreatic cells.

  19. Characterization of ionizing radiation effects on human skin allografts; Caracterizacao dos efeitos da radiacao ionizante em pele humana para aloenxerto

    Energy Technology Data Exchange (ETDEWEB)

    Bourroul, Selma Cecilia

    2004-07-01

    The skin has a fundamental role in the viability of the human body. In the cases of extensive wounds, allograft skin provides an alternative to cover temporarily the damaged areas. After donor screening and preservation in glycerol (above 85%), the skin can be stored in the Skin Banks. The glycerol at this concentration has a bacteriostatic effect after certain time of preservation. On the other hand, skin sterilization by ionizing radiation may reduces the quarantine period for transplantation in patients and its safety is considered excellent. The objectives of this work were to establish procedures using two sources of ionizing radiation for sterilization of human skin allograft, and to evaluate the skin after gamma and electron beam irradiation. The analysis of stress-strain intended to verify possible effects of the radiation on the structure of preserved grafts. Skin samples were submitted to doses of 25 kGy and 50 kGy in an irradiator of {sup 60}Co and in an electron beam accelerator. Morphology and ultra-structure studies were also accomplished. The samples irradiated with a dose of 25 kGy seemed to maintain the bio mechanic characteristics. The gamma irradiated samples with a dose of 50 kGy and submitted to an electron beam at doses of 25 kGy and 50 kGy presented significant differences in the values of the elasticity modulus, in relation to the control. The analysis of the ultramicrographies revealed modifications in the structure and alterations in the pattern of collagen fibrils periodicity of the irradiated samples. (author)

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

    Directory of Open Access Journals (Sweden)

    Volkert A L Huurman

    Full Text Available BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: In this cohort study, cellular islet-specific autoimmunity associates with clinical outcome of islet cell transplantation under ATG

  1. Biotin uptake by mouse and human pancreatic beta cells/islets: a regulated, lipopolysaccharide-sensitive carrier-mediated process.

    Science.gov (United States)

    Ghosal, Abhisek; Sekar, Thillai V; Said, Hamid M

    2014-08-01

    Biotin is essential for the normal function of pancreatic beta cells. These cells obtain biotin from their surroundings via transport across their cell membrane. Little is known about the uptake mechanism involved, how it is regulated, and how it is affected by internal and external factors. We addressed these issues using the mouse-derived pancreatic beta-TC-6 cells and freshly isolated mouse and human primary pancreatic beta cells as models. The results showed biotin uptake by pancreatic beta-TC-6 cells occurs via a Na(+)-dependent, carrier-mediated process, that is sensitive to desthiobiotin, as well as to pantothenic acid and lipoate; the process is also saturable as a function of concentration (apparent Km = 22.24 ± 5.5 μM). These cells express the sodium-dependent multivitamin transporter (SMVT), whose knockdown (with doxycycline-inducible shRNA) led to a sever inhibition in biotin uptake. Similarly, uptake of biotin by mouse and human primary pancreatic islets is Na(+)-dependent and carrier-mediated, and both cell types express SMVT. Biotin uptake by pancreatic beta-TC-6 cells is also adaptively regulated (via transcriptional mechanism) by extracellular substrate level. Chronic treatment of pancreatic beta-TC-6 cells with bacterial lipopolysaccharides (LPS) leads to inhibition in biotin uptake. This inhibition is mediated via a Toll-Like receptor 4-mediated process and involves a decrease in membrane expression of SMVT. These findings show, for the first time, that pancreatic beta cells/islets take up biotin via a specific and regulated carrier-mediated process, and that the process is sensitive to the effect of LPS.

  2. Discovery of novel glucose-regulated proteins in isolated human pancreatic islets using LC-MS/MS-based proteomics.

    Science.gov (United States)

    Schrimpe-Rutledge, Alexandra C; Fontès, Ghislaine; Gritsenko, Marina A; Norbeck, Angela D; Anderson, David J; Waters, Katrina M; Adkins, Joshua N; Smith, Richard D; Poitout, Vincent; Metz, Thomas O

    2012-07-06

    The prevalence of diabetes mellitus is increasing dramatically throughout the world, and the disease has become a major public health issue. The most common form of the disease, type 2 diabetes, is characterized by insulin resistance and insufficient insulin production from the pancreatic beta-cell. Since glucose is the most potent regulator of beta-cell function under physiological conditions, identification of the insulin secretory defect underlying type 2 diabetes requires a better understanding of glucose regulation of human beta-cell function. To this aim, a bottom-up LC-MS/MS-based proteomics approach was used to profile pooled islets from multiple donors under basal (5 mM) or high (15 mM) glucose conditions. Our analysis discovered 256 differentially abundant proteins (∼p < 0.05) after 24 h of high glucose exposure from more than 4500 identified in total. Several novel glucose-regulated proteins were elevated under high glucose conditions, including regulators of mRNA splicing (pleiotropic regulator 1), processing (retinoblastoma binding protein 6), and function (nuclear RNA export factor 1), in addition to neuron navigator 1 and plasminogen activator inhibitor 1. Proteins whose abundances markedly decreased during incubation at 15 mM glucose included Bax inhibitor 1 and synaptotagmin-17. Up-regulation of dicer 1 and SLC27A2 and down-regulation of phospholipase Cβ4 were confirmed by Western blots. Many proteins found to be differentially abundant after high glucose stimulation are annotated as uncharacterized or hypothetical. These findings expand our knowledge of glucose regulation of the human islet proteome and suggest many hitherto unknown responses to glucose that require additional studies to explore novel functional roles.

  3. Noninvasive imaging of islet grafts using positron-emission tomography

    Science.gov (United States)

    Lu, Yuxin; Dang, Hoa; Middleton, Blake; Zhang, Zesong; Washburn, Lorraine; Stout, David B.; Campbell-Thompson, Martha; Atkinson, Mark A.; Phelps, Michael; Gambhir, Sanjiv Sam; Tian, Jide; Kaufman, Daniel L.

    2006-07-01

    Islet transplantation offers a potential therapy to restore glucose homeostasis in type 1 diabetes patients. However, islet transplantation is not routinely successful because most islet recipients gradually lose graft function. Furthermore, serological markers of islet function are insensitive to islet loss until the latter stages of islet graft rejection. A noninvasive method of monitoring islet grafts would aid in the assessment of islet graft survival and the evaluation of interventions designed to prolong graft survival. Here, we show that recombinant adenovirus can engineer isolated islets to express a positron-emission tomography (PET) reporter gene and that these islets can be repeatedly imaged by using microPET after transplantation into mice. The magnitude of signal from engineered islets implanted into the axillary cavity was directly related to the implanted islet mass. PET signals attenuated over the following weeks because of the transient nature of adenovirus-mediated gene expression. Because the liver is the preferred site for islet implantation in humans, we also tested whether islets could be imaged after transfusion into the mouse liver. Control studies revealed that both intrahepatic islet transplantation and hyperglycemia altered the biodistribution kinetics of the PET probe systemically. Although transplanted islets were dispersed throughout the liver, clear signals from the liver region of mice receiving PET reporter-expressing islets were detectable for several weeks. Viral transduction, PET reporter expression, and repeated microPET imaging had no apparent deleterious effects on islet function after implantation. These studies lay a foundation for noninvasive quantitative assessments of islet graft survival using PET. diabetes | transplantation

  4. Extracellular matrix components supporting human islet function in alginate-based immunoprotective microcapsules for treatment of diabetes

    NARCIS (Netherlands)

    Llacua Carrasco, Luis; de Haan, Bart J; Smink, Sandra A; de Vos, Paul

    2016-01-01

    In the pancreas, extracellular matrix (ECM) components play an import role in providing mechanical and physiological support, and also contribute to the function of islets. These ECM-connections are damaged during islet-isolation from the pancreas and are not fully recovered after encapsulation and

  5. Discovery of novel glucose-regulated proteins in isolated human pancreatic islets using LC-MS/MS-based proteomics

    Energy Technology Data Exchange (ETDEWEB)

    Rutledge, Alexandra C.; Fontes, Ghislaine; Gritsenko, Marina A.; Norbeck, Angela D.; Anderson, David J.; Waters, Katrina M.; Adkins, Joshua N.; Smith, Richard D.; Poitout, Vincent; Metz, Thomas O.

    2012-07-06

    The prevalence of diabetes mellitus is increasing dramatically throughout the world, and the disease has become a major public health issue. The most common form of the disease, type 2 diabetes, is due in part to insufficient insulin production from the pancreatic beta-cell. Since glucose is the most potent and physiologically important regulators of beta-cell function under physiological conditions, understanding the insulin secretory defect underlying type 2 diabetes requires a better understanding of glucose regulation of beta-cell function. To this aim, a bottom-up LC-MS/MS-based proteomics approach was used to profile pooled islets from multiple donors under basal (5 mM) or high (15 mM) glucose conditions. Our analysis discovered 256 differentially abundant proteins ({approx}p < 0.05) after 24 h of high glucose exposure from more than 4500 identified in total. Several novel glucose-regulated proteins were elevated under high glucose conditions, including regulators of mRNA splicing (Pleiotropic regulator 1), processing (Retinoblastoma binding protein 6), and function (Nuclear RNA export factor 1), in addition to Neuron navigator 1 and Plasminogen activator inhibitor 1. Proteins whose abundances markedly decreased during incubation at 15 mM glucose included Bax inhibitor 1 and Synaptotagmin-17. Many proteins found to be differentially abundant after high glucose stimulation were uncharacterized or hypothetical. These findings expand our knowledge of glucose regulation of the human islet proteome and suggest many hitherto unknown responses to glucose that require additional studies to explore novel functional roles.

  6. Acidic pH retards the fibrillization of human islet amyloid polypeptide due to electrostatic repulsion of histidines

    Science.gov (United States)

    Li, Yang; Xu, Weixin; Mu, Yuguang; Zhang, John Z. H.

    2013-08-01

    The human Islet Amyloid Polypeptide (hIAPP) is the major constituent of amyloid deposits in pancreatic islets of type-II diabetes. IAPP is secreted together with insulin from the acidic secretory granules at a low pH of approximately 5.5 to the extracellular environment at a neutral pH. The increased accumulation of extracellular hIAPP in diabetes indicates that changes in pH may promote amyloid formation. To gain insights and underlying mechanisms of the pH effect on hIAPP fibrillogenesis, all-atom molecular dynamics simulations in explicit solvent model were performed to study the structural properties of five hIAPP protofibrillar oligomers, under acidic and neutral pH, respectively. In consistent with experimental findings, simulation results show that acidic pH is not conducive to the structural stability of these oligomers. This provides a direct evidence for a recent experiment [L. Khemtemourian, E. Domenech, J. P. F. Doux, M. C. Koorengevel, and J. A. Killian, J. Am. Chem. Soc. 133, 15598 (2011)], 10.1021/ja205007j, which suggests that acidic pH inhibits the fibril formation of hIAPP. In addition, a complementary coarse-grained simulation shows the repulsive electrostatic interactions among charged His18 residues slow down the dimerization process of hIAPP by twofold. Besides, our all-atom simulations reveal acidic pH mainly affects the local structure around residue His18 by destroying the surrounding hydrogen-bonding network, due to the repulsive interactions between protonated interchain His18 residues at acidic pH. It is also disclosed that the local interactions nearby His18 operating between adjacent β-strands trigger the structural transition, which gives hints to the experimental findings that the rate of hIAPP fibril formation and the morphologies of the fibrillar structures are strongly pH-dependent.

  7. Bone mononuclear cells and pancreatic islets allograft in the treatment of diabetic rats%异体大鼠骨髓单个核细胞胰岛细胞移植治疗糖尿病

    Institute of Scientific and Technical Information of China (English)

    韩玮; 王云海; 王太成; 陈启龙

    2010-01-01

    Objective To study the curative effect of bone marrow mononuclear cells and pancreatic islets transplantation through the approach of the liver and tail vein. Methods The pancreatic islets and bone marrow mononuclear cells were isolated by discontinuous gradients centrifugation. Twenty-eight diabetic rats were randomly divided into groups A, B, C, and D. The rats in group A received 1000 pancreatic islets transplantation beneath the liver capsule, those in group B received transplantation of pancreatic islets and 1 × 107 bone marrow mononuclear cells beneath the liver capsule, those in group C received transplantation of 1000 pancreatic islets through the tail vein, and those in group D received transplantation of 1000 pancreatic islets and 1 × 107 bone marrow mononuclear cells through the tail vein. Plasma glucose level was determined at different time points. The effectiveness of combined different cells and transplantation approaches was compared. Results The blood glucose levels in groups A and B started to decrease in 3 days after transplantation. The blood glucose level in group A could be reduced to the normal level (7. 98 ±2. 28) mmol/L and maintain (3.71 ±0. 95) days, and that in group B could be reduced to (7. 72 ±1.75) mmol/L and maintain (4. 86 ± 1.06) days. The blood glucose level in groups C and D started to decrease in 4 days after transplantation. The blood glucose level in group C could be reduced to (7. 35 ±1.40) mmol/L and maintain (7. 85 ± 1.46) days, and that in group D to (7.00 ± 0. 83 ) mmol/L and maintain ( 14. 10 ± 1.21 ) days. There was statistically significant difference in blood glucose level among all groups ( P < 0. 05 ). Conclusion Transplantation of pancreatic islets combined with mononuclear cells through the tail vein in rats maintains the normal plasma glucose level for the longest duration. The plasma glucose level is ideal.%目的 观察异体骨髓单个核细胞和胰岛细胞通过肝脏和静脉途径移植后

  8. Potential for clinical pancreatic islet xenotransplantation

    Directory of Open Access Journals (Sweden)

    Bottino R

    2014-09-01

    Full Text Available Rita Bottino,1 Santosh Nagaraju,2 Vikas Satyananda,2 Hidetaka Hara,2 Martin Wijkstrom,2 Massimo Trucco,1 David KC Cooper2 1Institute of Cellular Therapeutics, Allegheny Health Network, 2Thomas E Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh, Pittsburgh, PA, USA Abstract: Diabetes mellitus is increasing worldwide. Type 1 diabetes can be treated successfully by islet allotransplantation, the results of which are steadily improving. However, the number of islets that can be obtained from deceased human donors will never be sufficient to cure more than a very small percentage of patients who might benefit from transplantation. Although there are some differences in glucose metabolism between pigs and humans, the use of pigs could provide an unlimited supply of islets, and the insulin produced would undoubtedly control glucose levels. Transplantation of islets into the portal vein results in islets residing in the liver; however, an early inflammatory response and rejection remain problematic, even when the recipient is receiving immunosuppressive therapy. In the long term, immunosuppressive drugs may exhibit toxicities to patients and specifically harm the islet cells. In contrast, encapsulation techniques provide islets with a physical barrier that prevents antibodies binding to the islet graft while still allowing insulin to be released into the recipient's circulation; in theory, patients receiving encapsulated grafts might not require exogenous immunosuppressive therapy. Nonhuman primates with encapsulated pig islet transplants have remained insulin-independent for several weeks, but long-term efficacy remains uncertain. Furthermore, techniques are now available to knock out genes from the pig and/or insert human genes, thus rendering the antigenic structure of pigs closer to that of humans, and providing protection from the human immune response. Islet transplantation from genetically engineered pigs has been

  9. Apelin is a novel islet peptide

    DEFF Research Database (Denmark)

    Ringström, Camilla; Nitert, Marloes Dekker; Bennet, Hedvig;

    2010-01-01

    Apelin, a recently discovered peptide with wide tissue distribution, regulates feeding behavior, improves glucose utilization, and inhibits insulin secretion. We examined whether apelin is expressed in human islets, as well as in normal and type 2 diabetic (T2D) animal islets. Further, we studied...

  10. Secretion of neurotensin from a human pancreatic islet cell carcinoma cell line (QGP-1N).

    Science.gov (United States)

    Tateishi, K; Funakoshi, A; Kitayama, N; Matsuoka, Y

    1993-12-10

    Effects of various secretagogues on secretion of neurotensin from a pancreatic islet cell carcinoma cell line (QGP-1N) were examined. Carbachol stimulated secretion of neurotensin concentration-dependently in the range of 10(-6) - 10(-4) M. The neurotensin secretion stimulated with 10(-5) M carbachol was completely inhibited by atropine at 10(-5) M. Phorbol ester and calcium ionophore (A23187) stimulated secretion of neurotensin. The removal of extracellular Ca2+ suppressed the secretion through the stimulation with 10(-5) M carbachol. Fluoride, an activator of guanine nucleotide-binding (G) protein, stimulated secretion of neurotensin. Neurotensin released into culture medium through stimulation with carbachol coeluted with neurotensin 1-13 on a gel-chromatography. Our results suggest that secretion of neurotensin from QGP-1N cells is mainly regulated by acetylcholine through muscarinic receptors coupled to G protein and that an increase in intracellular Ca2+ and protein kinase C play an important role in stimulus-secretion coupling.

  11. Islet amyloid polypeptide, islet amyloid, and diabetes mellitus.

    Science.gov (United States)

    Westermark, Per; Andersson, Arne; Westermark, Gunilla T

    2011-07-01

    Islet amyloid polypeptide (IAPP, or amylin) is one of the major secretory products of β-cells of the pancreatic islets of Langerhans. It is a regulatory peptide with putative function both locally in the islets, where it inhibits insulin and glucagon secretion, and at distant targets. It has binding sites in the brain, possibly contributing also to satiety regulation and inhibits gastric emptying. Effects on several other organs have also been described. IAPP was discovered through its ability to aggregate into pancreatic islet amyloid deposits, which are seen particularly in association with type 2 diabetes in humans and with diabetes in a few other mammalian species, especially monkeys and cats. Aggregated IAPP has cytotoxic properties and is believed to be of critical importance for the loss of β-cells in type 2 diabetes and also in pancreatic islets transplanted into individuals with type 1 diabetes. This review deals both with physiological aspects of IAPP and with the pathophysiological role of aggregated forms of IAPP, including mechanisms whereby human IAPP forms toxic aggregates and amyloid fibrils.

  12. Human umbilical cord matrix-derived stem cells exert trophic effects on β-cell survival in diabetic rats and isolated islets

    Directory of Open Access Journals (Sweden)

    Yunting Zhou

    2015-12-01

    Full Text Available Human umbilical cord matrix-derived stem cells (uMSCs, owing to their cellular and procurement advantages compared with mesenchymal stem cells derived from other tissue sources, are in clinical trials to treat type 1 (T1D and type 2 diabetes (T2D. However, the therapeutic basis remains to be fully understood. The immunomodulatory property of uMSCs could explain the use in treating T1D; however, the mere immune modulation might not be sufficient to support the use in T2D. We thus tested whether uMSCs could exert direct trophic effects on β-cells. Infusion of uMSCs into chemically induced diabetic rats prevented hyperglycemic progression with a parallel preservation of islet size and cellularity, demonstrating the protective effect of uMSCs on β-cells. Mechanistic analyses revealed that uMSCs engrafted long-term in the injured pancreas and the engraftment markedly activated the pancreatic PI3K pathway and its downstream anti-apoptotic machinery. The pro-survival pathway activation was associated with the expression and secretion of β-cell growth factors by uMSCs, among which insulin-like growth factor 1 (IGF1 was highly abundant. To establish the causal relationship between the uMSC-secreted factors and β-cell survival, isolated rat islets were co-cultured with uMSCs in the transwell system. Co-culturing improved the islet viability and insulin secretion. Furthermore, reduction of uMSC-secreted IGF1 via siRNA knockdown diminished the protective effects on islets in the co-culture. Thus, our data support a model whereby uMSCs exert trophic effects on islets by secreting β-cell growth factors such as IGF1. The study reveals a novel therapeutic role of uMSCs and suggests that multiple mechanisms are employed by uMSCs to treat diabetes.

  13. Impact of exposure to low concentrations of nitric oxide on protein profile in murine and human pancreatic islet cells.

    Science.gov (United States)

    Tapia-Limonchi, Rafael; Díaz, Irene; Cahuana, Gladys M; Bautista, Mario; Martín, Franz; Soria, Bernat; Tejedo, Juan R; Bedoya, Francisco J

    2014-01-01

    Homeostatic levels of nitric oxide (NO) protect efficiently against apoptotic death in both human and rodent pancreatic β cells, but the protein profile of this action remains to be determined. We have applied a 2 dimensional LC-MS-MALDI-TOF/TOF-based analysis to study the impact of protective NO in rat insulin-producing RINm5F cell line and in mouse and human pancreatic islets (HPI) exposed to serum deprivation condition. 24 proteins in RINm5F and 22 in HPI were identified to undergo changes in at least one experimental condition. These include stress response mitochondrial proteins (UQCRC2, VDAC1, ATP5C1, ATP5A1) in RINm5F cells and stress response endoplasmic reticulum proteins (HSPA5, PDIA6, VCP, GANAB) in HPI. In addition, metabolic and structural proteins, oxidoreductases and chaperones related with protein metabolism are also regulated by NO treatment. Network analysis of differentially expressed proteins shows their interaction in glucocorticoid receptor and NRF2-mediated oxidative stress response pathways and eNOS signaling. The results indicate that exposure to exogenous NO counteracts the impact of serum deprivation on pancreatic β cell proteome. Species differences in the proteins involved are apparent.

  14. Impact of exposure to low concentrations of nitric oxide on protein profile in murine and human pancreatic islet cells

    Science.gov (United States)

    Tapia-Limonchi, Rafael; Díaz, Irene; Cahuana, Gladys M; Bautista, Mario; Martín, Franz; Soria, Bernat; Tejedo, Juan R; Bedoya, Francisco J

    2014-01-01

    Homeostatic levels of nitric oxide (NO) protect efficiently against apoptotic death in both human and rodent pancreatic β cells, but the protein profile of this action remains to be determined. We have applied a 2 dimensional LC-MS-MALDI-TOF/TOF-based analysis to study the impact of protective NO in rat insulin-producing RINm5F cell line and in mouse and human pancreatic islets (HPI) exposed to serum deprivation condition. 24 proteins in RINm5F and 22 in HPI were identified to undergo changes in at least one experimental condition. These include stress response mitochondrial proteins (UQCRC2, VDAC1, ATP5C1, ATP5A1) in RINm5F cells and stress response endoplasmic reticulum proteins (HSPA5, PDIA6, VCP, GANAB) in HPI. In addition, metabolic and structural proteins, oxidoreductases and chaperones related with protein metabolism are also regulated by NO treatment. Network analysis of differentially expressed proteins shows their interaction in glucocorticoid receptor and NRF2-mediated oxidative stress response pathways and eNOS signaling. The results indicate that exposure to exogenous NO counteracts the impact of serum deprivation on pancreatic β cell proteome. Species differences in the proteins involved are apparent. PMID:25658244

  15. Disturbed α-Cell Function in Mice with β-Cell Specific Overexpression of Human Islet Amyloid Polypeptide

    Directory of Open Access Journals (Sweden)

    Bo Ahrén

    2008-01-01

    Full Text Available Exogenous administration of islet amyloid polypeptide (IAPP has been shown to inhibit both insulin and glucagon secretion. This study examined α-cell function in mice with β-cell specific overexpression of human IAPP (hIAPP after an oral protein gavage (75 mg whey protein/mouse. Baseline glucagon levels were higher in transgenic mice (41±4.0 pg/mL, n=6 than in wildtype animals (19±5.1 pg/mL, n=5, P=.015. In contrast, the glucagon response to protein was impaired in transgenic animals (21±2.7 pg/mL in transgenic mice versus 38±5.7 pg/mL in wildtype mice at 15 minutes; P=.027. Baseline insulin levels did not differ between the groups, while the insulin response, as the glucagon response, was impaired after protein challenge (P=.018. Glucose levels were not different between the groups and did not change significantly after protein gavage. Acetaminophen was given through gavage to the animals (2 mg/mouse to estimate gastric emptying. The plasma acetaminophen profile was similar in the two groups of mice. We conclude that disturbances in glucagon secretion exist in mice with β-cell specific overexpression of human IAPP, which are not secondary to changes in gastric emptying. The reduced glucagon response to protein challenge may reflect a direct inhibitory influence of hIAPP on glucagon secretion.

  16. Characterisation of the insulinotropic activity of an aqueous extract of Gymnema sylvestre in mouse beta-cells and human islets of Langerhans.

    Science.gov (United States)

    Liu, Bo; Asare-Anane, Henry; Al-Romaiyan, Altaf; Huang, Guocai; Amiel, Stephanie A; Jones, Peter M; Persaud, Shanta J

    2009-01-01

    Leaves of the Gymnema sylvestre (GS) plant have been used to treat diabetes mellitus for millennia, but the previously documented insulin secretagogue effects of GS extracts in vitro may be non-physiological through damage to the beta-cells. We have now examined the effects of a novel GS extract (termed OSA) on insulin secretion from the MIN6 beta-cell line and isolated human islets of Langerhans. Insulin secretion from MIN6 cells was stimulated by OSA in a concentration-dependent manner, with low concentrations (0.06-0.25 mg/ml) having no deleterious effects on MIN6 cell viability, while higher concentrations (> or = 0.5 mg/ml) caused increased Trypan blue uptake. OSA increased beta-cell Ca2+ levels, an effect that was mediated by Ca2+ influx through voltage-operated calcium channels. OSA also reversibly stimulated insulin secretion from isolated human islets and its insulin secretagogue effects in MIN6 cells and human islets were partially dependent on the presence of extracellular Ca2+. These data indicate that low concentrations of the GS isolate OSA stimulate insulin secretion in vitro, at least in part as a consequence of Ca2+ influx, without compromising beta-cell viability. Identification of the component of the OSA extract that stimulates regulated insulin exocytosis, and further investigation of its mode(s) of action, may provide promising lead targets for Type 2 diabetes therapy.

  17. A somatostatin-secreting cell line established from a human pancreatic islet cell carcinoma (somatostatinoma): release experiment and immunohistochemical study.

    Science.gov (United States)

    Iguchi, H; Hayashi, I; Kono, A

    1990-06-15

    Production and secretion of somatostatin (SRIF) were studied using a carcinoembryonic antigen (CEA)-producing cell line (QGP-1) established from a human pancreatic islet cell carcinoma. High concentrations of SRIF (274 +/- 51 ng/mg of protein, mean +/- SD, n = 5) and CEA (3083 +/- 347 ng/mg of protein, mean +/- SD, n = 5) were present in QGP-1 cells, and the basal secretion rates of SRIF and CEA by the cells (n = 5) were 46.4 +/- 4.8 and 1690 +/- 78 pg/10(5) cells/h, respectively. Immunohistochemical studies revealed the presence of SRIF in xenografts of QGP-1 cells and colocalization of SRIF and CEA. Secretion of SRIF by QGP-1 cells was stimulated in the presence of high K+ (50 mmol) and theophylline (10 mmol), but arginine (10 mmol) and glucose (300 mg/dl) had no effect on the SRIF secretion. The QGP-1 cell line may be useful for studying the regulation mechanism of SRIF secretion.

  18. Mechanism of Inhibition of Human Islet Amyloid Polypeptide-Induced Membrane Damage by a Small Organic Fluorogen

    Science.gov (United States)

    Li, Xiaoxu; Wan, Mingwei; Gao, Lianghui; Fang, Weihai

    2016-02-01

    Human islet amyloid polypeptide (hIAPP) is believed to be responsible for the death of insulin-producing β-cells. However, the mechanism of membrane damage at the molecular level has not been fully elucidated. In this article, we employ coarse- grained dissipative particle dynamics simulations to study the interactions between a lipid bilayer membrane composed of 70% zwitterionic lipids and 30% anionic lipids and hIAPPs with α-helical structures. We demonstrated that the key factor controlling pore formation is the combination of peptide charge-induced electroporation and peptide hydrophobicity-induced lipid disordering and membrane thinning. According to these mechanisms, we suggest that a water-miscible tetraphenylethene BSPOTPE is a potent inhibitor to rescue hIAPP-induced cytotoxicity. Our simulations predict that BSPOTPE molecules can bind directly to the helical regions of hIAPP and form oligomers with separated hydrophobic cores and hydrophilic shells. The micelle-like hIAPP-BSPOTPE clusters tend to be retained in the water/membrane interface and aggregate therein rather than penetrate into the membrane. Electrostatic attraction between BSPOTPE and hIAPP also reduces the extent of hIAPP binding to the anionic lipid bilayer. These two modes work together and efficiently prevent membrane poration.

  19. Graft function 1 year after pregnancy in an islet-transplanted patient.

    Science.gov (United States)

    Schive, Simen W; Scholz, Hanne; Sahraoui, Afaf; Kloster-Jensen, Kristine; Hafsahl, Geir; Korsgren, Olle; Foss, Aksel; Jenssen, Trond G

    2015-10-01

    Pancreatic islet transplantation is a treatment option for patients with type 1 diabetes (T1D), but pregnancy has generally not been advised for women after receiving an islet allograft. We hereby describe what is to our knowledge the first successful pregnancy and persistent graft function in a woman 4 years after her initial islet transplantation. A 37-year-old woman with brittle type 1 diabetes was transplanted with two separate islet graft infusions, eventually becoming insulin independent. Ten months after her second transplantation, her immunosuppression was switched from tacrolimus and sirolimus to tacrolimus, azathioprine, and prednisolone, due to her wish to become pregnant. She became pregnant one year later, and after 38 weeks of uncomplicated pregnancy, she gave birth to a healthy child by C-section. The current report suggests that pregnancy and childbirth can be accomplished after islet transplantation without loss of islet graft function.

  20. Identification of novel type 1 diabetes candidate genes by integrating genome-wide association data, protein-protein interactions, and human pancreatic islet gene expression

    DEFF Research Database (Denmark)

    Bergholdt, Regine; Brorsson, Caroline; Palleja, Albert;

    2012-01-01

    Genome-wide association studies (GWAS) have heralded a new era in susceptibility locus discovery in complex diseases. For type 1 diabetes, >40 susceptibility loci have been discovered. However, GWAS do not inevitably lead to identification of the gene or genes in a given locus associated...... with disease, and they do not typically inform the broader context in which the disease genes operate. Here, we integrated type 1 diabetes GWAS data with protein-protein interactions to construct biological networks of relevance for disease. A total of 17 networks were identified. To prioritize...... and substantiate these networks, we performed expressional profiling in human pancreatic islets exposed to proinflammatory cytokines. Three networks were significantly enriched for cytokine-regulated genes and, thus, likely to play an important role for type 1 diabetes in pancreatic islets. Eight of the regulated...

  1. Discovery of molecular pathways mediating 1,25-dihydroxyvitamin D3 protection against cytokine-induced inflammation and damage of human and male mouse islets of Langerhans

    DEFF Research Database (Denmark)

    Wolden-Kirk, Heidi; Rondas, D; Bugliani, M

    2014-01-01

    . The aim of this study was to clarify the molecular mechanisms by which 1,25(OH)2D3 contributes to β-cell protection against cytokine-induced β-cell dysfunction and death. Human and mouse islets were exposed to IL-1β and interferon-γ in the presence or absence of 1,25(OH)2D3. Effects on insulin secretion....../phenotype. In conclusion, these findings demonstrate a direct protective effect of 1,25(OH)2D3 against inflammation-induced β-cell dysfunction and death in human and murine islets, with, in particular, alterations in chemokine production by the islets. These effects may contribute to the beneficial effects of 1,25(OH)2D3......Protection against insulitis and diabetes by active vitamin D, 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), in nonobese diabetic mice has until now mainly been attributed to its immunomodulatory effects, but also protective effects of this hormone on inflammation-induced β-cell death have been reported...

  2. Acceleration of Functional Maturation and Differentiation of Neonatal Porcine Islet Cell Monolayers Shortly In Vitro Cocultured with Microencapsulated Sertoli Cells

    Directory of Open Access Journals (Sweden)

    Francesca Mancuso

    2010-01-01

    Full Text Available The limited availability of cadaveric human donor pancreata as well as the incomplete success of the Edmonton protocol for human islet allografts fasten search for new sources of insulin the producing cells for substitution cell therapy of insulin-dependent diabetes mellitus (T1DM. Starting from isolated neonatal porcine pancreatic islets (NPIs, we have obtained cell monolayers that were exposed to microencapsulated monolayered Sertoli cells (ESCs for different time periods (7, 14, 21 days. To assess the development of the cocultured cell monolayers, we have studied either endocrine cell phenotype differentiation markers or c-kit, a hematopoietic stem cell marker, has recently been involved with growth and differentiation of β-cell subpopulations in human as well as rodent animal models. ESC which were found to either accelerate maturation and differentiation of the NPIs β-cell phenotype or identify an islet cell subpopulation that was marked positively for c-kit. The insulin/c-kit positive cells might represent a new, still unknown functionally immature β-cell like element in the porcine pancreas. Acceleration of maturation and differentiation of our NPI cell monolayers might generate a potential new opportunity to develop insulin-producing cells that may suite experimental trials for cell therapy of T1DM.

  3. Microwell scaffolds for the extrahepatic transplantation of islets of Langerhans.

    Directory of Open Access Journals (Sweden)

    Mijke Buitinga

    Full Text Available Allogeneic islet transplantation into the liver has the potential to restore normoglycemia in patients with type 1 diabetes. However, the suboptimal microenvironment for islets in the liver is likely to be involved in the progressive islet dysfunction that is often observed post-transplantation. This study validates a novel microwell scaffold platform to be used for the extrahepatic transplantation of islet of Langerhans. Scaffolds were fabricated from either a thin polymer film or an electrospun mesh of poly(ethylene oxide terephthalate-poly(butylene terephthalate (PEOT/PBT block copolymer (composition: 4000PEOT30PBT70 and were imprinted with microwells, ∼400 µm in diameter and ∼350 µm in depth. The water contact angle and water uptake were 39±2° and 52.1±4.0 wt%, respectively. The glucose flux through electrospun scaffolds was three times higher than for thin film scaffolds, indicating enhanced nutrient diffusion. Human islets cultured in microwell scaffolds for seven days showed insulin release and insulin content comparable to those of free-floating control islets. Islet morphology and insulin and glucagon expression were maintained during culture in the microwell scaffolds. Our results indicate that the microwell scaffold platform prevents islet aggregation by confinement of individual islets in separate microwells, preserves the islet's native rounded morphology, and provides a protective environment without impairing islet functionality, making it a promising platform for use in extrahepatic islet transplantation.

  4. The heterogeneity of islet autoantibodies and the progression of islet failure in type 1 diabetic patients.

    Science.gov (United States)

    Liu, Jin; Bian, Lingling; Ji, Li; Chen, Yang; Chen, Heng; Gu, Yong; Ma, Bingqin; Gu, Wei; Xu, Xinyu; Shi, Yun; Wang, Jian; Zhu, Dalong; Sun, Zilin; Ma, Jianhua; Jin, Hui; Shi, Xing; Miao, Heng; Xin, Bing; Zhu, Yan; Zhang, Zhenwen; Bu, Ruifang; Xu, Lan; Shi, Guangde; Tang, Wei; Li, Wei; Zhou, Dongmei; Liang, Jun; Cheng, Xingbo; Shi, Bimin; Dong, Jixiang; Hu, Ji; Fang, Chen; Zhong, Shao; Yu, Weinan; Lu, Weiping; Wu, Chenguang; Qian, Li; Yu, Jiancheng; Gao, Jialin; Fei, Xiaoqiang; Zhang, Qingqing; Wang, Xueqin; Cui, Shiwei; Cheng, Jinluo; Xu, Ning; Wang, Guofeng; Han, Guoqing; Xu, Chunrong; Xie, Yun; An, Minmin; Zhang, Wei; Wang, Zhixiao; Cai, Yun; Fu, Qi; Fu, Yu; Zheng, Shuai; Yang, Fan; Hu, Qingfang; Dai, Hao; Jin, Yu; Zhang, Zheng; Xu, Kuanfeng; Li, Yifan; Shen, Jie; Zhou, Hongwen; He, Wei; Zheng, Xuqin; Han, Xiao; Yu, Liping; She, Jinxiong; Zhang, Mei; Yang, Tao

    2016-09-01

    Type 1 diabetes mellitus is heterogeneous in many facets. The patients suffered from type 1 diabetes present several levels of islet function as well as variable number and type of islet-specific autoantibodies. This study was to investigate prevalence and heterogeneity of the islet autoantibodies and clinical phenotypes of type 1 diabetes mellitus; and also discussed the process of islet failure and its risk factors in Chinese type 1 diabetic patients. A total of 1,291 type 1 diabetic patients were enrolled in this study. Demographic information was collected. Laboratory tests including mixed-meal tolerance test, human leukocyte antigen alleles, hemoglobinA1c, lipids, thyroid function and islet autoantibodies were conducted. The frequency of islet-specific autoantibody in newly diagnosed T1DM patients (duration shorter than half year) was 73% in East China. According to binary logistic regressions, autoantibody positivity, longer duration and lower Body Mass Index were the risk factors of islet failure. As the disease developed, autoantibodies against glutamic acid decarboxylase declined as well as the other two autoantibodies against zinc transporter 8 and islet antigen 2. The decrease of autoantibodies was positively correlated with aggressive beta cell destruction. Autoantibodies can facilitate the identification of classic T1DM from other subtypes and predict the progression of islet failure. As there were obvious heterogeneity in autoantibodies and clinical manifestation in different phenotypes of the disease, we should take more factors into consideration when identifying type 1 diabetes mellitus.

  5. Differential interleukin-1 receptor antagonism on pancreatic beta and alpha cells. Studies in rodent and human islets and in normal rats

    DEFF Research Database (Denmark)

    Zumsteg, U; Reimers, J I; Pociot, F;

    1993-01-01

    The monokines interleukin-1 alpha and -beta have been implicated as effector molecules in the immune-mediated pancreatic beta-cell destruction leading to insulin-dependent diabetes mellitus. Here we investigated the effects of interleukin-1 receptor antagonism on insulin and glucagon release of rat......, mouse and human islets exposed to recombinant human interleukin-1 beta, and on interleukin-1 beta induced changes in blood glucose, serum insulin and serum glucagon levels in Wistar Kyoto rats. The interleukin-1 receptor antagonist reduced the co-mitogenic effect of interleukin-1 beta on mouse and rat...

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

    Directory of Open Access Journals (Sweden)

    Jyuhn-Huarng Juang

    Full Text Available OBJECT: 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. MATERIALS AND METHODS: 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. RESULTS: 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. CONCLUSION: Our results indicate that after syngeneic and allogeneic transplantation, islets labeled with CSPIO nanoparticles can be effectively and safely imaged by MR.

  7. Islet Xeno/transplantation and the risk of contagion: local responses from Canada and Australia to an emerging global technoscience.

    Science.gov (United States)

    Cheng, Myra

    2015-01-01

    This paper situates the public debate over the use of living animal organs and tissue for human therapies within the history of experimental islet transplantation. Specifically, the paper compares and contrasts the Canadian and Australian responses on xenotransplantation to consider what lessons can be learnt about the regulation of a complex and controversial biotechnology. Sobbrio and Jorqui described public engagement on xenotransplantation in these countries as 'important forms of experimental democracy.' While Canada experimented with a novel nation-wide public consultation, Australia sought public input within the context of a national inquiry. In both instances, the outcome was a temporary moratorium on all forms of clinical xenotransplantation comparable to the policies adopted in some European countries. In addition, the Australian xenotransplantation ban coincided with a temporary global ban on experimental islet allotransplantation in 2007. Through historical and comparative research, this paper investigates how public controversies over organ and tissue transplantation can inform our understanding of the mediation of interspeciality and the regulation of a highly contested technoscience. It offers an alternative perspective on the xenotransplantation controversy by exploring the ways in which coinciding moratoriums on islet allograft and xenograft challenge, complicate and confound our assumptions regarding the relationships between human and animal, between routine surgery and clinical experimentation, between biomedical science and social science, and between disease risks and material contagion.

  8. Coffee components inhibit amyloid formation of human islet amyloid polypeptide in vitro: possible link between coffee consumption and diabetes mellitus.

    Science.gov (United States)

    Cheng, Biao; Liu, Xinran; Gong, Hao; Huang, Lianqi; Chen, Hong; Zhang, Xin; Li, Chuanzhou; Yang, Muyang; Ma, Bingjun; Jiao, Lihua; Zheng, Ling; Huang, Kun

    2011-12-28

    Global epidemic studies have suggested that coffee consumption is reversely correlated with the incidence of type 2 diabetes mellitus (T2DM), a metabolic disease. The misfolding of human islet amyloid polypeptide (hIAPP) is regarded as one of the causative factors of T2DM. Coffee extracts have three major active components: caffeine, caffeic acid (CA), and chlorogenic acid (CGA). In this study, the effects of these major coffee components, as well as dihydrocaffeic acid (DHCA) (a major metabolite of CGA and CA), on the amyloidogenicity of hIAPP were investigated by thioflavin-T based fluorescence emission, transmission electronic microscopy, circular dichroism, light-induced cross-linking, dynamic light scattering, and MTT-based cell viability assays. The results suggest that all components show varied inhibitory effects on the formation of toxic hIAPP amyloids, in which CA shows the highest potency in delaying the conformational transition of the hIAPP molecule with the most prolonged lag time, whereas caffeine shows the lowest potency. At a 5-fold excess molar ratio of compound to hIAPP, all coffee-derived compounds affect the secondary structures of incubated hIAPP as suggested by the circular dichroism spectra and CDPro deconvolution analysis. Further photoinduced cross-linking based oligomerization and dynamic light scattering studies suggested CA and CGA significantly suppressed the formation of hIAPP oligomers, whereas caffeine showed no significant effect on oligomerization. Cell protection effects were also observed for all three compounds, with the protection efficiency being greatest for CA and least for CGA. These findings suggest that the beneficial effects of coffee consumption on T2DM may be partly due to the ability of the major coffee components and metabolites to inhibit the toxic aggregation of hIAPP.

  9. New insights into side effect of solvents on the aggregation of human islet amyloid polypeptide 11-20.

    Science.gov (United States)

    Mao, Yexuan; Yu, Lanlan; Yang, Ran; Ma, Chuanguo; Qu, Ling-bo; Harrington, Peter de B

    2016-02-01

    The formation of highly ordered fibrils for the human islet amyloid polypeptide (hIAPP) is considered as one of the precipitating factors of type 2 diabetes mellitus. In this study, an emerging new approach microscale thermophoresis and conventional ThT fluorescence assay were utilized to investigate the aggregation behavior of hIAPP(11-20), giving a new insight of the solvent effect on the aggregation of hIAPP(11-20). hIAPP(11-20) displayed different aggregation behaviors in various buffers, revealing that hIAPP(11-20) not only self-aggregates but also binds to solvent components. hIAPP(11-20) had a higher binding affinity for Tris than other selected buffers because multiple hydrogen bonds form, resulting in weaker self-aggregation of hIAPP(11-20) at the early stage of aggregation and prolonging the fibril formation process. hIAPP(11-20) displayed similar self-aggregation in both HEPES and pure water. Negatively charged phosphate ions in the PBS solution 'neutralize' the charges carried by hIAPP(11-20) itself to some extent, causing rapid aggregation of hIAPP(11-20), and leading to a shorter fibrillation process of hIAPP(11-20). These results revealed that solvents contribute to the aggregation of hIAPP(11-20) and demonstrated the affect of solvents on the activity of biomolecules. Additionally, as a new technique, microscale thermophoresis offers a powerful and promising approach to study the early stages of aggregation of peptides or proteins.

  10. Adult Human Pancreatic Islet Beta-Cells Display Limited Turnover and Long Lifespan as Determined by In-Vivo Thymidine Analog Incorporation and Radiocarbon Dating

    Energy Technology Data Exchange (ETDEWEB)

    Perl, S; Kushner, J A; Buchholz, B A; Meeker, A K; Stein, G M; Hsieh, M; Kirby, M; Pechhold, S; Liu, E H; Harlan, D M; Tisdale, J F

    2010-03-15

    Diabetes mellitus results from an absolute or relative deficiency of insulin producing pancreatic beta-cells. The adult human beta-cell's turnover rate remains unknown. We employed novel techniques to examine adult human islet beta-cell turnover and longevity in vivo. Subjects enrolled in NIH clinical trials received thymidine analogues [iododeoxyuridine (IdU) or bromodeoxyuridine (BrdU)] 8-days to 4-years prior to death. Archival autopsy samples from ten patients (aged 17-74 years) were employed to assess beta-cell turnover by scoring nuclear analog labeling within insulin staining cells. Human adult beta-cell longevity was determined by estimating the cells genomic DNA integration of atmospheric carbon-14 ({sup 14}C). DNA was purified from pancreatic islets isolated from cadaveric donors; whole islet prep DNA was obtained from a 15 year old donor, and purified beta-cell DNA was obtained from two donors (age 48 and 80 years). {sup 14}C levels were then determined using accelerator mass spectrometry (AMS). Cellular 'birth date' was determined by comparing the subject's DNA {sup 14}C content relative to a well-established {sup 14}C atmospheric prevalence curve. In the two subjects less than age 20 years, 1-2% of the beta-cell nuclei co-stained for BrdU/IdU. No beta-cell nuclei co-stained in the eight patients more than 30 years old. Consistent with the BrdU/IdU turnover data, beta-cell DNA {sup 14}C content indicated the cells 'birth date' occurred within the subject's first 30 years of life. Under typical circumstances, adult human beta-cells and their cellular precursors are established by young adulthood.

  11. Pig islets xenotransplantation: recent progress and current perspectives

    Directory of Open Access Journals (Sweden)

    Haitao eZhu

    2014-03-01

    Full Text Available Islet xenotransplantation is a prospective treatment to bridge the gap between available human cells and needs of patients with diabetes. Pig is the ideal candidate to obtain such available islet cells. However, potential clinical application of pig islet transplantation still faces obstacles such as inadequate yield of high-quality functional islets and xenorejection of the transplants. Adequate amounts of available islets can be obtained based on selection of a suitable pathogen-free source herd and the development of isolation and purification methods. Several studies demonstrated feasibility of successful pre-clinical pig islet xenotransplantation and provided insights and possible mechanisms of xenogeneic immune recognition and rejection. Particularly promising is the achievement of long-term insulin independence in diabetic models by means of distinct islet products and novel immunotherapeutic strategies. Nonetheless, further efforts are needed to obtain much more data on safety and efficacy to translate these findings into clinical practice

  12. The immunogenicity of human cardiac valve allografts in vitro and in vivo

    NARCIS (Netherlands)

    F.M.E. Hoekstra (Franciska)

    1999-01-01

    textabstractAllogeneic transplantation has become an accepted method for the replacement of diseased organs and tissues. The concept of cardiac valve transplantation for the replacement of semilunar valves was introduced by Lam et al in 1952'. In about 1960 the first human cardiac valve

  13. Lymphoid neogenesis in skin of human hand, nonhuman primate, and rat vascularized composite allografts.

    Science.gov (United States)

    Hautz, Theresa; Zelger, Bettina G; Nasr, Isam W; Mundinger, Gerhard S; Barth, Rolf N; Rodriguez, Eduardo D; Brandacher, Gerald; Weissenbacher, Annemarie; Zelger, Bernhard; Cavadas, Pedro; Margreiter, Raimund; Lee, W P Andrew; Pratschke, Johann; Lakkis, Fadi G; Schneeberger, Stefan

    2014-09-01

    The mechanisms of skin rejection in vascularized composite allotransplantation (VCA) remain incompletely understood. The formation of tertiary lymphoid organs (TLO) in hand transplantation has been recently described. We assess this phenomenon in experimental and clinical VCA rejection. Skin biopsies of human (n = 187), nonhuman primate (n = 11), and rat (n = 15) VCAs were analyzed for presence of TLO. A comprehensive immunohistochemical assessment (characterization of the cell infiltrate, expression of adhesion molecules) including staining for peripheral node addressin (PNAd) was performed and correlated with rejection and time post-transplantation. TLO were identified in human, nonhuman primate, and rat skin samples. Expression of PNAd was increased in the endothelium of vessels upon rejection in human skin (P = 0.003) and correlated with B- and T-lymphocyte numbers and LFA-1 expression. PNAd expression was observed at all time-points after transplantation and increased significantly after year 5. In nonhuman primate skin, PNAd expression was found during inflammatory conditions early and late after transplantation. In rat skin, PNAd expression was strongly associated with acute rejection and time post-transplantation. Lymphoid neogenesis and TLO formation can be uniformly found in experimental and human VCA. PNAd expression in vascular endothelium correlates with skin rejection and T- and B-cell infiltration.

  14. Saffold Virus, a Human Cardiovirus, and Risk of Persistent Islet Autoantibodies in the Longitudinal Birth Cohort Study MIDIA.

    Directory of Open Access Journals (Sweden)

    German Tapia

    Full Text Available The aim of this study was to describe the frequency and distribution of Saffold virus in longitudinal stool samples from children, and test for association with development of persistent autoantibodies predictive of type 1 diabetes. A cohort of Norwegian children carrying the HLA genotype associated with highest risk of type 1 diabetes ("DR4-DQ8/DR3-DQ2" was followed with monthly stool samples from 3 to 35 months of age. Blood samples were tested for autoantibodies to insulin, glutamic acid decarboxylase65 and Islet Antigen-2. 2077 stool samples from 27 children with ≥ 2 repeatedly positive islet autoantibodies (cases, and 53 matched controls were analysed for Saffold virus genomic RNA by semi-quantitative real-time reverse transcriptase PCR. Saffold virus was found in 53 of 2077 (2.6% samples, with similar proportions between cases (2.5% and controls (2.6%. The probability of being infected by 3 years of age was 28% (95% CI 0.18-0.40. Viral quantities ranged from <1 to almost 105 copies/μl. Estimated odds ratio between islet autoimmunity and infection episodes prior to seroconversion was 1.98 (95% CI: 0.57-6.91, p = 0.29. Saffold virus had no statistically significant association with islet autoimmunity.

  15. The role of islet neogenesis-associated protein (INGAP) in islet neogenesis.

    Science.gov (United States)

    Lipsett, Mark; Hanley, Stephen; Castellarin, Mauro; Austin, Emily; Suarez-Pinzon, Wilma L; Rabinovitch, Alex; Rosenberg, Lawrence

    2007-01-01

    Islet Neogenesis-Associated Protein (INGAP) is a member of the Reg family of proteins implicated in various settings of endogenous pancreatic regeneration. The expression of INGAP and other RegIII proteins has also been linked temporally and spatially with the induction of islet neogenesis in animal models of disease and regeneration. Furthermore, administration of a peptide fragment of INGAP (INGAP peptide) has been demonstrated to reverse chemically induced diabetes as well as improve glycemic control and survival in an animal model of type 1 diabetes. Cultured human pancreatic tissue has also been shown to be responsive to INGAP peptide, producing islet-like structures with function, architecture and gene expression matching that of freshly isolated islets. Likewise, studies in normoglycemic animals show evidence of islet neogenesis. Finally, recent clinical studies suggest an effect of INGAP peptide to improve insulin production in type 1 diabetes and glycemic control in type 2 diabetes.

  16. Isolation, characterization, and chromosomal mapping of the human Nkx6.1 gene (NKX6A), a new pancreatic islet homeobox gene

    Energy Technology Data Exchange (ETDEWEB)

    Inoue, Hiroshi; Permutt, M.A.; Veile, R. [Washington Univ. School of Medicine, St. Louis, MO (United States)] [and others

    1997-03-01

    Nkx6.1 (gene symbol NKX6A), a new member of the NK homeobox gene family, was recently identified in rodent pancreatic islet 13-cell lines. The pattern of expression suggested that this gene product might be important for control of islet development and/or regulation of insulin biosynthesis. We now report cloning of human NKX6A, characterization of its genomic structure, and its chromosomal localization. The predicted protein of human NKX6A contained 367 amino acids and had 97% identity to the hamster protein. The highly conserved NK decapeptide and homeodomain regions were identical between human and hamster, suggesting functional importance of these domains. The coding region spanned approximately 4.8 kb and was composed of three exons. The gene was localized to four CEPH {open_quotes}B{close_quotes} yeast artificial chromosome clones (914B4, 951G9, 981D6, and 847133), and a nearby polymorphic marker (D4S1538) on chromosome 4 was identified <1270 kb from the gene. Using fluorescence in situ hybridization, we also determined that NKX6A maps to 4q21.2-q22. 11 refs., 2 figs.

  17. The Choice of Enzyme for Human Pancreas Digestion Is a Critical Factor for Increasing the Success of Islet Isolation

    Science.gov (United States)

    Qi, Meirigeng; Valiente, Luis; McFadden, Brian; Omori, Keiko; Bilbao, Shiela; Juan, Jemily; Rawson, Jeffrey; Scott, Stephen; Ferreri, Kevin; Mullen, Yoko; El-Shahawy, Mohamed; Dafoe, Donald; Kandeel, Fouad; Al-Abdullah, Ismail H.

    2015-01-01

    Background We evaluated 3 commercially available enzymes for pancreatic digestion by comparing key parameters during the islet isolation process, as well as islet quality after isolation. Methods Retrospectively compared and analyzed islet isolations from pancreata using 3 different enzyme groups: liberase HI (n = 63), collagenase NB1/neutral protease (NP) (n = 43), and liberase mammalian tissue-free collagenase/thermolysin (MTF C/T) (n = 115). A standardized islet isolation and purification method was used. Islet quality assessment was carried out using islet count, viability, in vitro glucose-stimulated insulin secretion (GSIS), glucose-stimulated oxygen consumption rate, and in vivo transplantation model in mice. Results Donor characteristics were not significantly different among the 3 enzyme groups used in terms of age, sex, hospital stay duration, cause of death, body mass index, hemoglobin A1c, cold ischemia time, and pancreas weight. Digestion efficacy (percentage of digested tissue by weight) was significantly higher in the liberase MTF C/T group (73.5 ± 1.5 %) when compared to the liberase HI group (63.6 ± 2.3 %) (P < 0.001) and the collagenase NB1/NP group (61.7 ± 2.9%) (P < 0.001). The stimulation index for GSIS was significantly higher in the liberase MTF C/T group (5.3 ± 0.5) as compared to the liberase HI (2.9 ± 0.2) (P < 0.0001) and the collagenase NB1/NP (3.6 ± 2.9) (P = 0.012) groups. Furthermore, the liberase MTF C/T enzymes showed the highest success rate of transplantation in diabetic non-obese diabetic severe combined immunodeficiency mice (65%), which was significantly higher than the liberase HI (42%, P = 0.001) and the collagenase NB1/NP enzymes (41%, P < 0.001). Conclusions Liberase MTF C/T is superior to liberase HI and collagenase NB1/NP in terms of digestion efficacy and GSIS in vitro. Moreover, liberase MTF C/T had a significantly higher success rate of transplantation in diabetic NOD Scid mice compared to liberase HI and

  18. A prospective, randomised comparative study of weekly versus biweekly application of dehydrated human amnion/chorion membrane allograft in the management of diabetic foot ulcers

    Science.gov (United States)

    Zelen, Charles M; Serena, Thomas E; Snyder, Robert J

    2014-01-01

    The aim of this study is to determine if weekly application of dehydrated human amnion/chorion membrane allograft reduce time to heal more effectively than biweekly application for treatment of diabetic foot ulcers. This was an institutional review board-approved, registered, prospective, randomised, comparative, non-blinded, single-centre clinical trial. Patients with non-infected ulcers of ≥ 4 weeks duration were included for the study. They were randomised to receive weekly or biweekly application of allograft in addition to a non-adherent, moist dressing with compressive wrapping. All wounds were offloaded. The primary study outcome was mean time to healing. Overall, during the 12-week study period, 92·5% (37/40) ulcers completely healed. Mean time to complete healing was 4·1 ± 2·9 versus 2·4 ± 1·8 weeks (P = 0·039) in the biweekly versus weekly groups, respectively. Complete healing occurred in 50% versus 90% by 4 weeks in the biweekly and weekly groups, respectively (P = 0·014). Number of grafts applied to healed wounds was similar at 2·4 ± 1·5 and 2·3 ± 1·8 for biweekly versus weekly groups, respectively (P = 0·841). These results validate previous studies showing that the allograft is an effective treatment for diabetic ulcers and show that wounds treated with weekly application heal more rapidly than with biweekly application. More rapid healing may decrease clinical operational costs and prevent long-term medical complications. PMID:24618401

  19. Saffold Virus, a Human Cardiovirus, and Risk of Persistent Islet Autoantibodies in the Longitudinal Birth Cohort Study MIDIA.

    Science.gov (United States)

    Tapia, German; Bøås, Håkon; de Muinck, Eric J; Cinek, Ondrej; Stene, Lars C; Torjesen, Peter A; Rasmussen, Trond; Rønningen, Kjersti S

    2015-01-01

    The aim of this study was to describe the frequency and distribution of Saffold virus in longitudinal stool samples from children, and test for association with development of persistent autoantibodies predictive of type 1 diabetes. A cohort of Norwegian children carrying the HLA genotype associated with highest risk of type 1 diabetes ("DR4-DQ8/DR3-DQ2") was followed with monthly stool samples from 3 to 35 months of age. Blood samples were tested for autoantibodies to insulin, glutamic acid decarboxylase65 and Islet Antigen-2. 2077 stool samples from 27 children with ≥ 2 repeatedly positive islet autoantibodies (cases), and 53 matched controls were analysed for Saffold virus genomic RNA by semi-quantitative real-time reverse transcriptase PCR. Saffold virus was found in 53 of 2077 (2.6%) samples, with similar proportions between cases (2.5%) and controls (2.6%). The probability of being infected by 3 years of age was 28% (95% CI 0.18-0.40). Viral quantities ranged from Saffold virus had no statistically significant association with islet autoimmunity.

  20. Multiplexed color-coded probe-based gene expression assessment for clinical molecular diagnostics in formalin-fixed paraffin-embedded human renal allograft tissue.

    Science.gov (United States)

    Adam, Benjamin; Afzali, Bahman; Dominy, Katherine M; Chapman, Erin; Gill, Reeda; Hidalgo, Luis G; Roufosse, Candice; Sis, Banu; Mengel, Michael

    2016-03-01

    Histopathologic diagnoses in transplantation can be improved with molecular testing. Preferably, molecular diagnostics should fit into standard-of-care workflows for transplant biopsies, that is, formalin-fixed paraffin-embedded (FFPE) processing. The NanoString(®) gene expression platform has recently been shown to work with FFPE samples. We aimed to evaluate its methodological robustness and feasibility for gene expression studies in human FFPE renal allograft samples. A literature-derived antibody-mediated rejection (ABMR) 34-gene set, comprised of endothelial, NK cell, and inflammation transcripts, was analyzed in different retrospective biopsy cohorts and showed potential to molecularly discriminate ABMR cases, including FFPE samples. NanoString(®) results were reproducible across a range of RNA input quantities (r = 0.998), with different operators (r = 0.998), and between different reagent lots (r = 0.983). There was moderate correlation between NanoString(®) with FFPE tissue and quantitative reverse transcription polymerase chain reaction (qRT-PCR) with corresponding dedicated fresh-stabilized tissue (r = 0.487). Better overall correlation with histology was observed with NanoString(®) (r = 0.354) than with qRT-PCR (r = 0.146). Our results demonstrate the feasibility of multiplexed gene expression quantification from FFPE renal allograft tissue. This represents a method for prospective and retrospective validation of molecular diagnostics and its adoption in clinical transplantation pathology.

  1. A new approach for pancreatic tissue engineering: human endometrial stem cells encapsulated in fibrin gel can differentiate to pancreatic islet beta-cell.

    Science.gov (United States)

    Niknamasl, Azadeh; Ostad, Seyed Nasser; Soleimani, Mansoureh; Azami, Mahmoud; Salmani, Maryam Kabir; Lotfibakhshaiesh, Nasrin; Ebrahimi-Barough, Somayeh; Karimi, Roya; Roozafzoon, Reza; Ai, Jafar

    2014-10-01

    Metabolic diabetes mellitus as the most serious and prevalent metabolic disease in the world has various complications. The most effective treatment of type I diabetes seems to be islet cell transplantation. Shortage of donors and difficult procedures and high rate of rejection have always restricted this approach. Tissue engineering is a novel effective solution to many medical problems such as diabetes. Endometrial mesenchymal stem cells as a lineage which have the potential to differentiate to mesodermal and endodermal tissues seem to be suitable for this purpose. Fibrin hydrogel with a high degree of biocompatibility and specific properties making it similar to normal pancreas seems to be an ideal scaffold. After successfully isolating stem cells (hEnSCs) from human endometrium, a three-step protocol was used to differentiate them into pancreatic beta cells. Fibrin was used as 3D scaffold. After 2 weeks, cells formed clusters like islets cells, and secretion of insulin was measured by chemiluminescence. PDX1, proinsulin, and c-peptide as special markers of β cells were detected by immunofluorescence. Expression of glucagon, PDX1, and insulin genes in mRNA level was detected by Real time PCR and gel electrophoresis. The former showed higher levels of gene expression in 3D cultures. SEM analysis showed good integrity between cells and scaffold. No toxicity was detected with fibrin scaffold by MTT assay.

  2. Tat-biliverdin reductase A protects INS-1 cells from human islet amyloid polypeptide-induced cytotoxicity by alleviating oxidative stress and ER stress.

    Science.gov (United States)

    Lee, Su Jin; Kang, Hyung Kyung; Eum, Won Sik; Park, Jinseu; Choi, Soo Young; Kwon, Hyeok Yil

    2017-02-15

    Human islet amyloid polypeptide (hIAPP), a major constituent of islet amyloid deposits, induces pancreatic β-cell apoptosis and eventually contributes to β-cell deficit in patients with type 2 diabetes mellitus (T2DM). In this study, Tat-mediated transduction of biliverdin reductase A (BLVRA) was investigated in INS-1 cells to examine whether exogenous supplementation of BLVRA prevented hIAPP-induced apoptosis and dysfunction in insulin secretion in β-cells. Tat-BLVRA fusion protein was efficiently delivered into INS-1 cells in a time- and dose-dependent manner. Exposure of cells to hIAPP induced apoptotic cell death, which was dose-dependently inhibited by pre-treatment with Tat-BLVRA for 1 h. Transduced Tat-BLVRA reduced hIAPP-evoked generation of reactive oxygen species, a crucial mediator of β-cell destruction. Immunoblot analysis showed that Tat-BLVRA suppressed hIAPP-induced increase in the levels of proteins involved in endoplasmic reticulum (ER) stress and apoptosis signaling. Transduced Tat-BLVRA also recovered hIAPP-induced dysfunction in basal and glucose-stimulated insulin secretions. These results suggested that transduced Tat-BLVRA enhanced the tolerance of β-cells against IAPP-induced cytotoxicity by alleviating oxidative stress and ER stress. Therefore, Tat-mediated transduction of BLVRA may provide a potential tool to ameliorate β-cell deficit in pancreas with T2DM.

  3. Immature transformed rat islet beta-cells differentially express C-peptides derived from the genes coding for insulin I and II as well as a transfected human insulin gene

    DEFF Research Database (Denmark)

    Blume, N; Petersen, J S; Andersen, L C;

    1992-01-01

    Synthetic peptides representing unique sequences in rat proinsulin C-peptide I and II were used to generate highly specific antisera, which, when applied on sections of normal rat pancreas, confirm a homogeneous coexpression of the two C-peptides in all islet beta-cells. Insulin gene expression...... is induced in the transformed heterogeneous rat islet cell clone, NHI-6F, by transient in vivo passage. During this process a transfected human insulin gene is coactivated with the endogenous nonallelic rat insulin I and II genes. Newly established cultures from NHI-6F insulinomas having a high frequency...

  4. Differential interleukin-1 receptor antagonism on pancreatic beta and alpha cells. Studies in rodent and human islets and in normal rats

    DEFF Research Database (Denmark)

    Zumsteg, U; Reimers, J I; Pociot, F

    1993-01-01

    The monokines interleukin-1 alpha and -beta have been implicated as effector molecules in the immune-mediated pancreatic beta-cell destruction leading to insulin-dependent diabetes mellitus. Here we investigated the effects of interleukin-1 receptor antagonism on insulin and glucagon release of rat......, mouse and human islets exposed to recombinant human interleukin-1 beta, and on interleukin-1 beta induced changes in blood glucose, serum insulin and serum glucagon levels in Wistar Kyoto rats. The interleukin-1 receptor antagonist reduced the co-mitogenic effect of interleukin-1 beta on mouse and rat...... thymocytes with a 50% inhibitory concentration of 10- and 100-fold molar excess, respectively. Complete inhibition was obtained with a 100-1,000-fold molar excess. However, at a 100-fold molar excess the interleukin-1 receptor antagonist did not antagonise the potentiating effect of interleukin-1 beta on rat...

  5. Pancreatic beta cells and islets take up thiamin by a regulated carrier-mediated process: studies using mice and human pancreatic preparations

    Science.gov (United States)

    Mee, Lisa; Nabokina, Svetlana M.; Sekar, V. Thillai; Subramanian, Veedamali S.; Maedler, Kathrin; Said, Hamid M.

    2009-01-01

    Thiamin is essential for the normal function of the endocrine pancreas, but very little is known about uptake mechanism(s) and regulation by beta cells. We addressed these issues using mouse-derived pancreatic beta-TC-6 cells, and freshly isolated primary mouse and human pancreatic islets. Results showed that thiamin uptake by beta-TC-6 cells involves a pH (but not Na+)-dependent carrier-mediated process that is saturable at both the nanomolar (apparent Km = 37.17 ± 9.9 nM) and micromolar (apparent Km = 3.26 ± 0.86 μM) ranges, cis-inhibited by thiamin structural analogs, and trans-stimulated by unlabeled thiamin. Involvement of carrier-mediated process was also confirmed in primary mouse and human pancreatic islets. Both THTR-1 and THTR-2 were found to be expressed in these mouse and human pancreatic preparations. Maintaining beta-TC-6 cells in the presence of a high level of thiamin led to a significant (P < 0.01) decrease in thiamin uptake, which was associated with a significant downregulation in level of expression of THTR-1 and THTR-2 at the protein and mRNA levels and a decrease in transcriptional (promoter) activity. Modulators of intracellular Ca2+/calmodulin- and protein-tyrosine kinase-mediated pathways also altered thiamin uptake. Finally, confocal imaging of live beta-TC-6 cells showed that clinical mutants of THTR-1 have mixed expression phenotypes and all led to impairment in thiamin uptake. These studies demonstrate for the first time that thiamin uptake by the endocrine pancreas is carrier mediated and is adaptively regulated by the prevailing vitamin level via transcriptional mechanisms. Furthermore, clinical mutants of THTR-1 impair thiamin uptake via different mechanisms. PMID:19423748

  6. 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 < 0.001). Comparisons of individual methods showed good correlations between mean values of IEQ number (r(2) = 0.91) and total islet number (r(2) = 0.88) and thus increased to r(2) = 0.93 when islet surface area was estimated comparatively with IEQ number. The ADIA method showed very high intraobserver reproducibility compared to the standard manual method (p < 0.001). However, islet purity was routinely estimated as significantly higher with the manual method versus the ADIA method (p < 0.001). The ADIA method also detected small islets between 10 and 50 µm in size. Automated digital 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

  7. Towards engineering a novel transplantation site for pancreatic islets

    NARCIS (Netherlands)

    Smink, Alexandra Maria

    2016-01-01

    Intraportal pancreatic islet transplantation is a promising therapy for type 1 diabetes, but the liver is not an optimal site as it is associated with massive cell-death in the graft. Several alternative sites were investigated, but the human body does not contain an adequate islet transplantation s

  8. Pancreatic Islet Survival and Engraftment Is Promoted by Culture on Functionalized Spider Silk Matrices.

    Science.gov (United States)

    Johansson, Ulrika; Ria, Massimiliano; Åvall, Karin; Dekki Shalaly, Nancy; Zaitsev, Sergei V; Berggren, Per-Olof; Hedhammar, My

    2015-01-01

    Transplantation of pancreatic islets is one approach for treatment of diabetes, however, hampered by the low availability of viable islets. Islet isolation leads to disruption of the environment surrounding the endocrine cells, which contributes to eventual cell death. The reestablishment of this environment is vital, why we herein investigated the possibility of using recombinant spider silk to support islets in vitro after isolation. The spider silk protein 4RepCT was formulated into three different formats; 2D-film, fiber mesh and 3D-foam, in order to provide a matrix that can give the islets physical support in vitro. Moreover, cell-binding motifs from laminin were incorporated into the silk protein in order to create matrices that mimic the natural cell environment. Pancreatic mouse islets were thoroughly analyzed for adherence, necrosis and function after in vitro maintenance on the silk matrices. To investigate their suitability for transplantation, we utilized an eye model which allows in vivo imaging of engraftment. Interestingly, islets that had been maintained on silk foam during in vitro culture showed improved revascularization. This coincided with the observation of preserved islet architecture with endothelial cells present after in vitro culture on silk foam. Selected matrices were further evaluated for long-term preservation of human islets. Matrices with the cell-binding motif RGD improved human islet maintenance (from 36% to 79%) with preserved islets architecture and function for over 3 months in vitro. The islets established cell-matrix contacts and formed vessel-like structures along the silk. Moreover, RGD matrices promoted formation of new, insulin-positive islet-like clusters that were connected to the original islets via endothelial cells. On silk matrices with islets from younger donors (<35 year), the amount of newly formed islet-like clusters found after 1 month in culture were almost double compared to the initial number of islets

  9. Pancreatic Islet Survival and Engraftment Is Promoted by Culture on Functionalized Spider Silk Matrices.

    Directory of Open Access Journals (Sweden)

    Ulrika Johansson

    Full Text Available Transplantation of pancreatic islets is one approach for treatment of diabetes, however, hampered by the low availability of viable islets. Islet isolation leads to disruption of the environment surrounding the endocrine cells, which contributes to eventual cell death. The reestablishment of this environment is vital, why we herein investigated the possibility of using recombinant spider silk to support islets in vitro after isolation. The spider silk protein 4RepCT was formulated into three different formats; 2D-film, fiber mesh and 3D-foam, in order to provide a matrix that can give the islets physical support in vitro. Moreover, cell-binding motifs from laminin were incorporated into the silk protein in order to create matrices that mimic the natural cell environment. Pancreatic mouse islets were thoroughly analyzed for adherence, necrosis and function after in vitro maintenance on the silk matrices. To investigate their suitability for transplantation, we utilized an eye model which allows in vivo imaging of engraftment. Interestingly, islets that had been maintained on silk foam during in vitro culture showed improved revascularization. This coincided with the observation of preserved islet architecture with endothelial cells present after in vitro culture on silk foam. Selected matrices were further evaluated for long-term preservation of human islets. Matrices with the cell-binding motif RGD improved human islet maintenance (from 36% to 79% with preserved islets architecture and function for over 3 months in vitro. The islets established cell-matrix contacts and formed vessel-like structures along the silk. Moreover, RGD matrices promoted formation of new, insulin-positive islet-like clusters that were connected to the original islets via endothelial cells. On silk matrices with islets from younger donors (<35 year, the amount of newly formed islet-like clusters found after 1 month in culture were almost double compared to the initial

  10. Transcriptional profiling of type 1 diabetes genes on chromosome 21 in a rat beta-cell line and human pancreatic islets

    DEFF Research Database (Denmark)

    Bergholdt, R.; Karlsen, A.E.; Hagedorn, Peter;

    2007-01-01

    We recently finemapped a type 1 diabetes (T1D)-linked region on chromosome 21, indicating that one or more T1D-linked genes exist in this region with 33 annotated genes. In the current study, we have taken a novel approach using transcriptional profiling in predicting and prioritizing the most...... likely candidate genes influencing beta-cell function in this region. Two array-based approaches were used, a rat insulinoma cell line (INS-1alphabeta) overexpressing pancreatic duodenum homeobox 1 (pdx-1) and treated with interleukin 1beta (IL-1beta) as well as human pancreatic islets stimulated...... with a mixture of cytokines. Several candidate genes with likely functional significance in T1D were identified. Genes showing differential expression in the two approaches were highly similar, supporting the role of these specific gene products in cytokine-induced beta-cell damage. These were genes involved...

  11. PDZ-domain containing-2 (PDZD2) drives the maturity of human fetal pancreatic progenitor-derived islet-like cell clusters with functional responsiveness against membrane depolarization.

    Science.gov (United States)

    Leung, Kwan Keung; Suen, Po Man; Lau, Tse Kin; Ko, Wing Hung; Yao, Kwok Ming; Leung, Po Sing

    2009-09-01

    We recently reported the isolation and characterization of a population of pancreatic progenitor cells (PPCs) from early trimester human fetal pancreata. The PPCs, being the forerunners of adult pancreatic cell lineages, were amenable to growth and differentiation into insulin-secreting islet-like cell clusters (ICCs) upon stimulation by adequate morphogens. Of note, a novel morphogenic factor, PDZ-domain containing-2 (PDZD2) and its secreted form (sPDZD2) were ubiquitously expressed in the PPCs. Our goals for this study were to evaluate the potential role of sPDZD2 in stimulating PPC differentiation and to establish the optimal concentration for such stimulation. We found that 10(-9)M sPDZD2 promoted PPC differentiation, as evidenced by the upregulation of the pancreatic endocrine markers (PDX-1, NGN3, NEURO-D, ISL-1, NKX 2.2, NKX 6.1) and INSULIN mRNA. Inhibited endogenous production of sPDZD2 suppressed expression of these factors. Secreted PDZD2 treatment significantly elevated the C-peptide content of the ICCs and increased the basal rate of insulin secretion. However, they remained unresponsive to glucose stimulation, reflected by a minimal increase in GLUT-2 and GLUCOKINASE mRNA expression. Interestingly, sPDZD2 treatment induced increased expression of the L-type voltage-gated calcium channel (Ca(v)1.2) in the ICCs, triggering calcium ion influx under KCl stimulation and conferring an ability to secrete insulin in response to KCl. Pancreatic progenitor cells from 10- and 13-week fetal pancreata showed peak expression of endogenous sPDZD2, implying that sPDZD2 has a specific role in islet development during the first trimester. In conclusion, our data suggest that sPDZD2 promotes functional maturation of human fetal PPC-derived ICCs, thus enhancing its transplanting potentials.

  12. The morphology of islets within the porcine donor pancreas determines the isolation result: successful isolation of pancreatic islets can now be achieved from young market pigs.

    Science.gov (United States)

    Krickhahn, Mareike; Bühler, Christoph; Meyer, Thomas; Thiede, Arnulf; Ulrichs, Karin

    2002-01-01

    Clinical islet allotransplantation has become an increasingly efficient "routine" therapy in recent years. Shortage of human donor organs leads to porcine pancreatic islets as a potential source for islet xenotransplantation. Yet it is still very difficult to isolate sufficient numbers of intact porcine islets, particularly from young market pigs. In the following study islets were successfully isolated from retired breeders [4806 +/- 720 islet equivalents per gram organ (IEQ/g); n = 25; 2-3 years old; RB] and also from young hybrid pigs [2868 +/- 260 IEQ/g; n = 65; 4-6 months old; HY] using LiberasePI and a modified version of Ricordi's digestion-filtration technique. As expected, isolations from RB showed significantly better results (p organs from RB (80%) contained mainly large islets (diameter > 200 microm), in contrast to only 35% of all pancreases from HY. Remarkably, the islet size in situ, regardless whether detected in RB or HY, strongly determined the isolation result. A donor organ with predominantly large islets resulted in significantly higher numbers of IEQs compared with a donor organ with predominantly small islets [RB(Large Islets): 5680 +/- 3,318 IEQ/g (n= 20); RB(Small Islets): 1353 +/- 427 IEQ/g (n = 5); p organ prior to the isolation process. Under these conditions highly successful isolations can reliably be performed even from young market pigs.

  13. Exocrine contamination impairs implantation of pancreatic islets transplanted beneath the kidney capsule.

    Science.gov (United States)

    Gray, D W; Sutton, R; McShane, P; Peters, M; Morris, P J

    1988-11-01

    The effect of exocrine contamination on islets implanted under the kidney capsule has been studied by histological examination of pure or exocrine-contamination human, monkey, or rat islets transplanted to the kidney capsule of the nude rat, monkey, or rat, respectively. Exocrine contamination resulted in an appearance suggestive of impaired islet implantation, due to tissue necrosis and subsequent fibrosis. The effect of exocrine contamination was examined quantitatively in a rat islet isograft model in which handpicked DA rat islets were transplanted under the kidney capsule of normal DA rats. The islets were either pure or deliberately recontaminated with exocrine tissue (50 or 90% contamination). Four hundred pure islets were placed under one kidney capsule and 400 islets (of similar size and from the same islet preparation) were contaminated and then placed under the contralateral kidney capsule. After 2 weeks the kidneys were removed and extracted for insulin content. The insulin content of kidneys bearing islets contaminated by either 50 or 90% exocrine tissue was significantly reduced when compared to the contralateral kidney bearing pure islets. These findings support the view that exocrine contamination of islets resulted in impaired islet implantation when transplanted to a confined site such as the kidney subcapsule.

  14. Ultrastructural studies of time-course and cellular specificity of interleukin-1 mediated islet cytotoxicity

    DEFF Research Database (Denmark)

    Mandrup-Poulsen, T; Egeberg, J; Nerup, J

    1987-01-01

    Previous electron-microscopic studies of isolated islets of Langerhans exposed to the monokine interleukin-1 for 7 days have indicated that interleukin-1 is cytotoxic to all islet cells. To study the time-course and possible cellular specificity of interleukin-1 cytotoxicity to islets exposed...... to interleukin-1 for short time periods, isolated rat or human islets were incubated with or without 25 U/ml highly purified human interleukin-1 for 24 h. Samples of rat islets were taken after 5 min, 30 min, 1, 2, 4, 6, 8, 10, 12, 16, 20 and 24 h and samples of human islets after 5 min, 30 min and 24 h...... of incubation and examined by electron microscopy in a blinded fashion. Already after 30 min, accumulation of opaque intracytoplasmic bodies without apparent surrounding membranes, and autophagic vacuoles were seen in about 20% of the beta cells examined in rat islets exposed to interleukin-1. After 16 h...

  15. Renal Structural Changes after Kidney Allograft Transplantation

    NARCIS (Netherlands)

    Bakker, R.C.

    2005-01-01

    In vitro studies done on human proximal tubular epithelial cells showed no direct cytotoxicity of cyclosporine A. The 15-year results of an open randomized trial comparing cyclosporine withdrawal and conversion to azathioprine with continued cyclosporine treatment after kidney allograft transplantat

  16. Cathelicidin Antimicrobial Peptide: A Novel Regulator of Islet Function, Islet Regeneration, and Selected Gut Bacteria.

    Science.gov (United States)

    Pound, Lynley D; Patrick, Christopher; Eberhard, Chandra E; Mottawea, Walid; Wang, Gen-Sheng; Abujamel, Turki; Vandenbeek, Roxanne; Stintzi, Alain; Scott, Fraser W

    2015-12-01

    Cathelicidin antimicrobial peptide (CAMP) is a naturally occurring secreted peptide that is expressed in several organs with pleiotropic roles in immunomodulation, wound healing, and cell growth. We previously demonstrated that gut Camp expression is upregulated when type 1 diabetes-prone rats are protected from diabetes development. Unexpectedly, we have also identified novel CAMP expression in the pancreatic β-cells of rats, mice, and humans. CAMP was present even in sterile rat embryo islets, germ-free adult rat islets, and neogenic tubular complexes. Camp gene expression was downregulated in young BBdp rat islets before the onset of insulitis compared with control BBc rats. CAMP treatment of dispersed islets resulted in a significant increase in intracellular calcium mobilization, an effect that was both delayed and blunted in the absence of extracellular calcium. Additionally, CAMP treatment promoted insulin and glucagon secretion from isolated rat islets. Thus, CAMP is a promoter of islet paracrine signaling that enhances islet function and glucoregulation. Finally, daily treatment with the CAMP/LL-37 peptide in vivo in BBdp rats resulted in enhanced β-cell neogenesis and upregulation of potentially beneficial gut microbes. In particular, CAMP/LL-37 treatment shifted the abundance of specific bacterial populations, mitigating the gut dysbiosis observed in the BBdp rat. Taken together, these findings indicate a novel functional role for CAMP/LL-37 in islet biology and modification of gut microbiota. © 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

  17. Radiation sterilization of skin allograft

    Science.gov (United States)

    Kairiyama, E.; Horak, C.; Spinosa, M.; Pachado, J.; Schwint, O.

    2009-07-01

    In the treatment of burns or accidental loss of skin, cadaveric skin allografts provide an alternative to temporarily cover a wounded area. The skin bank facility is indispensable for burn care. The first human skin bank was established in Argentina in 1989; later, 3 more banks were established. A careful donor selection is carried out according to the national regulation in order to prevent transmissible diseases. As cadaveric human skin is naturally highly contaminated, a final sterilization is necessary to reach a sterility assurance level (SAL) of 10 -6. The sterilization dose for 106 batches of processed human skin was determined on the basis of the Code of Practice for the Radiation Sterilization of Tissue Allografts: Requirements for Validation and Routine Control (2004) and ISO 11137-2 (2006). They ranged from 17.6 to 33.4 kGy for bioburdens of >10-162.700 CFU/100 cm 2. The presence of Gram negative bacteria was checked for each produced batch. From the analysis of the experimental results, it was observed that the bioburden range was very wide and consequently the estimated sterilization doses too. If this is the case, the determination of a tissue-specific dose per production batch is necessary to achieve a specified requirement of SAL. Otherwise if the dose of 25 kGy is preselected, a standardized method for substantiation of this dose should be done to confirm the radiation sterilization process.

  18. Comparison of the regenerative potential of an allograft used alone and that in conjunction with an immunosuppressive drug in the treatment of human periodontal infrabony defects: A clinical and radiological study

    Directory of Open Access Journals (Sweden)

    Dhawan Shivani

    2010-01-01

    Full Text Available Background: Experimentation confirmed the conclusion that bone allografts, like other tissue and organ allografts, are immunogenic. These immune responses cause resorption of allograft, thus lowering the bone formation capacity of the graft. An attempt has been made in this study to prevent immune reactions and achieve enhanced regeneration of allograft-demineralized freeze-dried bone matrix by incorporating it with an immunosuppressive drug Cyclosporine-A (CsA in the treatment of human periodontal infrabony defects. Materials and Methods: Fifteen patients showing clinical evidence of almost bilateral infrabony defects requiring bone grafting procedures were selected. In each patient, the infrabony defect of one side of the arch was designated as Group A (control site and the infrabony defect of the contralateral side of the same arch was designated as Group B (test site. Results: On comparative evaluation of the two groups (by Student t-test, the mean values of reduction in probing depth (P=0.81 NS and gain in clinical attachment level (P=1.00 NS of Group B were found to be greater than that of Group-A, but the differences were statistically non-significant. The mean linear bone fill (P=0.010 FNx08 of Group B was also detected to be higher than that of Group A, and the difference was found to be statistically significant. Conclusion: Increase in linear bone fill in Group B signifies the role of CsA in augmenting the regenerative potential of allograft by eliminating immune reactions.

  19. Pancreas procurement from multiorgan donors for islet trasplantation.

    Science.gov (United States)

    Ricordi, C; Mazzeferro, V; Casavilla, A; Scotti, C; Pinna, A; Tzakis, A; Starzl, T E

    1992-01-01

    The outcome of human islet isolation procedures can be significantly effected by the technique used for pancreas procurement. In fact, the final step of islet purification using discontinuous density gradients requires a significant difference between the density of the islets and the density of the non-endocrine component of the gland. Therefore, any procedure during multi-organ procurement that will result in edema or degranulation of the acinar tissue will result in failure of the islet purification step. In this report a technique for combined harvesting of liver and pancreas is presented. The use of this procedure can be of assistance to avoid damage to the pancreas that could result in a compromised islet purification for improper handling of the gland even before it arrives to the isolation facility.

  20. Islet transplantation and antioxidant management A comprehensive review

    Institute of Scientific and Technical Information of China (English)

    Seyed Sajad Mohseni Salehi Monfared; Bagher Larijani; Mohammad Abdollahi

    2009-01-01

    Islet transplantation as a promising treatment for type 1 diabetes has received widespread attention.Oxidative stress plays an essential role in cell injury during islet isolation and transplantation procedures.Antioxidants have been used in various studies to improve islet transplantation procedures. The present study reviews the role of oxidative stress and the benefits of antioxidants in islet transplantation procedures. The bibliographical databases Pubmed and Scopus were searched up to November 2008.All relevant human and animal in-vivo and in-vitro studies, which investigated antioxidants on islets,were included. Almost all the tested antioxidants used in the in-vitro studies enhanced islet viability and insulin secretion. Better control of blood glucose after transplantation was the major outcome of antioxidant therapy in all in-vivo studies. The data also indicated that antioxidants improved islet transplantation procedures. Although there is still insufficient evidence to draw definitive conclusions about the efficacy of individual supplements, the benefits of antioxidants in islet isolation procedures cannot be ignored.

  1. 21 CFR 862.1163 - Cardiac allograft gene expression profiling test system.

    Science.gov (United States)

    2010-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1163 Cardiac allograft gene expression profiling test system....

  2. Mild exposure of RIN-5F β-cells to human islet amyloid polypeptide aggregates upregulates antioxidant enzymes via NADPH oxidase-RAGE: An hormetic stimulus

    Directory of Open Access Journals (Sweden)

    Elisabetta Borchi

    2014-01-01

    Full Text Available The presence of amyloid aggregates of human islet amyloid polypeptide (hIAPP, a hallmark of type 2 diabetes, contributes to pancreatic β-cell impairment, where oxidative stress plays a key role. A contribution of NADPH oxidase to reactive oxygen species (ROS generation after cell exposure to micromolar concentrations of hIAPP aggregates has been suggested. However, little is known about β-cells exposure to lower amounts of hIAPP aggregates, similar to those found in human pancreas. Thus, we aimed to investigate the events resulting from RIN-5F cells exposure to nanomolar concentrations of toxic hIAPP aggregates. We found an early and transient rise of NADPH oxidase activity resulting from increased Nox1 expression following the engagement of receptor for advanced glycation end-products (RAGE by hIAPP aggregates. Unexpectedly, NADPH oxidase activation was not accompanied by a significant ROS increase and the lipoperoxidation level was significantly reduced. Indeed, cell exposure to hIAPP aggregates affected the antioxidant defences, inducing a significant increase of the expression and activity of catalase and glutathione peroxidase. We conclude that exposure of pancreatic β-cells to nanomolar concentrations of hIAPP aggregates for a short time induces an hormetic response via the RAGE-Nox1 axis; the latter stimulates the enzymatic antioxidant defences that preserve the cells against oxidative stress damage.

  3. The International Xenotransplantation Association consensus statement on conditions for undertaking clinical trials of porcine islet products in type 1 diabetes--chapter 6: Patient selection for pilot clinical trials of islet xenotransplantation.

    Science.gov (United States)

    O'Connell, Philip J

    2009-01-01

    Several clinical scenarios would justify trials of islet xenotransplantation, including islet after kidney (IAK) transplantation and islet xenotransplantation for hypoglycemia unawareness. In selecting a patient cohort, several factors unique to xenotransplant trials must be considered, including the potential for unique infective complications, the risks of immunosuppression, and the requirement for life-long monitoring. These factors need to be balanced against the benefits of improved glycemic control and ability to deliver large numbers of high-quality islets, something more difficult to achieve in islet allotransplantation. The importance of these risks varies depending on the patient cohort selected and proposed therapy to be undertaken. In islet xenotransplantation for hypoglycemia unawareness, immunosuppressive risks, and problems with life-long monitoring need special attention in trial design. Whereas in IAK xenotransplantation, consideration must be given to ensure that the procedure does not impact on renal allograft outcomes and patient co-morbidity, which is already significant in renal transplant recipients. This review outlines the rationale for undertaking islet xenotransplantation in highly select patients with type 1 diabetes and addresses some of the unique challenges. At present, no clear consensus exists as to the most appropriate patient group for the first trials of islet xenotransplantation. Rather, this will depend on the type of treatment being proposed, the potential infective risks and the overall burden of immunosuppression.

  4. Type I and II Diabetic Adipose-Derived Stem Cells Respond In Vitro to Dehydrated Human Amnion/Chorion Membrane Allograft Treatment by Increasing Proliferation, Migration, and Altering Cytokine Secretion

    OpenAIRE

    Massee, Michelle; Chinn, Kathryn; Lim, Jeremy J; Godwin, Lisa; Young, Conan S.; Koob, Thomas J.

    2016-01-01

    Objective: Human amniotic membranes have been shown to be effective for healing diabetic foot ulcers clinically and to regulate stem cell activity in vitro and in vivo; however, diabetic stem cells may be impaired as a sequela of the disease. In this study, dehydrated human amnion/chorion membrane (dHACM) allografts (EpiFix®; MiMedx Group) were evaluated for their ability to regulate diabetic stem cells in vitro. Approach: Human adipose-derived stem cells (ADSCs) from normal, type I diabetic,...

  5. Renalase Gene Polymorphism in Patients After Renal Allograft Transplantation

    Directory of Open Access Journals (Sweden)

    Andrzej Pawlik

    2014-06-01

    Full Text Available Background/Aims: Renalase is a recently discovered protein, which is likely involved in regulation of blood pressure in humans and animals. Previous studies suggest that renalase reflects kidney functioning. A common missense polymorphism in the flavin-adenine dinucleotide-binding domain of human renalase (Glu37Asp has been described. In this study we examined the association between (Glu37Asp polymorphism (rs2296545 in renalase gene and kidney allograft function. Methods: The study enrolled 270 Caucasian kidney allograft recipients. SNP within the renalase was genotyped using TaqMan genotyping assays. Results: There were no statistically significant associations between renalase gene rs2296545 polymorphism and delayed graft function, acute rejection, chronic allograft dysfunction as well as creatinine serum concentrations and blood pressure values after transplantation. Conclusions: The results of this study suggest, that renalase gene rs2296545 polymorphism is not important factor determining renal allograft function.

  6. Bisphenol A accelerates toxic amyloid formation of human islet amyloid polypeptide: a possible link between bisphenol A exposure and type 2 diabetes.

    Science.gov (United States)

    Gong, Hao; Zhang, Xin; Cheng, Biao; Sun, Yue; Li, Chuanzhou; Li, Ting; Zheng, Ling; Huang, Kun

    2013-01-01

    Bisphenol A (BPA) is a chemical compound widely used in manufacturing plastic products. Recent epidemiological studies suggest BPA exposure is positively associated with the incidence of type 2 diabetes mellitus (T2DM), however the mechanisms underlying this link remain unclear. Human islet amyloid polypeptide (hIAPP) is a hormone synthesized and secreted by the pancreatic β-cells. Misfolding of hIAPP into toxic oligomers and mature fibrils can disrupt cell membrane and lead to β-cell death, which is regarded as one of the causative factors of T2DM. To test whether there are any connections between BPA exposure and hIAPP misfolding, we investigated the effects of BPA on hIAPP aggregation using thioflavin-T based fluorescence, transmission electronic microscopy, circular dichroism, dynamic light scattering, size-exclusion chromatography, fluorescence-dye leakage assay in an artificial micelle system and the generation of reactive oxygen species in INS-1 cells. We demonstrated that BPA not only dose-dependently promotes the aggregation of hIAPP and enhances the membrane disruption effects of hIAPP, but also promotes the extent of hIAPP aggregation related oxidative stress. Taken together, our results suggest that BPA exposure increased T2DM risk may involve the exacerbated toxic aggregation of hIAPP.

  7. Targeted Mass Spectrometry Approach Enabled Discovery of O-Glycosylated Insulin and Related Signaling Peptides in Mouse and Human Pancreatic Islets.

    Science.gov (United States)

    Yu, Qing; Canales, Alejandra; Glover, Matthew S; Das, Rahul; Shi, Xudong; Liu, Yang; Keller, Mark P; Attie, Alan D; Li, Lingjun

    2017-08-07

    O-Linked glycosylation often involves the covalent attachment of sugar moieties to the hydroxyl group of serine or threonine on proteins/peptides. Despite growing interest in glycoproteins, little attention has been directed to glycosylated signaling peptides, largely due to lack of enabling analytical tools. Here we explore the occurrence of naturally O-linked glycosylation on the signaling peptides extracted from mouse and human pancreatic islets using mass spectrometry (MS). A novel targeted MS-based method is developed to increase the likelihood of capturing these modified signaling peptides and to provide improved sequence coverage and accurate glycosite localization, enabling the first large-scale discovery of O-glycosylation on signaling peptides. Several glycosylated signaling peptides with multiple glycoforms are identified, including the first report of glycosylated insulin-B chain and insulin-C peptide and BigLEN. This discovery may reveal potential novel functions as glycosylation could influence their conformation and biostability. Given the importance of insulin and its related peptide hormones and previous studies of glycosylated insulin analogues, this natural glycosylation may provide important insights into diabetes research and therapeutic treatments.

  8. Effects of cholesterol on pore formation in lipid bilayers induced by human islet amyloid polypeptide fragments: A coarse-grained molecular dynamics study

    Science.gov (United States)

    Xu, Weixin; Wei, Guanghong; Su, Haibin; Nordenskiöld, Lars; Mu, Yuguang

    2011-11-01

    Disruption of the cellular membrane by the amyloidogenic peptide, islet amyloid polypeptide (IAPP), has been considered as one of the mechanisms of β-cell death during type 2 diabetes. The N-terminal region (residues 1-19) of the human version of IAPP is suggested to be primarily responsible for the membrane-disrupting effect of the full-length hIAPP peptide. However, the detailed assembly mode of hIAPP1-19 with membrane remains unclear. To gain insight into the interactions of hIAPP1-19 oligomer with the model membrane, we have employed coarse-grained molecular dynamics self-assembly simulations to study the aggregation of hIAPP1-19 fragments in the binary lipid made of zwitterionic dipalmitoylphosphatidylcholine (DPPC) and anionic dipalmitoylphosphatidylserine (DPPS) in the presence and absence of different levels of cholesterol content. The membrane-destabilizing effect of hIAPP1-19 is found to be modulated by the presence of cholesterol. In the absence of cholesterol, hIAPP1-19 aggregates prefer to locate inside the bilayer, forming pore-like assemblies. While in the presence of cholesterol molecules, the lipid bilayer becomes more ordered and stiff, and the hIAPP1-19 aggregates are dominantly positioned at the bilayer-water interface. The action of cholesterol may suggest a possible way to maintain the membrane integrity by small molecule interference.

  9. Targeted delivery of interleukin-10 to chronic cardiac allograft rejection using a human antibody specific to the extra domain A of fibronectin.

    Science.gov (United States)

    Franz, Marcus; Doll, Fabia; Grün, Katja; Richter, Petra; Köse, Nilay; Ziffels, Barbara; Schubert, Harald; Figulla, Hans R; Jung, Christian; Gummert, Jan; Renner, André; Neri, Dario; Berndt, Alexander

    2015-09-15

    Management of chronic rejection is challenging since there are not sufficient preventive or therapeutic strategies. The rejection process leads to overexpression of ED-A(+) fibronectin (ED-A(+) Fn). The human antibody F8, specific to ED-A(+) Fn, may serve as a vehicle for targeted delivery of bioactive payloads, e.g. interleukin 10 (IL-10). The aim of this study was to investigate the therapeutic effects of the fusion protein F8-interleukin-10 (F8-IL10) in the process of chronic rejection development. A heterotopic rat heart transplantation model was used to induce chronic rejection. For therapeutic interventions, the immunocytokines F8-humanIL10 (DEKAVIL), F8-ratIL10 as well as KSF-humanIL10 (irrelevant antigen-specificity) were used. Treatment was performed weekly for 10 weeks starting at day 7 after transplantation (1mg/animal). In the cardiac allografts, treatment with F8-huIL10 or F8-ratIL10 was associated with increased heart weights, a higher grade of chronic rejection, increased CIF, higher protein expression levels of alpha-smooth muscle actin (α-SMA), an augmented infiltration with inflammatory cells (CD4+, CD8+ and CD68+ cells) and higher serum levels of brain natriuretic peptide (BNP) compared to the control groups. All observed treatment effects are transplantation-specific since the F8 antibody is specific to ED-A(+) Fn that is not expressed in healthy hearts. A clear targeting effect of F8-huIL10 as well as F8-ratIL10 could be proven. Against that background, a further study is needed to address the question, if F8-IL10 treatment is capable to reduce CAV and CIF starting at a time point when chronic rejection has fully developed (therapeutic approach). Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  10. Microencapsulation of pancreatic islets for use in a bioartificial pancreas.

    Science.gov (United States)

    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.

  11. B7-H4 Pathway in Islet Transplantation and β-Cell Replacement Therapies

    Directory of Open Access Journals (Sweden)

    Xiaojie Wang

    2011-01-01

    Full Text Available Type 1 diabetes (T1D is a chronic autoimmune disease and characterized by absolute insulin deficiency. β-cell replacement by islet cell transplantation has been established as a feasible treatment option for T1D. The two main obstacles after islet transplantation are alloreactive T-cell-mediated graft rejection and recurrence of autoimmune diabetes mellitus in recipients. T cells play a central role in determining the outcome of both autoimmune responses and allograft survival. B7-H4, a newly identified B7 homolog, plays a key role in maintaining T-cell homeostasis by reducing T-cell proliferation and cytokine production. The relationship between B7-H4 and allograft survival/autoimmunity has been investigated recently in both islet transplantation and the nonobese diabetic (NOD mouse models. B7-H4 protects allograft survival and generates donor-specific tolerance. It also prevents the development of autoimmune diabetes. More importantly, B7-H4 plays an indispensable role in alloimmunity in the absence of the classic CD28/CTLA-4 : B7 pathway, suggesting a synergistic/additive effect with other agents such as CTLA-4 on inhibition of unwanted immune responses.

  12. Role of the Galectin-7 expression in cardiac allografts in human and mouse%Galectin-7在人和鼠同种异体心脏移植物中的表达及其意义

    Institute of Scientific and Technical Information of China (English)

    雒真龙; 周鸿敏; 黄霞; 方静; 徐洪来; 高义; 杨超; 潘铁成; 陈忠华

    2013-01-01

    Objective To explore the function of galectin-7 in acute rejection through detecting the expression galectin-7 in cardiac allografts in human and mouse.Methods Mouse models of cardiac allografts and isografts were constructed.Western blot and Immunohistochemistry (IHC) were used for detecting galectin-7 expression in the grafts.IHC was used for detecting galectin-7 expression in nyocardial tissues of 15 heart transplant patients with acute rejection and 10 normal individuals.Results The expression of galectin-7 in mouse allografts was significantly higher than that in isografts (1.34 ± 0.18,0.77 ± 0.03,P < 0.01).Galectin-7 mainly located in lymphocytes and vascular endothelial cells in human allografts.Conclusion Galectin-7 is related to the acute rejection in human and mouse cardiac allografts.%目的:通过检测发生急性排斥反应的小鼠及人移植心脏的galectin-7表达,进而分析galectin-7在急排发生过程中的作用.方法:建立小鼠心脏移植模型,分为鼠急排组和鼠对照组.Western blot法和免疫组化法检测小鼠移植心脏galectin-7的表达情况.选择15例心脏移植急排患者(人急排组)和10例正常人(人对照组),免疫组化法检测心肌组织galectin-7的表达.结果:鼠急排组gatectin-7表达明显高于鼠对照组(1.34±0.18,0.77±0.03,P< 0.01).人急排组心脏galectin-7表达明显高于人对照组,且galectin-7主要定位于人急排心脏的浸润淋巴细胞及血管内皮细胞.结论:Galectin-7与人和鼠同种异体心脏移植急性排斥反应有相关性.

  13. The influence of intraperitoneal transplantation of free and encapsulated Langerhans islets on the second set phenomenon.

    Science.gov (United States)

    Orłowski, Tadeusz; Godlewska, Ewa; Mościcka, Maria; Sitarek, Elzbieta

    2003-12-01

    To protect the allografts or xenografts against transplant rejection special semipermeable membranes are applied. So far, there are only a few studies on the influence of an immunoisolated graft on the recipient immune system. Therefore, the possibility that an intraperitoneally grafted alginate/poly L-lysine/alginate (APA) coated pancreatic islets graft can effectively sensitize the recipient and provoke second set phenomenon was studied. C3H male mice and male WAG rats were used as donors of full-thickness skin and of free or encapsulated islet intraperitoneal grafts. Male BALB/c mice served as recipients. Skin grafts were performed following the method of Billingham and Medawar. The length of the second skin graft survival time served as the criterion for the sensitizing capacity of the primary graft. APA encapsulation of islets delayed but has not prevented the development of the second set phenomenon. However, the second skin graft rejection time was significantly longer after grafting of encapsulated islets than after free islets transplantation. APA microencapsulation of intraperitoneally transplanted islets delayed but did not prevent the development of the second set phenomenon. Encapsulation does not ensure complete immunoisolation, but only creates "an artificially immunoprivileged site of transplantation."

  14. Pig islets for islet xenotransplantation: current status and future perspectives

    Institute of Scientific and Technical Information of China (English)

    Hu Qinghua; Liu Zhongwei; Zhu Haitao

    2014-01-01

    Objective To review the current status and progress on pig islet xenotransplantation.Data sources Data used in this review were mainly from English literature of Pubmed database.The search terms were "pig islet" and "xenotransplantation".Study selection The original articles and critical reviews selected were relevant to this review's theme.Results Pigs are suggested to be an ideal candidate for obtaining available islet cells for transplantation.However,the potential clinical application of pig islet is still facing challenges including inadequate yield of high-quality functional islets and xenorejection of the transplants.The former can be overcome mainly by selection of a suitable pathogen-free source herd and the development of isolation and purification technology.While the feasibility of successful preclinical pig islet xenotranplantation provides insights in the possible mechanisms of xenogeneic immune recognition and rejection to overwhelm the latter.In addition,the achievement of long-term insulin independence in diabetic models by means of distinct islet products and novel immunotherapeutic strategies is promising.Conclusions Pig islet xenotransplantation is one of the prospective treatments to bridge the gap between the needs of transplantation in patients with diabetes and available islet cells.Nonetheless,further studies and efforts are needed to translate obtained findings into tangible applications.

  15. Hemo oxygenase-1 induction in vitro and in vivo can yield pancreas islet xenograft survival and improve islet function

    Institute of Scientific and Technical Information of China (English)

    CHEN Xi; SU Chang; ZHANG Zheng-yun; ZHANG Ming-jun; GU Wei-qiong; LI Xiao-ying; LI Hong-wei; ZHOU Guang-wen

    2011-01-01

    Background The induced expression of heme oxygenase-1 (HO-1) in donor islets improves allograft survival.Cobalt protoporphyrin (CoPP) could significantly enhance the expression of HO-1 mRNA and protein in rat islet safely.Our work was to study how to protect pancreatic islet xenograft by CoPP-induction.Methods Islet xenografts treated with CoPP-induction and CoPP+ Zinc protoporphyrin (ZnPP) in vitro and in vivo were randomly transplanted into murine subrenal capsule; then the graft survival time was compared by blood glucose level and pathological examination and meanwhile the interferon γ (IFN-γ),tumor necrosis factor a (TNF-α),interleukin 10 (IL-10) and IL-1β level in serum and their mRNA and HO-1 mRNA and protein expression were examined.Results Islets with CoPP-induction under low- and high-glucose stimulation exhibited much higher insulin secretion compared with other three groups.CoPP-induction could increase higher expression of HO-1 (mRNA:3.33- and 76.09-fold in vitro and in vivo; protein:2.85- and 58.72-fold).The normoglycemia time in induction groups ((14.63±1.19) and (16.88+1.64) days) was significantly longer.The pathological examination showed less lymphocyte infiltration in induction groups.The IL-10 level and its mRNA in induction groups were significantly higher.Conclusions The HO-1 induced by CoPP would significantly improve function,prolong normoglycemia time and reduce lymphocyte infiltration.Meanwhile CoPP-induction in vivo had more beneficial effects than in vitro.Its mechanism could be related to immune-modulation of IL-10.

  16. Chronic allograft nephropathy.

    Science.gov (United States)

    Vadivel, Nidyanandh; Tullius, Stefan G; Chandraker, Anil

    2007-07-01

    Chronic allograft nephropathy (CAN) remains the Achilles heel of renal transplantation. In spite of the significant strides achieved in one-year renal allograft survival with newer immunosuppressant strategies, the fate of long-term renal allograft survival remains unchanged. The number of renal transplant recipients returning to dialysis has doubled in the past decade. This is especially important since these patients pose a significantly increased likelihood of dying while on the waiting list for retransplantation, due to increasing disparity between donor organ availability versus demand and longer waiting time secondary to heightened immunologic sensitization from their prior transplants. In this review we analyze the latest literature in detail and discuss the definition, natural history, pathophysiology, alloantigen dependent and independent factors that play a crucial role in CAN and the potential newer therapeutic targets on the horizon. This article highlights the importance of early identification and careful management of all the potential contributing factors with particular emphasis on prevention rather than cure of CAN as the core management strategy.

  17. Islet transplantation in rodents: do encapsulated islets really work?

    Directory of Open Access Journals (Sweden)

    Yngrid Ellyn Dias Maciel de Souza

    2011-06-01

    Full Text Available CONTEXT: Diabetes mellitus type I affects around 240 million people in the world and only in the USA 7.8% of the population. It has been estimated that the costs of its complications account for 5% to 10% of the total healthcare spending around the world. According to World Health Organization, 300 million people are expected to develop diabetes mellitus by the year 2025. The pancreatic islet transplantation is expected to be less invasive than a pancreas transplant, which is currently the most commonly used approach. OBJECTIVES: To compare the encapsulated and free islet transplantation in rodents looking at sites of islet implantation, number of injected islets, viability and immunosuppression. METHODS: A literature search was conducted using MEDLINE/PUBMED and SCIELO with terms about islet transplantation in the rodent from 2000 to 2010. We found 2,636 articles but only 56 articles from 2000 to 2010 were selected. RESULTS: In these 56 articles used, 34% were encapsulated and 66% were nonencapsulated islets. Analyzing both types of islets transplantation, the majority of the encapsulated islets were implanted into the peritoneal cavity and the nonencapsulated islets into the liver, through the portal vein. In addition, the great advantage of the peritoneal cavity as the site of islet transplantation is its blood supply. Both vascular endothelial cells and vascular endothelial growth factor were used to stimulate angiogenesis of the islet grafts, increasing the vascularization rapidly after implantation. It also has been proven that there is influence of the capsules, since the larger the capsule more chances there are of central necrosis. In some articles, the use of immunosuppression demonstrated to increase the life expectancy of the graft. CONCLUSION: While significant progress has been made in the islets transplantation field, many obstacles remain to be overcome. Microencapsulation provides a means to transplant islets without

  18. Increased expression of miR-187 in human islets from individuals with type 2 diabetes is associated with reduced glucose-stimulated insulin secretion.

    Science.gov (United States)

    Locke, J M; da Silva Xavier, G; Dawe, H R; Rutter, G A; Harries, L W

    2014-01-01

    Type 2 diabetes is characterised by progressive beta cell dysfunction, with changes in gene expression playing a crucial role in its development. MicroRNAs (miRNAs) are post-transcriptional regulators of gene expression and therefore alterations in miRNA levels may be involved in the deterioration of beta cell function. Global TaqMan arrays and individual TaqMan assays were used to measure islet miRNA expression in discovery (n = 20) and replication (n = 20) cohorts from individuals with and without type 2 diabetes. The role of specific dysregulated miRNAs in regulating insulin secretion, content and apoptosis was subsequently investigated in primary rat islets and INS-1 cells. Identification of miRNA targets was assessed using luciferase assays and by measuring mRNA levels. In the discovery and replication cohorts miR-187 expression was found to be significantly increased in islets from individuals with type 2 diabetes compared with matched controls. An inverse correlation between miR-187 levels and glucose-stimulated insulin secretion (GSIS) was observed in islets from normoglycaemic donors. This correlation paralleled findings in primary rat islets and INS-1 cells where overexpression of miR-187 markedly decreased GSIS without affecting insulin content or apoptotic index. Finally, the gene encoding homeodomain-interacting protein kinase-3 (HIPK3), a known regulator of insulin secretion, was identified as a direct target of miR-187 and displayed reduced expression in islets from individuals with type 2 diabetes. Our findings suggest a role for miR-187 in the blunting of insulin secretion, potentially involving regulation of HIPK3, which occurs during the pathogenesis of type 2 diabetes.

  19. Utilization of organs from donors after circulatory death for vascularized pancreas and islet of Langerhans transplantation : recommendations from an expert group

    NARCIS (Netherlands)

    Berney, Thierry; Boffa, Catherine; Augustine, Titus; Badet, Lionel; de Koning, Eelco; Pratschke, Johann; Socci, Carlo; Friend, Peter

    2015-01-01

    Donation after circulatory death (DCD) donors are increasingly being used as a source of pancreas allografts for vascularized organ and islet transplantation. We provide practice guidelines aiming to increase DCD pancreas utilization. We review risk assessment and donor selection criteria. We report

  20. De novo expression of fetal ED-A(+) fibronectin and B (+) tenascin-C splicing variants in human cardiac allografts: potential impact for targeted therapy of rejection.

    Science.gov (United States)

    Franz, Marcus; Matusiak-Brückner, Monika; Richter, Petra; Grün, Katja; Ziffels, Barbara; Neri, Dario; Maschek, Hansjörg; Schulz, Uwe; Pfeil, Alexander; Jung, Christian; Figulla, Hans R; Gummert, Jan; Berndt, Alexander; Renner, André

    2014-10-01

    Management of acute and especially chronic rejection after human cardiac transplantation is still challenging. Chronic rejection, represented by allograft vasculopathy (CAV) and cardiac interstitial fibrosis (CIF) is known to cause severe long-term complications. Rejection associated tissue-remodelling entails the reoccurrence of fetal variants of Fibronectin (Fn) and Tenascin-C (Tn-C), which are virtually absent in adult human organs. In a rat model, an extensive re-expression could be demonstrated for ED-A(+) Fn with spatial association to CAV and CIF. Thus, it is of great interest to investigate the cardiac tissue expression and distribution in human samples. From 48 heart transplanted patients, 64 tissue specimens derived from right ventricular biopsies were available. Histopathological analysis was performed according to the International Society for Heart and Lung Transplantation (ISHLT) guidelines for the detection of acute rejection. By immunohistochemistry, protein expression of ED-A(+) Fn, B(+) Tn-C, alpha-smooth muscle actin, CD31 and CD45 was assessed and analysed semiquantitatively. Co-localisation studies were performed by means of immunofluorescence double labelling. Histopathological analysis of the 64 samples revealed different ISHLT grades (0R in 36 cases, 1R in 20 cases and 2R in 8 cases). There was a distinct and quantitatively relevant re-occurrence of ED-A(+) Fn and B(+) Tn-C in most samples. Semi-quantitative evaluation did not show any correlation to the acute rejection grade for all markers. Interestingly, significant correlations to the extent of inflammation could be shown for ED-A(+) Fn (r = 0.442, p = 0.000) and B(+) Tn-C (r = 0.408, p = 0.001) as well as between both proteins (r = 0.663, p = 0.000). A spatial association of ED-A(+) Fn and B(+) Tn-C to CAV and CIF could be demonstrated. A relevant re-occurrence of ED-A(+) Fn and B(+) Tn-C following human heart transplantation could be demonstrated with spatial association to

  1. Long-term function of islets encapsulated in a re-designed alginate microcapsule construct in omentum pouches of immune-competent diabetic rats

    Science.gov (United States)

    Pareta, Rajesh; McQuilling, John P; Sittadjody, Sivanandane; Jenkins, Randy; Bowden, Stephen; Orlando, Giuseppe; Farney, Alan C; Brey, Eric M; Opara, Emmanuel C

    2014-01-01

    Objectives Our study aim was to determine encapsulated islet graft viability in an omentum pouch and the effect of FGF-1 released from our redesigned alginate microcapsules on the function of the graft. Methods Isolated rat islets were encapsulated in an inner core made with 1.5% low-viscosity high-mannuronic acid (LVM) alginate followed by an external layer made with 1.25% low-viscosity high-guluronic acid (LVG) alginate with or without FGF-1, in microcapsules measuring 300 – 400 μm in diameter. The two alginate layers were separated by a perm-selective membrane made with 0.1 % Poly-L-Ornithine (PLO), and the inner LVM core was partially chelated using 55 mM sodium citrate for 2 min. Results A marginal mass of encapsulated islet allografts (~2000 islets/kg) in Streptozotocin-diabetic Lewis rats caused significant reduction in blood glucose levels similar to the effect observed with encapsulated islet isografts. Transplantation of allo-islets co-encapsulated with FGF-1 did not result in better glycemic control, but induced greater body weight maintenance in transplant recipients compared to those that received only allo-islets. Histological examination of the retrieved tissue demonstrated morphologically and functionally intact islets in the microcapsules, with no signs of fibrosis. Conclusion We conclude that the omentum is a viable site for encapsulated islet transplantation. PMID:24681880

  2. Human allogeneic CD2+lymphocytes activate airway-derived epithelial cells to produce interleukin-6 and interleukin-8. Possible role for the epithelium in chronic allograft rejection

    NARCIS (Netherlands)

    Borger, P; Kauffman, HF; Scholma, J; Timmerman, JAB; Koeter, GH

    2002-01-01

    Background: The adhesion of lymphocytes to the epithelium and the release of proinflammatory cytokines are important features observed during acute and chronic allograft rejection. Development of chronic rejection in lung-transplantation patients is preceded by high levels of interleukin (IL)-6 and

  3. Lymphoid-Like Structures with Distinct B Cell Areas in Kidney Allografts are not Predictive for Graft Rejection. A Non-human Primate Study

    NARCIS (Netherlands)

    Jonker, Margreet; Wubben, Jacqueline A. M.; 't Hart, Bert A.; Haanstra, Krista G.

    2015-01-01

    Kidney allograft biopsies were analyzed for the presence of B cell clusters/aggregates using CD20 staining. Few B cells were found in the diffuse interstitial infiltrates, but clusters of B cells were found in nodular infiltrates. These nodular infiltrates were smaller shortly after transplantation,

  4. Impact of human leukocyte antigen matching and recipients' panel reactive antibodies on two-year outcome in presensitized renal allograft recipients

    Institute of Scientific and Technical Information of China (English)

    MENG Hui-lin; JIN Xun-bo; LI Xiang-tie; WANG Hong-wei; L(U) Jia-ju

    2009-01-01

    Background Renal transplantation in sensitized candidates remains a highly significant challenge worldwide. The production of panel reactive antibody (PRA) against human leukocyte antigen (HLA) is a major risk factor in presensitized recipients. The aim of this study was to evaluate the impact of HLA matching and recipients' PRA on two-year outcome in presensitized renal allograft recipients.Methods We determined the percentage of panel reactivity and specificity of anti-HLA immunoglobulin (Ig) G antibodies in 73 presensitized renal allograft recipients compared with 81 unsensitized recipients (control group). HLA genotyping of both recipients and corresponding donors was performed by PCR with sequence-specific primers (PCR-SSP). We analyzed the factors influencing the early graft outcome (two-year rejection rates and survival rates of the grafts), including HLA mismatching, class and degree of panel reactivity, and target antigen of donors.Results Presensitized recipients had a worse two-year outcome than unsensitized recipients (P=0.019 for rejection rate, P=0.01 for survival rate). The difference in number of HLA-mismatched alleles with either 6-antigen matching (Ag M) standard or amino acid residue matching (Res M) standard was not significant between the rejection and non-rejection groups of presensitized recipients or between the graft survival group and graft loss group. Compared with the control group, recipients with both PRA-Ⅰ and PRA-Ⅱ antibodies had a significantly worse two-year outcome (P=0.001 for rejection rate, P=0.002 for survival rate). The two-year outcomes of the peak PRA ≥50% group and its subgroup, at-transplant PRA ≥50% group, were significantly worse compared with the control group (P=0.025 and P=0.001 for rejection rate, P=0.043 and P=0.024 for survival rate). The rejection rates of the at-transplant target antigen positive group and its subgroup, HLA-Ⅰ target antigen positive group, were significantly higher than the control

  5. Tissue typing in human limb allograft%同种异体肢体移植配型的初步探讨

    Institute of Scientific and Technical Information of China (English)

    朱立军; 裴国献; 顾立强; 于立新

    2001-01-01

    目的研究组织配型在同种异体肢体移植中的意义和潜力。方法对准备接受同种异体手移植的10例单手或双手缺失准备接受异体手移植的患者和潜在的供者进行ABO血型和Rh血型、群体反应抗体(PRA)检测及混合淋巴细胞培养试验,采用聚合酶链反应(PCR)方法测定人类白细胞抗原(HLA)。结果为其中2例患者各找到1名ABO血型(A-A、O-O)、Rh血型相符,PRA和混合淋巴细胞培养试验均为阴性,人类白细胞抗原(HLA)配型半相合的供者。手术后联合应用免疫抑制剂,移植手顺利成活,免疫监测和皮肤活检证实无排斥反应发生。结论理想的组织配型是避免异体手移植发生排斥反应的前提条件;异体手移植理想的组织配型条件是ABO血型、Rh血型相符,淋巴细胞毒性交叉配合试验为阴性;PRA水平越低、HLA位点配合的越多,越能提高异体手移植的成功率和存活率。%Objectives To find donors with ideal tissue compatibility and study the potential of tissue typing in human hand allotransplantation. Methods ABO, Rh and PRA were studied in ten transplant candidates with one or two-hand defect and potential donors. HLA was confirmed by PRC in these people. Mixed leukocyte culture(MLC) was performed to determine T-cell alloreactivity between donor and recipient. After surgery, immunosuppressants such as antithymocyte globins, FK506, mycophenolic acid and prednisone were used. Results Two patients were matched respectively to two ABO- and Rh-compatible,HLA- half compatible brain-dead donors, with negative PRA and MLC. The postoperative course was uneventful. Ten months after surgery, the two hands recovered and test of C-reactive protein, immunoglobins, CD25 and HLA-DR in the blood and skin biopsy confirmed no occurre no of rejection occurre. Conclusions Ideal tissue compatibility is a prerequisite for hand allografting to prevent graft

  6. Macrophage-to-Myofibroblast Transition Contributes to Interstitial Fibrosis in Chronic Renal Allograft Injury.

    Science.gov (United States)

    Wang, Ying-Ying; Jiang, Hong; Pan, Jun; Huang, Xiao-Ru; Wang, Yu-Cheng; Huang, Hong-Feng; To, Ka-Fai; Nikolic-Paterson, David J; Lan, Hui-Yao; Chen, Jiang-Hua

    2017-02-16

    Interstitial fibrosis is an important contributor to graft loss in chronic renal allograft injury. Inflammatory macrophages are associated with fibrosis in renal allografts, but how these cells contribute to this damaging response is not clearly understood. Here, we investigated the role of macrophage-to-myofibroblast transition in interstitial fibrosis in human and experimental chronic renal allograft injury. In biopsy specimens from patients with active chronic allograft rejection, we identified cells undergoing macrophage-to-myofibroblast transition by the coexpression of macrophage (CD68) and myofibroblast (α-smooth muscle actin [α-SMA]) markers. CD68(+)/α-SMA(+) cells accounted for approximately 50% of the myofibroblast population, and the number of these cells correlated with allograft function and the severity of interstitial fibrosis. Similarly, in C57BL/6J mice with a BALB/c renal allograft, cells coexpressing macrophage markers (CD68 or F4/80) and α-SMA composed a significant population in the interstitium of allografts undergoing chronic rejection. Fate-mapping in Lyz2-Cre/Rosa26-Tomato mice showed that approximately half of α-SMA(+) myofibroblasts in renal allografts originated from recipient bone marrow-derived macrophages. Knockout of Smad3 protected against interstitial fibrosis in renal allografts and substantially reduced the number of macrophage-to-myofibroblast transition cells. Furthermore, the majority of macrophage-to-myofibroblast transition cells in human and experimental renal allograft rejection coexpressed the M2-type macrophage marker CD206, and this expression was considerably reduced in Smad3-knockout recipients. In conclusion, our studies indicate that macrophage-to-myofibroblast transition contributes to interstitial fibrosis in chronic renal allograft injury. Moreover, the transition of bone marrow-derived M2-type macrophages to myofibroblasts in the renal allograft is regulated via a Smad3-dependent mechanism.

  7. The Pattern of Neural Elements in the Islets of Normal and Diseased Pancreas and in Isolated Islets

    Directory of Open Access Journals (Sweden)

    Parviz M Pour

    2011-07-01

    Full Text Available Context The association between islet cells and neural elements, the so-called “neuro-insular complex”, has been known for centuries. Objective We examined the expression of beta-III tubulin, in normal pancreases from organ donors, surgical specimens of chronic pancreatitis, surgical specimens of ductal type carcinoma, isolated and purified islets of a 57-year-old male and the pancreases of adult Syrian golden hamsters by immunohistochemistry using a monoclonal antibody to beta-tubulin. Results In the normal pancreas of humans and hamsters, beta-III tubulin was expressed in alpha- and beta-cells, but not in PP cells, neural fibers and gangliae. Occasionally, intra-and peri-insular neural elements were also found. In chronic pancreatitis and pancreatic cancer samples, the number of beta-cells and the immunoreactivity of the beta-III tubulin antibody in islet cells were decreased in most cases. In cultured human islets, devoid of neural elements, no correlation was found between the expression of beta-III tubulin and islet cell hormones. Conclusion Beta-III tubulin is only expressed in the islets derived from the dorsal pancreas and in neural elements. In chronic pancreatitis and pancreatic cancer swelling of intra- and peri-insular nerves occurs, possibly in response to the loss of beta-cells. The secretion of insulin and the expression of beta-tubulin seem to be regulated by nerves.

  8. Ultrastructural studies of time-course and cellular specificity of interleukin-1 mediated islet cytotoxicity

    DEFF Research Database (Denmark)

    Mandrup-Poulsen, T; Egeberg, J; Nerup, J

    1987-01-01

    of incubation and examined by electron microscopy in a blinded fashion. Already after 30 min, accumulation of opaque intracytoplasmic bodies without apparent surrounding membranes, and autophagic vacuoles were seen in about 20% of the beta cells examined in rat islets exposed to interleukin-1. After 16 h......Previous electron-microscopic studies of isolated islets of Langerhans exposed to the monokine interleukin-1 for 7 days have indicated that interleukin-1 is cytotoxic to all islet cells. To study the time-course and possible cellular specificity of interleukin-1 cytotoxicity to islets exposed...... of incubation with interleukin-1, more than 80% of rat beta cells showed signs of degeneration. Beta cell specific changes similar to those observed in rat islets exposed to IL-1 for 30 min were seen in human islets exposed to IL-1 for 24 h. The described changes were not observed in alpha cells in interleukin...

  9. Islet expression of the DNA repair enzyme 8-oxoguanosine DNA glycosylase (Ogg1) in human type 2 diabetes

    OpenAIRE

    Yoon Kun-Ho; Wang-Rodriguez Jessica; Dib Sergio A.; Anachkov Kamen A; Tyrberg Björn; Levine Fred

    2002-01-01

    Abstract Background It has become increasingly clear that β-cell failure plays a critical role in the pathogenesis of type 2 diabetes. Free-radical mediated β-cell damage has been intensively studied in type 1 diabetes, but not in human type 2 diabetes. Therefore, we studied the protein expression of the DNA repair enzyme Ogg1 in pancreases from type 2 diabetics. Ogg1 was studied because it is the major enzyme involved in repairing 7,8-dihydro-8-oxoguanosine DNA adducts, a lesion previously o...

  10. 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

    contents of insulin and glucagon in a dose-dependent manner. A maximal effect on islet function was obtained with supernatant concentrations down to 5%. Supernatants of mononuclear cells stimulated with tuberculin were more potent than supernatants produced by lectin stimulation. Culture medium......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...

  11. Effect of interleukin-1 on the biosynthesis of proinsulin and insulin in isolated rat pancreatic islets

    DEFF Research Database (Denmark)

    Hansen, Birgit Sehested; Linde, S; Spinas, G A

    1988-01-01

    Insulin dependent diabetes mellitus (IDDM) is often preceded or associated with lymphocytic infiltration in the islets of Langerhans (insulitis). We recently demonstrated that interleukin-1 (IL-1) produced by activated macrophages exerts a bimodal effect on insulin release and biosynthesis...... in isolated rat islets. In the present study we have further analysed the effect of recombinant human interleukin-1 beta (rIL-1) on the biosynthesis and conversion of proinsulin 1 and 2 in rat islets. By RP-HPLC-analysis of islets labelled with [3H]leucine we found that exposure to 6 ng/ml of IL-1 for 24 h...

  12. Adaptation of pancreatic islet cyto-architecture during development

    Science.gov (United States)

    Striegel, Deborah A.; Hara, Manami; Periwal, Vipul

    2016-04-01

    Plasma glucose in mammals is regulated by hormones secreted by the islets of Langerhans embedded in the exocrine pancreas. Islets consist of endocrine cells, primarily α, β, and δ cells, which secrete glucagon, insulin, and somatostatin, respectively. β cells form irregular locally connected clusters within islets that act in concert to secrete insulin upon glucose stimulation. Varying demands and available nutrients during development produce changes in the local connectivity of β cells in an islet. We showed in earlier work that graph theory provides a framework for the quantification of the seemingly stochastic cyto-architecture of β cells in an islet. To quantify the dynamics of endocrine connectivity during development requires a framework for characterizing changes in the probability distribution on the space of possible graphs, essentially a Fokker-Planck formalism on graphs. With large-scale imaging data for hundreds of thousands of islets containing millions of cells from human specimens, we show that this dynamics can be determined quantitatively. Requiring that rearrangement and cell addition processes match the observed dynamic developmental changes in quantitative topological graph characteristics strongly constrained possible processes. Our results suggest that there is a transient shift in preferred connectivity for β cells between 1-35 weeks and 12-24 months.

  13. Islet cell development.

    Science.gov (United States)

    Rojas, Anabel; Khoo, Adrian; Tejedo, Juan R; Bedoya, Francisco J; Soria, Bernat; Martín, Franz

    2010-01-01

    Over the last years, there has been great success in driving stem cells toward insulin-expressing cells. However, the protocols developed to date have some limitations, such as low reliability and low insulin production. The most successful protocols used for generation of insulin-producing cells from stem cells mimic in vitro pancreatic organogenesis by directing the stem cells through stages that resemble several pancreatic developmental stages. Islet cell fate is coordinated by a complex network of inductive signals and regulatory transcription factors that, in a combinatorial way, determine pancreatic organ specification, differentiation, growth, and lineage. Together, these signals and factors direct the progression from multipotent progenitor cells to mature pancreatic cells. Later in development and adult life, several of these factors also contribute to maintain the differentiated phenotype of islet cells. A detailed understanding of the processes that operate in the pancreas during embryogenesis will help us to develop a suitable source of cells for diabetes therapy. In this chapter, we will discuss the main transcription factors involved in pancreas specification and beta-cell formation.

  14. Protection of rat islet viability following heme oxygenase-1 gene transfection via adenoviral vector in vitro

    Institute of Scientific and Technical Information of China (English)

    Xiaobo Chen; Yongxiang Li; Weiping Dong; Yang Jiao; Jianming Tan

    2007-01-01

    Objective: To investigate the effect of Heme oxygenase-1 (HO-1) gene transfection on the viability of cultured rat islets, and to explore the potential value of HO-1 gene in islet transplantation. Methods:Recombinant adenovirus vector containing human HO-1 gene(Ad-HO-1 ) or enhanced green fluorescent protein gene(Ad-EGFP) was generated by using AdEasy system respectively.The rat islets were transfected with Ad-HO-1, Ad-EGFP or blank vector and then cultured for 7 days. Transfection was confirmed by expression of EGFP and human HO-1 protein detected by fluorescence photographs and western blot, respectively. The insulin release upon different concentration of glucose stimulation was detected using insulin radioimmunoassay kit, and stimulation index (SI) was calculated. Glucose-stimulated insulin release was usedto assess islet viability. Results:Adenovirus vector successfully transferred HO-1 gene to rat islet cells in vitro, and the insulin release upon high level of glucose stimulation and stimulation index(SI) of Ad-HO-1-infected islets were significantly higher than those of Ad-EGFP-infected islets and control islets(P < 0.05).Conclusion: Adenovirus-mediated HO-1 gene transfection is a feasible strategy to confer cytoprotection and therefore protect the viability of cultured rat islets.

  15. Macro-or microencapsulation of pig islets to cure type 1 diabetes

    Institute of Scientific and Technical Information of China (English)

    Denis Dufrane; Pierre Gianello

    2012-01-01

    Although allogeneic islet transplantation can successfully cure type 1 diabetes,it has limited applicability.For example,organs are in short supply; several human pancreas donors are often needed to treat one diabetic recipient; the intrahepatic site may not be the most appropriate site for islet implantation; and immunosuppressive regimens,which are associated with side effects,are often required to prolong survival of the islet graft.An altemative source of insulinproducing cells would therefore be of major interest.Pigs represent a possible alternative source of beta cells.Grafting of pig islets may appear difficult because of the immunologic species barrier,but pig islets have been shown to function in primates for at least 6 mo with clinically incompatible immunosuppression.Therefore,a bioartificial pancreas made of encapsulated pig islets may resolve issues associated with islet allotransplantation.Although several groups have shown that encapsulated pig islets are functional in small-animal models,less is known about the use of bioartificial pancreases in large-animal models.In this review,we summarize current knowledge of encapsulated pig islets,to determine obstacles to implantation in humans and possible solutions to overcome these obstacles.

  16. Chronic Antibody-Mediated Rejection in Nonhuman Primate Renal Allografts: Validation of Human Histological and Molecular Phenotypes.

    Science.gov (United States)

    Adam, B A; Smith, R N; Rosales, I A; Matsunami, M; Afzali, B; Oura, T; Cosimi, A B; Kawai, T; Colvin, R B; Mengel, M

    2017-04-26

    Molecular testing represents a promising adjunct for the diagnosis of antibody-mediated rejection (AMR). Here, we apply a novel gene expression platform in sequential formalin-fixed paraffin-embedded samples from nonhuman primate (NHP) renal transplants. We analyzed 34 previously described gene transcripts related to AMR in humans in 197 archival NHP samples, including 102 from recipients that developed chronic AMR, 80 from recipients without AMR, and 15 normal native nephrectomies. Three endothelial genes (VWF, DARC, and CAV1), derived from 10-fold cross-validation receiver operating characteristic curve analysis, demonstrated excellent discrimination between AMR and non-AMR samples (area under the curve = 0.92). This three-gene set correlated with classic features of AMR, including glomerulitis, capillaritis, glomerulopathy, C4d deposition, and DSAs (r = 0.39-0.63, p < 0.001). Principal component analysis confirmed the association between three-gene set expression and AMR and highlighted the ambiguity of v lesions and ptc lesions between AMR and T cell-mediated rejection (TCMR). Elevated three-gene set expression corresponded with the development of immunopathological evidence of rejection and often preceded it. Many recipients demonstrated mixed AMR and TCMR, suggesting that this represents the natural pattern of rejection. These data provide NHP animal model validation of recent updates to the Banff classification including the assessment of molecular markers for diagnosing AMR. © 2017 The American Society of Transplantation and the American Society of Transplant Surgeons.

  17. New Insight in Copper-Ion Binding to Human Islet Amyloid: The Contribution of Metal-Complex Speciation To Reveal the Polypeptide Toxicity.

    Science.gov (United States)

    Magrì, Antonio; La Mendola, Diego; Nicoletti, Vincenzo Giuseppe; Pappalardo, Giuseppe; Rizzarelli, Enrico

    2016-09-05

    Type-2 diabetes (T2D) is considered to be a potential threat on a global level. Recently, T2D has been listed as a misfolding disease, such as Alzheimer's and Parkinson's diseases. Human islet amyloid polypeptide (hIAPP) is a molecule cosecreted in pancreatic β cells and represents the main constituent of an aggregated amyloid found in individuals affected by T2D. The trace-element serum level is significantly influenced during the development of diabetes. In particular, the dys-homeostasis of Cu(2+) ions may adversely affect the course of the disease. Conflicting results have been reported on the protective role played by complex species formed by Cu(2+) ions with hIAPP or its peptide fragments in vitro. The histidine (His) residue at position 18 represents the main binding site for the metal ion, but contrasting results have been reported on other residues involved in metal-ion coordination, in particular those toward the N or C terminus. Sequences that encompass regions 17-29 and 14-22 were used to discriminate between the two models of the hIAPP coordination mode. Due to poor solubility in water, poly(ethylene glycol) (PEG) derivatives were synthesized. A peptide fragment that encompasses the 17-29 region of rat amylin (rIAPP) in which the arginine residue at position 18 was substituted by a histidine residue was also obtained to assess that the PEG moiety does not alter the peptide secondary structure. The complex species formed by Cu(2+) ions with Ac-PEG-hIAPP(17-29)-NH2 , Ac-rIAPP(17-29)R18H-NH2 , and Ac-PEG-hIAPP(14-22)-NH2 were studied by using potentiometric titrations coupled with spectroscopic methods (UV/Vis, circular dichroism, and EPR). The combined thermodynamic and spectroscopic approach allowed us to demonstrate that hIAPP is able to bind Cu(2+) ions starting from the His18 imidazole nitrogen atom toward the N-terminus domain. The stability constants of copper(II) complexes with Ac-PEG-hIAPP(14-22)-NH2 were used to simulate the different

  18. Hypertension in Renal Allograft Recipients

    Directory of Open Access Journals (Sweden)

    Waiser Johannes

    1999-01-01

    Full Text Available Hypertension is a frequent complication after renal transplantation. It contributes to the considerable cardiovascular morbidity and mortality in renal allograft recipients. Additionally, it has a major impact on long-term allograft survival. The pathogenesis of post transplant hypertension is multifactorial. Besides common risk factors, renal allograft recipients accumulate specific risk factors related to the original renal disease, renal transplantation per se and the immunosuppressive regimen. Chronic allograft dysfunction is the main cause of post transplant hypertension. The introduction of calcineurin inhibitors, such as cyclosporine, has increased the prevalence of hypertension. At present, the growing manual of diagnostic and therapeutic tools enables us to adapt better antihypertensive therapy. Tight monitoring, individualization of the immunosuppressive protocol, inclusion of non-pharmacological measures and aggressive antihypertensive treatment should help to minimize the negative implications of post transplant hypertension. Probably, this goal can only be reached by "normalization" of systolic and diastolic blood pressure to below 135/85 mmHg.

  19. Radionuclide diagnosis of allograft rejection

    Energy Technology Data Exchange (ETDEWEB)

    George, E.A.

    1982-10-01

    Interaction with one or more anatomical and physiopathological characteristics of the rejecting renal allograft is suggested by those radioagents utilized specifically for the diagnosis of allograft rejection. Rejection, the most common cause of declining allograft function, is frequently mimicked clinically or masked by other immediate or long term post transplant complications. Understanding of the anatomical pathological features and kinetics of rejection and their modification by immunosuppressive maintenance and therapy are important for the proper clinical utilization of these radioagents. Furthermore, in selecting these radionuclides, one has to consider the comparative availability, preparatory and procedural simplicity, acquisition and display techniques and the possibility of timely report. The clinical utilities of radiofibrinogen, /sup 99m/Tc sulfur colloid and /sup 67/Ga in the diagnosis of allograft rejection have been evaluated to a variable extent in the past. The potential usefulness of the recently developed preparations of /sup 111/In labeled autologous leukocytes and platelets are presently under investigation.

  20. Islet Xenotransplantation and Xeno-antigenicity: studies in a preclinical model

    NARCIS (Netherlands)

    P.P.M. Rood (Pleunie)

    2008-01-01

    textabstractShortage of human donor organs is the major limiting factor for clinical islet allotransplantation. Xenotransplantation, using the pig as the source of islets is considered a potential solution to this problem. Since the development of pigs homozygous for α1,3-galactosyltransfer

  1. The role of the disulfide bond in the interaction of islet amyloid polypeptide with membranes

    NARCIS (Netherlands)

    Khemtemourian, L.P.; Engel, M.F.M.; Kruijtzer, J.A.W.; Hoppener, J.W.M.; Liskamp, R.M.J.; Killian, J.A.

    2010-01-01

    Human islet amyloid polypeptide (hIAPP) forms amyloid fibrils in pancreatic islets of patients with type 2 diabetes mellitus. It has been suggested that the N-terminal part, which contains a conserved intramolecular disulfide bond between residues 2 and 7, interacts with membranes, ultimately

  2. An imidazoline compound completely counteracts interleukin-1[beta] toxic effects to rat pancreatic islet [beta] cells

    DEFF Research Database (Denmark)

    Papaccio, Gianpaolo; Nicoletti, Ferdinando; Pisanti, Francesco A

    2002-01-01

    In vitro studies have demonstrated that interleukin (IL)-1beta decreases insulin and DNA contents in pancreatic islet beta cells, causing structural damage, that it is toxic to cultured human islet beta cells and that it is able to induce apoptosis in these cells....

  3. Pancreatic islets engineered with SA-FasL protein establish robust localized tolerance by inducing regulatory T cells in mice.

    Science.gov (United States)

    Yolcu, Esma S; Zhao, Hong; Bandura-Morgan, Laura; Lacelle, Chantale; Woodward, Kyle B; Askenasy, Nadir; Shirwan, Haval

    2011-12-01

    Allogeneic islet transplantation is an important therapeutic approach for the treatment of type 1 diabetes. Clinical application of this approach, however, is severely curtailed by allograft rejection primarily initiated by pathogenic effector T cells regardless of chronic use of immunosuppression. Given the role of Fas-mediated signaling in regulating effector T cell responses, we tested if pancreatic islets can be engineered ex vivo to display on their surface an apoptotic form of Fas ligand protein chimeric with streptavidin (SA-FasL) and whether such engineered islets induce tolerance in allogeneic hosts. Islets were modified with biotin following efficient engineering with SA-FasL protein that persisted on the surface of islets for >1 wk in vitro. SA-FasL-engineered islet grafts established euglycemia in chemically diabetic syngeneic mice indefinitely, demonstrating functionality and lack of acute toxicity. Most importantly, the transplantation of SA-FasL-engineered BALB/c islet grafts in conjunction with a short course of rapamycin treatment resulted in robust localized tolerance in 100% of C57BL/6 recipients. Tolerance was initiated and maintained by CD4(+)CD25(+)Foxp3(+) regulatory T (Treg) cells, as their depletion early during tolerance induction or late after established tolerance resulted in prompt graft rejection. Furthermore, Treg cells sorted from graft-draining lymph nodes, but not spleen, of long-term graft recipients prevented the rejection of unmodified allogeneic islets in an adoptive transfer model, further confirming the Treg role in established tolerance. Engineering islets ex vivo in a rapid and efficient manner to display on their surface immunomodulatory proteins represents a novel, safe, and clinically applicable approach with important implications for the treatment of type 1 diabetes.

  4. Pancreatic Islets Engineered with SA-FasL Protein Establish Robust Localized Tolerance by Inducing T Regulatory Cells in Mice

    Science.gov (United States)

    Yolcu, Esma S; Zhao, Hong; Bandura-Morgan, Laura; Lacelle, Chantale; Woodward, Kyle B; Askenasy, Nadir; Shirwan, Haval

    2011-01-01

    Allogeneic islet transplantation is an important therapeutic approach for the treatment of T1D. Clinical application of this approach, however, is severely curtailed by allograft rejection primarily initiated by pathogenic T effector cells regardless of chronic use of immunosuppression. Given the role of Fas-mediated signaling in regulating T effector cell responses, we tested if pancreatic islets can be engineered ex vivo to display on their surface an apoptotic form of FasL protein chimeric with streptavidin (SA-FasL), and whether such engineered islets induce tolerance in allogeneic hosts. Islets were modified with biotin following efficient engineering with SA-FasL protein that persisted on the surface of islets for over a week in vitro. SA-FasL-engineered islet grafts established euglycemia in chemically diabetic syngeneic mice indefinitely, demonstrating functionality and lack of acute toxicity. Most importantly, the transplantation of SA-FasL-engineered BALB/c islet grafts in conjunction with a short course of rapamycin treatment resulted in robust localized tolerance in 100% C57BL/6 recipients. Tolerance was initiated and maintained by CD4+CD25+FoxP3+ T regulatory (Treg) cells as their depletion early during tolerance induction or late after established tolerance resulted in prompt graft rejection. Furthermore, Treg cells sorted from graft-draining lymph nodes, but not spleen, of long-term graft recipients prevented the rejection of unmodified allogeneic islets in an adoptive transfer model, further confirming the Treg role in established tolerance. Engineering islets ex vivo in a rapid and efficient manner to display on their surface immunomodulatory proteins represents a novel, safe, and clinically applicable approach with important implications for the treatment of T1D. PMID:22068235

  5. Recent insights in islet amyloid polypeptide-induced membrane disruption and its role in β-cell death in type II diabetes mellitus

    NARCIS (Netherlands)

    Khemtémourian, L.P.|info:eu-repo/dai/nl/304824372; Killian, J.A.|info:eu-repo/dai/nl/071792317; Höppener, J.W.M.; Engel, M.F.M.|info:eu-repo/dai/nl/263614190

    2008-01-01

    The presence of fibrillar protein deposits (amyloid) of human islet amyloid polypeptide (hIAPP) in the pancreatic islets of Langerhans is thought to be related to death of the insulin-producing islet β-cells in type 2 diabetes mellitus (DM2). The mechanism of hIAPP-induced β-cell death is not

  6. Immunosuppression for islet transplantation.

    Directory of Open Access Journals (Sweden)

    Noguchi,Hirofumi

    2006-04-01

    Full Text Available The development by the Edmonton group of a sirolimus-based, steroid-free, low-tacrolimus regimen is a significant breakthrough that allows the rate of insulin independence after islet transplantation to increase from 13% to 80% at 1 year ; however, the rate is reduced to 50% at 3 years, attributed to prolonged tacrolimus exposure. Recently, immunosuppression agents such as cyclosporine, mycophenolate mofetil, and the novel agent FTY 720 have been used instead of tacrolimus. Lymphocytedepleting antibodies such as anti-thymocyte globulin, alemtuzumab, and hOKT3gamma 1 (ala, ala have been launched, and a costimulatory blockade of anti-CD40 monoclonal antibodies and CTLA4-Ig will be attempted in the near future. Moreover, the potential of a novel immunosuppressing peptide could now be realized using new technology called the protein transduction system. In this review, we show some of the most recent contributions to the advancement of knowledge in this field.

  7. 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.

    Directory of Open Access Journals (Sweden)

    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.

  8. Angiopoetin-2 Signals Do Not Mediate the Hypervascularization of Islets in Type 2 Diabetes

    Science.gov (United States)

    Shah, Payal; Lueschen, Navina; Ardestani, Amin; Oberholzer, Jose; Olerud, Johan; Carlsson, Per-Ola; Maedler, Kathrin

    2016-01-01

    Aims Changes in the islet vasculature have been implicated in the regulation of β-cell survival and function during the progression to type 2 diabetes (T2D). Failure of the β-cell to compensate for the increased insulin demand in obesity eventually leads to diabetes; as a result of the complex interplay of genetic and environmental factors (e.g. ongoing inflammation within the islets) and impaired vascular function. The Angiopoietin/Tie (Ang/Tie) angiogenic system maintains vasculature and is closely related to organ inflammation and angiogenesis. In this study we aimed to identify whether the vessel area within the islets changes in diabetes and whether such changes would be triggered by the Tie-antagonist Ang-2. Methods Immunohistochemical and qPCR analyses to follow islet vascularization and Ang/Tie levels were performed in human pancreatic autopsies and isolated human and mouse islets. The effect of Ang-2 was assessed in β-cell-specific Ang-2 overexpressing mice during high fat diet (HFD) feeding. Results Islet vessel area was increased in autopsy pancreases from patients with T2D. The vessel markers Tie-1, Tie-2 and CD31 were upregulated in mouse islets upon HFD feeding from 8 to 24 weeks. Ang-2 was transiently upregulated in mouse islets at 8 weeks of HFD and under glucolipotoxic conditions (22.2 mM glucose/ 0.5 mM palmitate) in vitro in human and mouse islets, in contrast to its downregulation by cytokines (IL-1β, IFN-ɣ and TNF-α). Ang-1 on the other hand was oppositely regulated, with a significant loss under glucolipotoxic condition, a trend to reduce in islets from patients with T2D and an upregulation by cytokines. Modulation of such changes in Ang-2 by its overexpression or the inhibition of its receptor Tie-2 impaired β-cell function at basal conditions but protected islets from cytokine induced apoptosis. In vivo, β-cell-specific Ang-2 overexpression in mice induced hypervascularization under normal diet but contrastingly led to

  9. Quantitative Assessment of Islets of Langerhans Encapsulated in Alginate

    Science.gov (United States)

    Johnson, Amy S.; O'Sullivan, Esther; D'Aoust, Laura N.; Omer, Abdulkadir; Bonner-Weir, Susan; Fisher, Robert J.; Weir, Gordon C.

    2011-01-01

    Improved methods have recently been developed for assessing islet viability and quantity in human islet preparations for transplantation, and these measurements have proven useful for predicting transplantation outcome. The objectives of this study were to adapt these methods for use with microencapsulated islets, to verify that they provide meaningful quantitative measurements, and to test them with two model systems: (1) barium alginate and (2) barium alginate containing a 70% (w/v) perfluorocarbon (PFC) emulsion, which presents challenges to use of these assays and is of interest in its own right as a means for reducing oxygen supply limitations to encapsulated tissue. Mitochondrial function was assessed by oxygen consumption rate measurements, and the analysis of data was modified to account for the increased solubility of oxygen in the PFC-alginate capsules. Capsules were dissolved and tissue recovered for nuclei counting to measure the number of cells. Capsule volume was determined from alginate or PFC content and used to normalize measurements. After low oxygen culture for 2 days, islets in normal alginate lost substantial viable tissue and displayed necrotic cores, whereas most of the original oxygen consumption rate was recovered with PFC alginate, and little necrosis was observed. All nuclei were recovered with normal alginate, but some nuclei from nonrespiring cells were lost with PFC alginate. Biocompatibility tests revealed toxicity at the islet periphery associated with the lipid emulsion used to provide surfactants during the emulsification process. We conclude that these new assay methods can be applied to islets encapsulated in materials as complex as PFC-alginate. Measurements made with these materials revealed that enhancement of oxygen permeability of the encapsulating material with a concentrated PFC emulsion improves survival of encapsulated islets under hypoxic conditions, but reformulation of the PFC emulsion is needed to reduce toxicity

  10. Unraveling pancreatic islet biology by quantitative proteomics

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Jianying; Dann, Geoffrey P.; Liew, Chong W.; Smith, Richard D.; Kulkarni, Rohit N.; Qian, Weijun

    2011-08-01

    The pancreatic islets of Langerhans play a critical role in maintaining blood glucose homeostasis by secreting insulin and several other important peptide hormones. Impaired insulin secretion due to islet dysfunction is linked to the pathogenesis underlying both Type 1 and Type 2 diabetes. Over the past 5 years, emerging proteomic technologies have been applied to dissect the signaling pathways that regulate islet functions and gain an understanding of the mechanisms of islet dysfunction relevant to diabetes. Herein, we briefly review some of the recent quantitative proteomic studies involving pancreatic islets geared towards gaining a better understanding of islet biology relevant to metabolic diseases.

  11. Lymphoid-Like Structures with Distinct B Cell Areas in Kidney Allografts are not Predictive for Graft Rejection. A Non-human Primate Study.

    Science.gov (United States)

    Jonker, Margreet; Wubben, Jacqueline A M; 't Hart, Bert A; Haanstra, Krista G

    2015-12-01

    Kidney allograft biopsies were analyzed for the presence of B cell clusters/aggregates using CD20 staining. Few B cells were found in the diffuse interstitial infiltrates, but clusters of B cells were found in nodular infiltrates. These nodular infiltrates were smaller shortly after transplantation, and their size increased over time. At the time of clinical rejection, the nodules often presented as tertiary lymphoid structures (TLS) with lymphoid-like follicles. The presence of small B cell clusters during the first 2 months after transplantation was not associated with early rejection. Even in animals that did not reject their allograft, TLS-like structures were present and could disappear over time. Although TLS were more often found in samples with interstitial fibrosis and tubular atrophy (IFTA), TLS were also present in samples without IFTA. The presence and density of clusters resembling tertiary lymphoid structures most likely reflect an ongoing immune response inside the graft and do not necessarily signify a poor graft outcome or IFTA.

  12. Fetal endocannabinoids orchestrate the organization of pancreatic islet microarchitecture.

    Science.gov (United States)

    Malenczyk, Katarzyna; Keimpema, Erik; Piscitelli, Fabiana; Calvigioni, Daniela; Björklund, Peyman; Mackie, Kenneth; Di Marzo, Vincenzo; Hökfelt, Tomas G M; Dobrzyn, Agnieszka; Harkany, Tibor

    2015-11-10

    Endocannabinoids are implicated in the control of glucose utilization and energy homeostasis by orchestrating pancreatic hormone release. Moreover, in some cell niches, endocannabinoids regulate cell proliferation, fate determination, and migration. Nevertheless, endocannabinoid contributions to the development of the endocrine pancreas remain unknown. Here, we show that α cells produce the endocannabinoid 2-arachidonoylglycerol (2-AG) in mouse fetuses and human pancreatic islets, which primes the recruitment of β cells by CB1 cannabinoid receptor (CB1R) engagement. Using subtractive pharmacology, we extend these findings to anandamide, a promiscuous endocannabinoid/endovanilloid ligand, which impacts both the determination of islet size by cell proliferation and α/β cell sorting by differential activation of transient receptor potential cation channel subfamily V member 1 (TRPV1) and CB1Rs. Accordingly, genetic disruption of TRPV1 channels increases islet size whereas CB1R knockout augments cellular heterogeneity and favors insulin over glucagon release. Dietary enrichment in ω-3 fatty acids during pregnancy and lactation in mice, which permanently reduces endocannabinoid levels in the offspring, phenocopies CB1R(-/-) islet microstructure and improves coordinated hormone secretion. Overall, our data mechanistically link endocannabinoids to cell proliferation and sorting during pancreatic islet formation, as well as to life-long programming of hormonal determinants of glucose homeostasis.

  13. Inhibition of nuclear factor-κB activation in pancreatic β-cells has a protective effect on allogeneic pancreatic islet graft survival.

    Directory of Open Access Journals (Sweden)

    Roy Eldor

    Full Text Available Pancreatic islet transplantation, a treatment for type 1 diabetes, has met significant challenges, as a substantial fraction of the islet mass fails to engraft, partly due to death by apoptosis in the peri- and post-transplantation periods. Previous evidence has suggested that NF-κB activation is involved in cytokine-mediated β-cell apoptosis and regulates the expression of pro-inflammatory and chemokine genes. We therefore sought to explore the effects of β-cell-specific inhibition of NF-κB activation as a means of cytoprotection in an allogeneic model of islet transplantation. To this end, we used islets isolated from the ToI-β transgenic mouse, where NF-κB signalling can specifically and conditionally be inhibited in β-cells by expressing an inducible and non-degradable form of IκBα regulated by the tet-on system. Our results show that β-cell-specific blockade of NF-κB led to a prolonged islet graft survival, with a relative higher preservation of the engrafted endocrine tissue and reduced inflammation. Importantly, a longer delay in allograft rejection was achieved when mice were systemically treated with the proteasome inhibitor, Bortezomib. Our findings emphasize the contribution of NF-κB activation in the allograft rejection process, and suggest an involvement of the CXCL10/IP-10 chemokine. Furthermore, we suggest a potential, readily available therapeutic agent that may temper this process.

  14. Inhibition of nuclear factor-κB activation in pancreatic β-cells has a protective effect on allogeneic pancreatic islet graft survival.

    Science.gov (United States)

    Eldor, Roy; Abel, Roy; Sever, Dror; Sadoun, Gad; Peled, Amnon; Sionov, Ronit; Melloul, Danielle

    2013-01-01

    Pancreatic islet transplantation, a treatment for type 1 diabetes, has met significant challenges, as a substantial fraction of the islet mass fails to engraft, partly due to death by apoptosis in the peri- and post-transplantation periods. Previous evidence has suggested that NF-κB activation is involved in cytokine-mediated β-cell apoptosis and regulates the expression of pro-inflammatory and chemokine genes. We therefore sought to explore the effects of β-cell-specific inhibition of NF-κB activation as a means of cytoprotection in an allogeneic model of islet transplantation. To this end, we used islets isolated from the ToI-β transgenic mouse, where NF-κB signalling can specifically and conditionally be inhibited in β-cells by expressing an inducible and non-degradable form of IκBα regulated by the tet-on system. Our results show that β-cell-specific blockade of NF-κB led to a prolonged islet graft survival, with a relative higher preservation of the engrafted endocrine tissue and reduced inflammation. Importantly, a longer delay in allograft rejection was achieved when mice were systemically treated with the proteasome inhibitor, Bortezomib. Our findings emphasize the contribution of NF-κB activation in the allograft rejection process, and suggest an involvement of the CXCL10/IP-10 chemokine. Furthermore, we suggest a potential, readily available therapeutic agent that may temper this process.

  15. Fibroblast populated collagen matrix promotes islet survival and reduces the number of islets required for diabetes reversal.

    Science.gov (United States)

    Jalili, Reza B; Moeen Rezakhanlou, Alireza; Hosseini-Tabatabaei, Azadeh; Ao, Ziliang; Warnock, Garth L; Ghahary, Aziz

    2011-07-01

    Islet transplantation represents a viable treatment for type 1 diabetes. However, due to loss of substantial mass of islets early after transplantation, islets from two or more donors are required to achieve insulin independence. Islet-extracellular matrix disengagement, which occurs during islet isolation process, leads to subsequent islet cell apoptosis and is an important contributing factor to early islet loss. In this study, we developed a fibroblast populated collagen matrix (FPCM) as a novel scaffold to improve islet cell viability and function post-transplantation. FPCM was developed by embedding fibroblasts within type-I collagen and used as scaffold for islet grafts. Viability and insulin secretory function of islets embedded within FPCM was evaluated in vitro and in a syngeneic murine islet transplantation model. Islets embedded within acellular matrix or naked islets were used as control. Islet cell survival and function was markedly improved particularly after embedding within FPCM. The composite scaffold significantly promoted islet isograft survival and reduced the critical islet mass required for diabetes reversal by half (from 200 to 100 islets per recipient). Fibroblast embedded within FPCM produced fibronectin and growth factors and induced islet cell proliferation. No evidence of fibroblast over-growth within composite grafts was noticed. These results confirm that FPCM significantly promotes islet viability and functionality, enhances engraftment of islet grafts and decreases the critical islet mass needed to reverse hyperglycemia. This promising finding offers a new approach to reducing the number of islet donors per recipient and improving islet transplant outcome.

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

    Science.gov (United States)

    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.

  17. Experimental studies on islet isolation and islet graft function

    NARCIS (Netherlands)

    Suijlichem, Paul Tjepke Robert van

    1994-01-01

    In the first part of the introduction (Chapter 1) of this thesis an overview of the numerous techniques used in islet isolation procedures is presented. The differing lines of approach for the dissociation of the pancreas which have been applied, and are still being further developed, indicate that

  18. Composite mandibular allografts in canines

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective: To evaluate the feasibility of transplanting composite mandibular allografts to repair large mandibular defects. Methods: Three composite mandibular transplantation models were established. The first model consisted of hemimandible with the attached teeth, muscle and skin, and oral mucosa. The second model was transplanted in the same way with the first one excluding oral mucosa and some teeth, and third one excluding the oral mucosa and all dental crowns. Fourteen transplanting operations were performed in canines. Cyclosporine A and methylprednisone were given for immunosuppression. Results: The composite mandibular organs had an effective and closed return circuit. Transplantation of vascularized allograft of mandibular compound organs was feasible. Two longest time survivors of 67 d and 76 d were in the third model group. Cyclosporine A was successful in suppressing rejection of transplanted composite allograft and prolonging survival time of transplantation models. Conclusions: The composite mandibular allografts were available with large block of living composite tissue,and helpful in restoration of appearance and function for severe mandibular defects.

  19. Emphysema in the renal allograft

    Energy Technology Data Exchange (ETDEWEB)

    Potter, J.L.; Sullivan, B.M.; Fluornoy, J.G.; Gerza, C.

    1985-04-01

    Two diabetic patients in whom emphysematous pyelonephritis developed after renal transplantation are described. Clinical recognition of this unusual and serious infection is masked by the effects of immunosuppression. Abdominal radiographic, ultrasound, and computed tomography findings are discussed. The clinical presentation includes urinary tract infection, sepsis, and acute tubular malfunction of the allograft in insulin-dependent diabetics.

  20. Classification of microscopy images of Langerhans islets

    Science.gov (United States)

    Å vihlík, Jan; Kybic, Jan; Habart, David; Berková, Zuzana; Girman, Peter; Kříž, Jan; Zacharovová, Klára

    2014-03-01

    Evaluation of images of Langerhans islets is a crucial procedure for planning an islet transplantation, which is a promising diabetes treatment. This paper deals with segmentation of microscopy images of Langerhans islets and evaluation of islet parameters such as area, diameter, or volume (IE). For all the available images, the ground truth and the islet parameters were independently evaluated by four medical experts. We use a pixelwise linear classifier (perceptron algorithm) and SVM (support vector machine) for image segmentation. The volume is estimated based on circle or ellipse fitting to individual islets. The segmentations were compared with the corresponding ground truth. Quantitative islet parameters were also evaluated and compared with parameters given by medical experts. We can conclude that accuracy of the presented fully automatic algorithm is fully comparable with medical experts.

  1. Adefovir nephrotoxicity in a renal allograft recipient

    Directory of Open Access Journals (Sweden)

    N George

    2015-01-01

    Full Text Available Adefovir dipivoxil, an oral prodrug of adefovir, is used in the treatment of lamivudine-resistant hepatitis B virus (HBV infection. Nephrotoxicity manifesting as proximal renal tubular dysfunction and acute tubular necrosis (ATN were commonly reported in the past, when higher doses were used for the treatment of human immunodeficiency virus infection. However, nephrotoxicity is rare at lower doses that are currently recommended for the treatment of HBV infection. A 31-year-old female was detected to be hepatitis B surface antigen positive months after a kidney transplant. The patient was initiated on lamivudine, but developed resistance after 1 year of treatment, at which time low-dose adefovir was added. The patient developed renal allograft dysfunction after 10 months of starting adefovir. Serum creatinine increased from 1.1 mg/dl to 1.9 mg/dl, along with progressively increasing sub-nephrotic proteinuria. Renal allograft biopsy revealed features of ATN. After discontinuation of adefovir, proteinuria resolved and renal dysfunction improved slowly over the next 2 years. Adefovir-induced nephrotoxicity, although uncommon at lower doses, needs to be considered in the differential diagnosis of renal dysfunction and sub-nephrotic proteinuria occurring in patients receiving adefovir for prolonged periods.

  2. Interleukin 1 dose-dependently affects the biosynthesis of (pro)insulin in isolated rat islets of Langerhans

    DEFF Research Database (Denmark)

    Spinas, G A; Hansen, B S; Linde, S

    1987-01-01

    Human crude and recombinant interleukin 1 (IL-1) was found to dose- and time-dependently affect the biosynthesis of (pro)insulin in isolated rat islets of Langerhans. Incubation of rat islets with either 0.5 U/ml or 5 U/ml of crude IL-1 for 1 h had no detectable effect on (pro)insulin biosynthesis...

  3. The Beta Cell in Its Cluster: Stochastic Graphs of Beta Cell Connectivity in the Islets of Langerhans.

    Directory of Open Access Journals (Sweden)

    Deborah A Striegel

    2015-08-01

    Full Text Available Pancreatic islets of Langerhans consist of endocrine cells, primarily α, β and δ cells, which secrete glucagon, insulin, and somatostatin, respectively, to regulate plasma glucose. β cells form irregular locally connected clusters within islets that act in concert to secrete insulin upon glucose stimulation. Due to the central functional significance of this local connectivity in the placement of β cells in an islet, it is important to characterize it quantitatively. However, quantification of the seemingly stochastic cytoarchitecture of β cells in an islet requires mathematical methods that can capture topological connectivity in the entire β-cell population in an islet. Graph theory provides such a framework. Using large-scale imaging data for thousands of islets containing hundreds of thousands of cells in human organ donor pancreata, we show that quantitative graph characteristics differ between control and type 2 diabetic islets. Further insight into the processes that shape and maintain this architecture is obtained by formulating a stochastic theory of β-cell rearrangement in whole islets, just as the normal equilibrium distribution of the Ornstein-Uhlenbeck process can be viewed as the result of the interplay between a random walk and a linear restoring force. Requiring that rearrangements maintain the observed quantitative topological graph characteristics strongly constrained possible processes. Our results suggest that β-cell rearrangement is dependent on its connectivity in order to maintain an optimal cluster size in both normal and T2D islets.

  4. Islet cryopreservation: improved recovery following taurine pretreatment.

    Science.gov (United States)

    Hardikar, A A; Risbud, M V; Remacle, C; Reusens, B; Hoet, J J; Bhonde, R R

    2001-01-01

    Simple and efficient freezing methods with maximal postthawing recovery form the basis of ideal cryopreservation. Taurine (2-amino ethanesulfonic acid), an end-product of sulphur amino acid metabolism, is one of the most abundant free amino acids in the body. The membrane stabilizing, free radical scavenging, and osmoregulatory roles of taurine have been well documented. We studied the effect of physiological and supra-physiological concentrations (0.3 and 3.0 mM) of taurine on islet cryopreservation. Islet viability on cryopreservation was significantly improved in both the taurine-treated groups (91.9 +/- 2.3% in 0.3 mM and 94.6 +/- 1.58% in 3.0 mM group, p taurine group, as examined under phase contrast and quantified by islet morphometric analysis (p Taurine-treated islets showed significant reduction in lipid peroxidation (0.905 and 0.848 nM MDA/microg protein for 0.3 and 3.0 mM taurine, respectively, p 200 mg/dl) following removal of the graft. Suboptimal islet transplantation using 250 IE suggests that the grafted islet mass was inadequate for diabetes reversal. In addition, no significant differences were observed in the islet insulin content between the three groups following cryopreservation of the islets at -196 degrees C. Our studies indicate that taurine pretreatment and its continued presence during islet cryopreservation improves the postthawing viable recovery of islets.

  5. Effects of mycophenolate mofetil vs cyclosporine administration on graft survival and function after islet allotransplantation in diabetic rats

    Institute of Scientific and Technical Information of China (English)

    Constantin Fotiadis; Paraskevi Xekouki; Apostolos E Papalois; Pantelis T Antonakis; Ioannis Sfiniadakis; Dimitrios Flogeras; Eleutheria Karampela; George Zografos

    2005-01-01

    AIM: To develop an experimental model of islet allotransplantation in diabetic rats and to determine the positive or adverse effects of MMF as a single agent. METHODS: Thirty-six male Wistar rats and 18 male Lewis rats were used as recipients and donors respectively. Diabetes was induced by the use of streptozotocin (60 mg/kg) intraperitoneally. Unpurified islets were isolated using the collagenase digestion technique and transplanted into the splenic parenchyma. The recipients were randomly assigned to one of the following three groups: group A (control group) had no immunosuppression; group B received cyclosporine (CsA) (5 mg/kg); group C receivedmycophenolate mofetil (MMF) (20 mg/kg). The animalswere killed on the 12th d. Blood and grafted tissues were obtained for laboratory and histological assessment. RESULTS: Median allograft survival was significantly higher in the two therapy groups than that in the controls (10 and 12 d for CsA and MMF respectively vs 0 d for the control group, P<0.01). No difference in allograft survival between the CsA and MMF groups was found. However,MMF had less renal and hepatic toxicity and allowed weight gain.CONCLUSION: Monotherapy with MMF for immunosu ppression was safe in an experimental model of islet allotransplantation and was equally effective with cyclosporine, with less toxicity.

  6. Progress in Clinical Encapsulated Islet Xenotransplantation.

    Science.gov (United States)

    Cooper, David K C; Matsumoto, Shinichi; Abalovich, Adrian; Itoh, Takeshi; Mourad, Nizar I; Gianello, Pierre R; Wolf, Eckhard; Cozzi, Emanuele

    2016-11-01

    At the 2015 combined congress of the Cell Transplant Society, International Pancreas and Islet Transplant Association, and International Xenotransplantation Association, a symposium was held to discuss recent progress in pig islet xenotransplantation. The presentations focused on 5 major topics - (1) the results of 2 recent clinical trials of encapsulated pig islet transplantation, (2) the inflammatory response to encapsulated pig islets, (3) methods to improve the secretion of insulin by pig islets, (4) genetic modifications to the islet-source pigs aimed to protect the islets from the primate immune and/or inflammatory responses, and (5) regulatory aspects of clinical pig islet xenotransplantation. Trials of microencapsulated porcine islet transplantation to treat unstable type 1 diabetic patients have been associated with encouraging preliminary results. Further advances to improve efficacy may include (1) transplantation into a site other than the peritoneal cavity, which might result in better access to blood, oxygen, and nutrients; (2) the development of a more biocompatible capsule and/or the minimization of a foreign body reaction; (3) pig genetic modification to induce a greater secretion of insulin by the islets, and/or to reduce the immune response to islets released from damaged capsules; and (4) reduction of the inflammatory response to the capsules/islets by improvements in the structure of the capsules and/or in genetic engineering of the pigs and/or in some form of drug therapy. Ethical and regulatory frameworks for islet xenotransplantation are already available in several countries, and there is now a wider international perception of the importance of developing an internationally harmonized ethical and regulatory framework.

  7. Anterior cruciate ligament reconstruction with allograft tendons.

    Science.gov (United States)

    Strickland, Sabrina M; MacGillivray, John D; Warren, Russell F

    2003-01-01

    Allograft tissue allows reconstruction of the ACL without the donor site morbidity that can be caused by autograft harvesting. Patients who must kneel as a part of their occupation or chosen sport are particularly good candidates for allograft reconstruction. Patients over 45 years of age and those requiring revision ACL surgery can also benefit from the use and availability of allograft tendons. In some cases, patients or surgeons may opt for allograft tendons to maximize the result or morbidity ratio. Despite advances in cadaver screening and graft preparation, there remain risks of disease transmission and joint infection after allograft implantation. Detailed explanation and informed consent is vitally important in cases in which allograft tissue is used.

  8. Mucormycosis (zygomycosis) of renal allograft.

    Science.gov (United States)

    Gupta, Krishan L; Joshi, Kusum; Kohli, Harbir S; Jha, Vivekanand; Sakhuja, Vinay

    2012-12-01

    Fungal infection is relatively common among renal transplant recipients from developing countries. Mucormycosis, also known as zygomycosis, is one of the most serious fungal infections in these patients. The most common of presentation is rhino-cerebral. Isolated involvement of a renal allograft is very rare. A thorough search of literature and our medical records yielded a total of 24 cases with mucormycosis of the transplanted kidney. There was an association with cytomegalovirus (CMV) infection and anti-rejection treatment in these patients and most of these transplants were performed in the developing countries from unrelated donors. The outcome was very poor with an early mortality in 13 (54.5%) patients. Renal allograft mucormycosis is a relatively rare and potentially fatal complication following renal transplantation. Early diagnosis, graft nephrectomy and appropriate antifungal therapy may result in an improved prognosis for these patients.

  9. Leiomyoma in a Renal Allograft

    Directory of Open Access Journals (Sweden)

    Yan Jun Li

    2016-01-01

    Full Text Available Leiomyomas are smooth muscle tumours that are rarely found in the kidney. There is one report of a leiomyoma in a kidney transplant in a paediatric recipient. Here, we report an adult renal transplant recipient who developed an Epstein-Barr virus-positive leiomyoma in his allograft 15 years after transplantation. The patient was converted to everolimus for posttransplant immunosuppression management and there was no sign of progression over a year.

  10. Genetic regulatory signatures underlying islet gene expression and type 2 diabetes

    Science.gov (United States)

    Varshney, Arushi; Scott, Laura J.; Welch, Ryan P.; Erdos, Michael R.; Chines, Peter S.; Narisu, Narisu; Albanus, Ricardo D’O.; Orchard, Peter; Wolford, Brooke N.; Kursawe, Romy; Vadlamudi, Swarooparani; Cannon, Maren E.; Didion, John P.; Hensley, John; Kirilusha, Anthony; Bonnycastle, Lori L.; Taylor, D. Leland; Watanabe, Richard; Mohlke, Karen L.; Boehnke, Michael; Collins, Francis S.; Parker, Stephen C. J.; Stitzel, Michael L.

    2017-01-01

    Genome-wide association studies (GWAS) have identified >100 independent SNPs that modulate the risk of type 2 diabetes (T2D) and related traits. However, the pathogenic mechanisms of most of these SNPs remain elusive. Here, we examined genomic, epigenomic, and transcriptomic profiles in human pancreatic islets to understand the links between genetic variation, chromatin landscape, and gene expression in the context of T2D. We first integrated genome and transcriptome variation across 112 islet samples to produce dense cis-expression quantitative trait loci (cis-eQTL) maps. Additional integration with chromatin-state maps for islets and other diverse tissue types revealed that cis-eQTLs for islet-specific genes are specifically and significantly enriched in islet stretch enhancers. High-resolution chromatin accessibility profiling using assay for transposase-accessible chromatin sequencing (ATAC-seq) in two islet samples enabled us to identify specific transcription factor (TF) footprints embedded in active regulatory elements, which are highly enriched for islet cis-eQTL. Aggregate allelic bias signatures in TF footprints enabled us de novo to reconstruct TF binding affinities genetically, which support the high-quality nature of the TF footprint predictions. Interestingly, we found that T2D GWAS loci were strikingly and specifically enriched in islet Regulatory Factor X (RFX) footprints. Remarkably, within and across independent loci, T2D risk alleles that overlap with RFX footprints uniformly disrupt the RFX motifs at high-information content positions. Together, these results suggest that common regulatory variations have shaped islet TF footprints and the transcriptome and that a confluent RFX regulatory grammar plays a significant role in the genetic component of T2D predisposition. PMID:28193859

  11. 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.

  12. Fibrillar dimer formation of islet amyloid polypeptides

    Energy Technology Data Exchange (ETDEWEB)

    Chiu, Chi-cheng [Univ. of Chicago, IL (United States); Argonne National Lab. (ANL), Argonne, IL (United States); de Pablo, Juan J. [Univ. of Chicago, IL (United States); Argonne National Lab. (ANL), Argonne, IL (United States)

    2015-05-08

    Amyloid deposits of human islet amyloid polypeptide (hIAPP), a 37-residue hormone co-produced with insulin, have been implicated in the development of type 2 diabetes. Residues 20 – 29 of hIAPP have been proposed to constitute the amyloidogenic core for the aggregation process, yet the segment is mostly unstructured in the mature fibril, according to solid-state NMR data. Here we use molecular simulations combined with bias-exchange metadynamics to characterize the conformational free energies of hIAPP fibrillar dimer and its derivative, pramlintide. We show that residues 20 – 29 are involved in an intermediate that exhibits transient β-sheets, consistent with recent experimental and simulation results. By comparing the aggregation of hIAPP and pramlintide, we illustrate the effects of proline residues on inhibition of the dimerization of IAPP. The mechanistic insights presented here could be useful for development of therapeutic inhibitors of hIAPP amyloid formation.

  13. Calpain-10 expression is elevated in pancreatic islets from patients with type 2 diabetes.

    Directory of Open Access Journals (Sweden)

    Charlotte Ling

    Full Text Available BACKGROUND: Calpain-10 was the first gene to be identified influencing the risk of type 2 diabetes (T2D by positioning cloning. Studies in beta-cell lines and rodent islets suggest that calpain-10 may act as a regulator of insulin secretion. However, its role in human pancreatic islets remains unclear. The aim of this study was to examine if calpain-10 expression is altered in islets from patients with T2D and if the transcript level correlates with insulin release. We also tested if polymorphisms in the CAPN10 gene are associated with gene expression and insulin secretion in vitro. METHODOLOGY/PRINCIPAL FINDINGS: Calpain-10 mRNA expression was analysed in human pancreatic islets from 34 non-diabetic and 10 T2D multi-organ donors. CAPN10 SNP-43 and SNP-44 were genotyped and related to gene expression and insulin release in response to glucose, arginine and glibenclamide. The mRNA level of calpain-10 was elevated by 64% in pancreatic islets from patients with T2D compared with non-diabetic donors (P = 0.01. Moreover, the calpain-10 expression correlated positively with arginine-stimulated insulin release in islets from non-diabetic donors (r = 0.45, P = 0.015. However, this correlation was lost in islets from patients with T2D (r = 0.09; P = 0.8. The G/G variant of SNP-43 was associated with reduced insulin release in response to glucose (Phuman islets, this correlation is lost in T2D suggesting that a stimulatory effect of calpain-10 could be lost in patients with T2D.

  14. A health-economic analysis of porcine islet xenotransplantation.

    Science.gov (United States)

    Beckwith, Jessica; Nyman, John A; Flanagan, Brian; Schrover, Rudolf; Schuurman, Henk-Jan

    2010-01-01

    Islet cell transplantation is a promising treatment for type 1 diabetes. To overcome the shortage of deceased human pancreas donors, porcine islet cell xenotransplantation is being developed as an alternative to allotransplantation. The objective of this study was to perform a cost-effectiveness analysis of porcine islet transplantation in comparison with standard insulin therapy. The patient population for this study was young adults, ages 20 to 40, for whom standard medical care is inadequate in controlling blood glucose levels (hypoglycemia unawareness). Since trial data were lacking, estimates used extrapolations from data found in the literature and ongoing trials in clinical allotransplantation. Cost estimates were based on the data available in the USA. Markov modeling and Monte Carlo simulations using software specifically developed for health-economic evaluations were used. Outcomes data for ongoing clinical islet allotransplantation from the University of Minnesota were used, along with probabilities of complications from the Diabetes Control and Complications Trial. Quality-adjusted life years (QALYs) were the effectiveness measure. The upper limit of being cost-effective is $100,000 per QALY. Cost data from the literature were used and adjusted to 2007 US dollars using the medical care portion of the Consumer Price Index. In both Markov modeling and Monte Carlo simulations, porcine islet xenotransplantation was both more effective and less costly over the course of the 20-yr model. For standard insulin therapy, cumulative cost per patient was $661,000, while cumulative effectiveness was 9.4 QALYs, for a cost of $71,100 per QALY. Transplantation had a cumulative cost of $659 000 per patient, a cumulative effectiveness of 10.9 QALYs, and a cost per QALY of $60,700. Islet transplantation became cost-effective at 4 yr after transplantation, and was more cost-effective than standard insulin treatment at 14 yr. These findings are related to relative high

  15. Expression of ICAM-1 and VCAM-1 in human chronic renal allograft rejection%细胞间粘附分子-1和血管细胞粘附分子-1在慢性排斥反应中的表达

    Institute of Scientific and Technical Information of China (English)

    潘晓鸣; 陈勇; 邢俊平

    1998-01-01

    To study the mechanism of human chronic renal allograft rejection, kidney tissues were taken from 16 patients with chronic renal allograft rejection and from 5 healthy subjects, and underwent the frazed section staining for ICAM-1 and VCAM-1 to anti-ICAM-1 and anti-VCAM-1 respectively by using immunohistochemistry(ABC).The results showed that there were differ-ent distribution of ICAM-1 and VCAM-1 expression in nomal kidney and renal allograft during chronic rejection.It was suggested that ICAM-1 and VCAM-1 might play an important role in the pathogenesis of human chronic renal allograft rejection.%为了探讨移植肾慢性排斥反应的发病机制,应用免疫组化技术(ABC法)对16例肾移植术后发生慢性排斥反应患者的移植肾组织及5例正常肾组织行细胞间粘附分子-1(ICAM-1)、血管细胞粘附分子-1(VCAM-1)染色及HE染色.结果表明ICAM-1、VCAM-1在正常肾脏和慢性排斥反应移植肾脏上的表达分布不同;结果提示,它们在移植肾慢性排斥反应的发生、发展过程中起重要作用

  16. Optimization and Implementation of Long Nerve Allografts

    Science.gov (United States)

    2014-10-01

    nerve tissue requires a graft to restore continuity and promote nerve regeneration and recovery of function. Presently, there is no acceptable nerve ...for nerve regeneration and meaningful recovering of nerve function that, in several cases was better than autografting. Other decellularized allografts... nerve graft, allograft, nerve regeneration , rehabilitation 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18. NUMBER OF PAGES 19a. NAME

  17. Control of islet intercellular adhesion molecule-1 expression by interferon-alpha and hypoxia.

    Science.gov (United States)

    Chakrabarti, D; Huang, X; Beck, J; Henrich, J; McFarland, N; James, R F; Stewart, T A

    1996-10-01

    The ability of interferon-alpha (IFN-alpha) to induce the adhesion molecules that characterize the islets of patients with type I diabetes has been investigated. We have found that all tested recombinant IFN-as will induce major histocompatibility complex (MHC) class I on arterial endothelial cells. Some but not all IFN-as will induce intercellular adhesion molecule-1 (ICAM-1). However, there is only a transient and modest increase in VCAM on arterial endothelial cells. IFN-alpha has very little effect on endothelial MHC class II expression but will induce these proteins on monocytes. Thus, there is a close concordance between the biological actions of IFN-alpha and the appearance of those adhesion molecules induced in the islets of patients with type I diabetes. IFN-alpha is also produced in normal human islets during short-term cultures, probably as a result of the ischemia present at the center of the islet. This induction of IFN-alpha by hypoxia may explain the previously reported spontaneous induction of ICAM-1 in human islets and may also be a contributing factor to the failure of islet grafts.

  18. The Mitochondrial Peptidase Pitrilysin Degrades Islet Amyloid Polypeptide in Beta-Cells.

    Directory of Open Access Journals (Sweden)

    Hanjun Guan

    Full Text Available Amyloid formation and mitochondrial dysfunction are characteristics of type 2 diabetes. The major peptide constituent of the amyloid deposits in type 2 diabetes is islet amyloid polypeptide (IAPP. In this study, we found that pitrilysin, a zinc metallopeptidase of the inverzincin family, degrades monomeric, but not oligomeric, islet amyloid polypeptide in vitro. In insulinoma cells when pitrilysin expression was decreased to 5% of normal levels, there was a 60% increase in islet amyloid polypeptide-induced apoptosis. In contrast, overexpression of pitrilysin protects insulinoma cells from human islet amyloid polypeptide-induced apoptosis. Since pitrilysin is a mitochondrial protein, we used immunofluorescence staining of pancreases from human IAPP transgenic mice and Western blot analysis of IAPP in isolated mitochondria from insulinoma cells to provide evidence for a putative intramitochondrial pool of IAPP. These results suggest that pitrilysin regulates islet amyloid polypeptide in beta cells and suggest the presence of an intramitochondrial pool of islet amyloid polypeptide involved in beta-cell apoptosis.

  19. Acellular nerve allograft promotes selective regeneration

    Institute of Scientific and Technical Information of China (English)

    Haili Xin; Guanjun Wang; Xinrong He; Jiang Peng; Quanyi Guo; Wenjing Xu

    2011-01-01

    Acellular nerve allograft preserves the basilar membrane tube and extracellular matrix, which pro-motes selective regeneration of neural defects via bridging. In the present study, a Sprague Dawley rat sciatic nerve was utilized to prepare acellular nerve allografts through the use of the chemical extraction method. Subsequently, the allograft was transplanted into a 10-mm sciatic nerve defect in Wistar rats, while autologous nerve grafts from Wistar rats served as controls. Compared with autologous nerve grafts, the acellular nerve allografts induced a greater number of degenerated nerve fibers from sural nerves, as well as a reduced misconnect rate in motor fibers, fewer acetyl-choline esterase-positive sural nerves, and a greater number of carbonic anhydrase-positive senso-ry nerve fibers. Results demonstrated that the acellular nerve allograft exhibited significant neural selective regeneration in the process of bridging nerve defects.

  20. A stereological study of effects of aqueous extract of Tamarindus indica seeds on pancreatic islets in streptozotocin-induced diabetic rats.

    Science.gov (United States)

    Hamidreza, Hamidreza; Heidari, Zahra; Shahraki, Mohammadreza; Moudi, Bita

    2010-10-01

    Tamarindus indica Linn was used as a traditional medicine for the management of diabetes mellitus in human and experimental animals. This study investigated effects of aqueous extract of Tamarindus indica seeds (AETIS) against STZ-induced damages in pancreatic islands by means of stereological methods. sixty matured normoglycemic male Wistar rats, weighing 200-250 gr, were selected and randomly divided into 6 groups (n=10). Control, STZ-induced diabetic; by intraperitoneal injection of 55 mg/Kg streptozotocin, Treated control group (TC); received AETIS at a dose of 200mg/kg/day, and AETIS treated diabetic groups (TD1-3); received respectively AETIS at the dose of 50, 100,and 200 mg/kg/day by gavage from one week after induction of diabetes by STZ. After 8 weeks of experiment, stereological estimation of volume density and total volume of islets and beta cells, volume weighted mean islets volume, mass of beta cells, islets, and pancreas and total number of islets were done. Volume density and total volume of islets, volume weighted mean islets volume, volume density islets/pancreas, volume density beta cells/islet, mass of islets and pancreas of treated diabetic groups (TD1-3) were significantly higher than untreated diabetic group (P0.05). Total number of islets, pancreas wet weight and volume did not show any significant changes between control and experimental groups (P>0.05). Results suggested that AETIS partially restores pancreatic beta cells and repairs STZ-induced damages in rats.

  1. Type I and II Diabetic Adipose-Derived Stem Cells Respond In Vitro to Dehydrated Human Amnion/Chorion Membrane Allograft Treatment by Increasing Proliferation, Migration, and Altering Cytokine Secretion.

    Science.gov (United States)

    Massee, Michelle; Chinn, Kathryn; Lim, Jeremy J; Godwin, Lisa; Young, Conan S; Koob, Thomas J

    2016-02-01

    Objective: Human amniotic membranes have been shown to be effective for healing diabetic foot ulcers clinically and to regulate stem cell activity in vitro and in vivo; however, diabetic stem cells may be impaired as a sequela of the disease. In this study, dehydrated human amnion/chorion membrane (dHACM) allografts (EpiFix(®); MiMedx Group) were evaluated for their ability to regulate diabetic stem cells in vitro. Approach: Human adipose-derived stem cells (ADSCs) from normal, type I diabetic, and type II diabetic donors were treated with soluble extracts of dHACM and evaluated for proliferation after 3 days by DNA assay, chemotactic migration after 1 day by transwell assay, cytokine secretion after 3 days by multiplex ELISA, and gene expression after 5 days by reverse transcription-polymerase chain reaction. Results: Although diabetic ADSCs demonstrated decreased responses compared to normal ADSCs, dHACM treatment stimulated diabetic ADSCs to proliferate after 3 days and enhanced migration over 24 h, similar to normal ADSCs. dHACM-treated diabetic ADSCs modulated secretion of soluble signals, including regulators of inflammation, angiogenesis, and healing. All ADSCs evaluated also responded to dHACM treatment with altered expression of immunomodulatory genes, including interleukins (IL)-1α, IL-1β, and IL-1RA. Innovation: This is the first reported case demonstrating that diabetic ADSCs respond to novel amniotic membrane therapies, specifically treatment with dHACM. Conclusion: dHACM stimulated diabetic ADSCs to migrate, proliferate, and alter cytokine expression suggesting that, despite their diabetic origin, ADSCs may respond to dHACM to accelerate diabetic wound healing.

  2. Rac1 regulates pancreatic islet morphogenesis.

    OpenAIRE

    2009-01-01

    Abstract Background Pancreatic islets of Langerhans originate from endocrine progenitors within the pancreatic ductal epithelium. Concomitant with differentiation of these progenitors into hormone-producing cells such cells delaminate, aggregate and migrate away from the ductal epithelium. The cellular and molecular mechanisms regulating islet cell delamination and cell migration are poorly understood. Extensive biochemical and cell biological studies using cultured cells demonstrated that Ra...

  3. Late Failing Heart Allografts: Pathology of Cardiac Allograft Vasculopathy and Association With Antibody-Mediated Rejection.

    Science.gov (United States)

    Loupy, A; Toquet, C; Rouvier, P; Beuscart, T; Bories, M C; Varnous, S; Guillemain, R; Pattier, S; Suberbielle, C; Leprince, P; Lefaucheur, C; Jouven, X; Bruneval, P; Duong Van Huyen, J P

    2016-01-01

    In heart transplantation, there is a lack of robust evidence of the specific causes of late allograft failure. We hypothesized that a substantial fraction of failing heart allografts may be associated with antibody-mediated injury and immune-mediated coronary arteriosclerosis. We included all patients undergoing a retransplantation for late terminal heart allograft failure in three referral centers. We performed an integrative strategy of heart allograft phenotyping by assessing the heart vascular tree including histopathology and immunohistochemistry together with circulating donor-specific antibodies. The main analysis included 40 explanted heart allografts patients and 402 endomyocardial biopsies performed before allograft loss. Overall, antibody-mediated rejection was observed in 19 (47.5%) failing heart allografts including 16 patients (40%) in whom unrecognized previous episodes of subclinical antibody-mediated rejection occurred 4.5 ± 3.5 years before allograft loss. Explanted allografts with evidence of antibody-mediated rejection demonstrated higher endothelitis and microvascular inflammation scores (0.89 ± 0.26 and 2.25 ± 0.28, respectively) compared with explanted allografts without antibody-mediated rejection (0.42 ± 0.11 and 0.36 ± 0.09, p = 0.046 and p < 0.0001, respectively). Antibody-mediated injury was observed in 62.1% of failing allografts with pure coronary arteriosclerosis and mixed (arteriosclerosis and atherosclerosis) pattern, while it was not observed in patients with pure coronary atherosclerosis (p = 0.0076). We demonstrate that antibody-mediated rejection is operating in a substantial fraction of failing heart allografts and is associated with severe coronary arteriosclerosis. Unrecognized subclinical antibody-mediated rejection episodes may be observed years before allograft failure.

  4. Canine islets in an ultrafiltered environment.

    Science.gov (United States)

    Merrell, R C; Basadonna, G

    1985-11-01

    Molecular sieve membranes can protect pancreatic islets against immune recognition in diabetic patients treated by endocrine tissue replacement. These biocompatible membranes permit the passage of small peptides such as insulin, and preclude the diffusion of immunoglobulins and immunogenic molecules. However, the tissue must function indefinitely in an ultrafiltered environment determined by the sequestering membranes. The chronic perifusion of canine islet tissue was compared in ultrafiltered and microfiltered chambers. The biphasic pattern of insulin release by similar numbers of islets from the same pancrease preparation was not significantly different when tissue was cultured in a micro- or an ultrafiltered environment. The cumulative insulin output of the two systems was quite similar over 3 days of culture. Canine islet tissue can be sustained in an ultrafiltered environment with maintenance of insulin release to glucose stimulation, which is quantitatively similar to islet tissue maintained in chronic perifusion without ultrafiltration.

  5. Rac1 regulates pancreatic islet morphogenesis

    Directory of Open Access Journals (Sweden)

    Ståhlberg Anders

    2009-01-01

    Full Text Available Abstract Background Pancreatic islets of Langerhans originate from endocrine progenitors within the pancreatic ductal epithelium. Concomitant with differentiation of these progenitors into hormone-producing cells such cells delaminate, aggregate and migrate away from the ductal epithelium. The cellular and molecular mechanisms regulating islet cell delamination and cell migration are poorly understood. Extensive biochemical and cell biological studies using cultured cells demonstrated that Rac1, a member of the Rho family of small GTPases, acts as a key regulator of cell migration. Results To address the functional role of Rac1 in islet morphogenesis, we generated transgenic mice expressing dominant negative Rac1 under regulation of the Rat Insulin Promoter. Blocking Rac1 function in beta cells inhibited their migration away from the ductal epithelium in vivo. Consistently, transgenic islet cell spreading was compromised in vitro. We also show that the EGF-receptor ligand betacellulin induced actin remodelling and cell spreading in wild-type islets, but not in transgenic islets. Finally, we demonstrate that cell-cell contact E-cadherin increased as a consequence of blocking Rac1 activity. Conclusion Our data support a model where Rac1 signalling controls islet cell migration by modulating E-cadherin-mediated cell-cell adhesion. Furthermore, in vitro experiments show that betacellulin stimulated islet cell spreading and actin remodelling is compromised in transgenic islets, suggesting that betacellulin may act as a regulator of Rac1 activity and islet migration in vivo. Our results further emphasize Rac1 as a key regulator of cell migration and cell adhesion during tissue and organ morphogenesis.

  6. Pancreatic islet renin angiotensin system: its novel roles in islet function and in diabetes mellitus.

    Science.gov (United States)

    Leung, Po Sing; Carlsson, Per-Ola

    2005-05-01

    Several regulatory systems are implicated in the regulation of islet function and beta cell mass. Of great interest in this context are some endocrine, paracrine/autocrine, and intracrine regulators. These include, to name but a few, the gut peptides, growth factors, prostaglandins, and some vasoactive mediators such as nitric oxide, bradykinins, endothelins, and angiotensins. Apart from its potent vasoconstrictor actions, the renin-angiotensin system (RAS) that generates angiotensin II has several novel functions-stimulation and inhibition of cell proliferation; induction of apoptosis; generation of reactive oxygen species; regulation of hormone secretion; and proinflammatory and profibrogenic actions. In the pancreas, recent evidence supports the presence of an islet RAS, which is subject to activation by islet transplantation and diabetes. Such a local islet RAS, if activated, may drive islet fibrosis and reduce islet blood flow, oxygen tension, and insulin biosynthesis. Moreover, activation of an islet RAS may drive the synthesis of reactive oxygen species, cause oxidative stress-induced beta cell dysfunction and apoptosis, and thus contribute to the islet dysfunction seen in type 2 diabetes and after islet transplantation. Blockade of the RAS could contribute to the development of novel therapeutic strategies in the prevention and treatment of patients with diabetes and in islet transplantation.

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

    Directory of Open Access Journals (Sweden)

    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.

  8. Allograft safety in anterior cruciate ligament reconstruction.

    Science.gov (United States)

    Cohen, Steven B; Sekiya, Jon K

    2007-10-01

    Allograft tissue seems to provide an excellent option for reconstruction of the ACL in the primary and revision setting. Although in general the risks of using allograft tissue in ACL reconstruction are low, the consequences of complications associated with disease or infection transmission or of recurrent instability secondary to graft failure are large. Surgeons should provide patients with the information available regarding allograft risks and should have thorough knowledge of the source and preparation of the grafts by their tissue bank before implantation for ACL reconstruction.

  9. Clinical use of fructosamine in islet transplantation.

    Science.gov (United States)

    Tharavanij, Thipaporn; Froud, Tatiana; Leitao, Cristiane B; Baidal, David A; Paz-Pabon, Charlotte N; Shari, Messinger; Cure, Pablo; Bernetti, Karina; Ricordi, Camillo; Alejandro, Rodolfo

    2009-01-01

    Many islet transplant recipients have medical conditions that could interfere with the accuracy of HbA1c measurements (e.g., anemia/dapsone use). Fructosamine is less prone to have clinical interferences and reflects glucose control in a shorter period of time than HbA1c. This study aimed to validate fructosamine use in islet transplant subjects and to evaluate its effectiveness as a predictor for islet graft dysfunction. Thirty-three islet transplant recipients who had concomitant fructosamine and HbA1c data available were retrospectively analyzed. HbA1c, fructosamine, mean capillary blood glucose, and islet graft function (fasting C-peptide/glucose ratio) were assessed. There was a significant and positive association between fructosamine and HbA1c (p 6% was predictive of this outcome 1 month in advance (OR 2.95, p = 0.003). However, although significantly associated with graft dysfunction, use of this cutoff as a predictor of dysfunction has poor sensitivity (50%) and specificity (77.6%). Fructosamine above the normal range (>270 mumol/L Quest Diagnostics) was also predictive of ensuing dysfunction (OR 2.47, p = 0.03); however, it had similarly poor sensitivity (62%) and specificity (64%). Fructosamine can be used as an alternative to HbA1c for glycemic assessment in islet transplant recipients in situations with HbA1c assay interference. Neither HbA1c nor fructosamine are good predictors of islet graft dysfunction.

  10. A clinical and radiological evaluation of the relative efficacy of demineralized freeze-dried bone allograft versus anorganic bovine bone xenograft in the treatment of human infrabony periodontal defects: A 6 months follow-up study

    Directory of Open Access Journals (Sweden)

    Vikram Blaggana

    2014-01-01

    Full Text Available Background: The ultimate goal of periodontal therapy entails regeneration of the periodontal tissues lost as a consequence of periodontitis. Predictable correction of vertical osseous defects has however posed as a constant therapeutic challenge. The aim of our present study is to evaluate the relative efficacy of demineralized freeze-dried bone allograft (DFDBA vs anorganic bovine bone xenograft (ABBX in the treatment of human infrabony periodontal defects. Materials and Methods: 15 patients with 30 bilaterally symmetrical defect sites in either of the arches, in the age group of 25-50 years were selected as part of split-mouth study design. Defect-A (right side was grafted with DFDBA while Defect-B (left side was grafted with ABBX. Various clinical and radiographic parameters viz. probing depth(PD, clinical attachment level(CAL and linear bone fill were recorded preoperatively, 12- & 24-weeks postoperatively. Results: Both defect-A & defect-B sites exhibited a highly significant reduction in probing depth, and gain in clinical attachment level and linear bone fill at 12-weeks & at the end of 24-weeks. Comparative evaluation between the study groups revealed a statistically non-significant reduction in probing depth (P<0.1 and mean gain in linear bone fill (P<0.1. However, there was a statistically significant gain in clinical attachment level (P <0.05 in Defect-A (CD=0.356 as compared to Defect-B (CD=0.346. Conclusions: Within the limits of this study, both the materials viz. ABBX and DFDBA are beneficial for the treatment of periodontal infrabony defects. Both the materials were found to be equally effective in all respects except the gain in attachment level, which was found to be more with DFDBA. Long-term studies are suggested to evaluate further the relative efficacy of the two grafts.

  11. Renal allograft rejection: sonography and scintigraphy

    Energy Technology Data Exchange (ETDEWEB)

    Singh, A.; Cohen, W.N.

    1980-07-01

    A total of 30 renal allograft patients who had sonographic B scanning and radionuclide studies of the transplant was studied as to whether: (1) the allograft rejection was associated with any consistent and reliable sonographic features and (2) the sonograms complemented the radionuclide studies. Focal areas of decreased parenchymal echogenicity were the most striking and consistent sonographic finding in chymal echogenicity were the most striking and consistens sonographic finding in allograft rejection. This was observed in most of the patients exhibiting moderate or severe rejection, but was frequently absent with mild rejection. Areas of decreased parenchymal echogenicity were not seen during episodes of acute tubular necrosis. Therefore, sonography showing zones of decreased parenchymal echogenicity was complementary to radionuclide studies in the diagnosis of allograft rejection versus acute tubular necrosis. Corticomedullary demarcation was difficult to interpret because of technical variables, and was inconsistently related to rejection in this series.

  12. Phycocyanin protects INS-1E pancreatic beta cells against human islet amyloid polypeptide-induced apoptosis through attenuating oxidative stress and modulating JNK and p38 mitogen-activated protein kinase pathways.

    Science.gov (United States)

    Li, Xiao-Ling; Xu, Gang; Chen, Tianfeng; Wong, Yum-Shing; Zhao, Hai-Lu; Fan, Rong-Rong; Gu, Xue-Mei; Tong, Peter C Y; Chan, Juliana C N

    2009-07-01

    It is widely accepted that human islet amyloid polypeptide (hIAPP) aggregation plays an important role in the loss of insulin-producing pancreatic beta cells. hIAPP-induced cytotoxicity is mediated by generation of reactive oxygen species (ROS). Phycocyanin (PC) is a natural compound from blue-green algae that is widely used as food supplement. Currently, little is known about the effects of PC on beta cells with the presence of hIAPP. The aim of this study was to investigate the in vitro protective effects of PC on INS-1E rat insulinoma beta cells against hIAPP-induced cell death, as well as the underlying mechanisms. Our results showed that hIAPP-induced cell death with apoptotic characteristics including growth inhibition, chromatin condensation and DNA fragmentation. However, cytotoxicity of hIAPP was significantly attenuated by co-incubation of the cells with PC. The results of Western blotting showed that activation of caspase-3 and cleavage of poly (ADP-ribose) polymerase (PARP) in hIAPP-treated cells was blocked by PC. Moreover, PC significantly prevented the hIAPP-induced overproduction of intracellular ROS and malondialdehyde (MDA), as well as changes in activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) enzymes. Furthermore, hIAPP triggered the activation of mitogen-activated protein kinases (MAPKs), and these effects were effectively suppressed by PC. Taken together, our results suggest that PC protects INS-1E pancreatic beta cells against hIAPP-induced apoptotic cell death through attenuating oxidative stress and modulating c-Jun N-terminal kinase (JNK) and p38 pathways.

  13. Selenium-enriched Spirulina protects INS-1E pancreatic beta cells from human islet amyloid polypeptide-induced apoptosis through suppression of ROS-mediated mitochondrial dysfunction and PI3/AKT pathway.

    Science.gov (United States)

    Li, Xiao-Ling; Wong, Yum-Shing; Xu, Gang; Chan, Juliana C N

    2015-06-01

    Human islet amyloid polypeptide (hIAPP) aggregation is linked to loss of pancreatic beta cells in type 2 diabetes, in part due to oxidative stress. Currently, little is known about the effects of selenium-enriched Spirulina on beta cells with the presence of hIAPP. In this study, INS-1E rat insulinoma cells were used as a model to evaluate in vitro protective effects of Se-enriched Spirulina extract (Se-SE) against hIAPP-induced cell death, as well as the underlying mechanisms. Flow cytometric analysis was used to evaluate cell apoptosis, mitochondrial membrane potential (ΔΨm) and ROS generation. Caspase activity was measured using a fluorometric method. Western blotting was applied to detect protein expression. Our results showed that exposure of INS-1E cells to hIAPP resulted in cell viability loss, LDH release and appearance of sub-G peak. However, cytotoxicity of hIAPP was significantly attenuated by co-treatment with Se-SE. Se-SE also inhibited hIAPP-induced activation of caspase-3, -8 and -9. Additionally, hIAPP-induced accumulation of ROS and superoxide was suppressed by co-treatment with Se-SE. Moreover, Se-SE was able to prevent hIAPP-induced depletion of ΔΨm and intracellular ATP, reduction in mitochondrial mass, changes in the expression of Bcl-2 family members, release of mitochondrial apoptogenic factors. Furthermore, hIAPP-mediated AKT inhibition was restored by co-treatment with Se-SE. Our results showed that Se-SE protects INS-1E cells from hIAPP-induced cell death through preventing ROS overproduction, mitochondrial dysfunction and modulating PI3K/AKT pathway.

  14. Suppression of islet homeostasis protein thwarts diabetes mellitus progression.

    Science.gov (United States)

    Oh, Seh-Hoon; Jorgensen, Marda L; Wasserfall, Clive H; Gjymishka, Altin; Petersen, Bryon E

    2017-05-01

    During progression to type 1 diabetes, insulin-producing β-cells are lost through an autoimmune attack resulting in unrestrained glucagon expression and secretion, activation of glycogenolysis, and escalating hyperglycemia. We recently identified a protein, designated islet homeostasis protein (IHoP), which specifically co-localizes within glucagon-positive α-cells and is overexpressed in the islets of both post-onset non-obese diabetic (NOD) mice and type 1 diabetes patients. Here we report that in the αTC1.9 mouse α-cell line, IHoP was released in response to high-glucose challenge and was found to regulate secretion of glucagon. We also show that in NOD mice with diabetes, major histocompatibility complex class II was upregulated in islets. In addition hyperglycemia was modulated in NOD mice via suppression of IHoP utilizing small interfering RNA (IHoP-siRNA) constructs/approaches. Suppression of IHoP in the pre-diabetes setting maintained normoglycemia, glyconeolysis, and fostered β-cell restoration in NOD mice 35 weeks post treatment. Furthermore, we performed adoptive transfer experiments using splenocytes from IHoP-siRNA-treated NOD/ShiLtJ mice, which thwarted the development of hyperglycemia and the extent of insulitis seen in recipient mice. Last, IHoP can be detected in the serum of human type 1 diabetes patients and could potentially serve as an early novel biomarker for type 1 diabetes in patients.

  15. Currently available useful immunohistochemical markers of renal pathology for the diagnosis of renal allograft rejection.

    Science.gov (United States)

    Kanzaki, Go; Shimizu, Akira

    2015-07-01

    Renal allograft dysfunction may be induced by various causes, including alloimmune rejection, viral infection, urinary tract obstruction, calcineurin inhibitor nephrotoxicity and/or recurrent renal disease. In order to determine the underlying cause, a renal biopsy is performed and the renal transplant pathology is diagnosed using the internationally consensus Banff classification. Although a progressive understanding of allograft rejection has provided numerous immunohistochemical markers, only the C4d is regarded to be a sufficiently useful marker for antibody-mediated allograft rejection according to the Banff classification. This review summarizes currently available useful immunohistochemical markers of renal transplant pathology, including C4d, with diagnostic implications for human renal allograft rejection. In particular, we discuss immunohistochemical markers in the following three categories: immunohistochemical markers of renal pathology used to (i) analyze the mechanisms of alloimmune rejection, (ii) monitor cell injury and/or inflammation associated with rejection and (iii) identify renal components in order to improve the diagnosis of rejection. In addition, recent progress in the field of renal transplant pathology includes the development of a new method for assessing molecular pathology using OMICS analyses. As the recent findings of various studies in patients undergoing renal transplantation are very encouraging, novel immunohistochemical markers must be also developed and combined with new technologies for the diagnosis of human renal allograft rejection.

  16. Specific unresponsiveness to skin allografts in burns.

    Science.gov (United States)

    Clark, G T; Moon, D J; Cunningham, P R; Johnson, T D; Thomas, J M; Thomas, F T

    1989-05-01

    We have examined the potential to provide long-term or even permanent wound coverage in a mouse model of a 30% total body surface area burn using skin allografts. Treatment of the recipient mouse with rabbit anti-mouse thymocyte serum (ATS) followed by donor bone marrow infusion induces a state of specific unresponsiveness to the skin allograft without the need for chronic immunosuppression. Specifically, a B6AF1 mouse receives a burn on Day -2 relative to grafting, ATS on Day -1, and Day +2, a skin allograft from a C3H/He mouse on Day 0, and infusion of C3H/He donor bone marrow on Day +6. We studied three groups of burned mice: Group I, allograft control (n = 5); Group II, allograft plus ATS (n = 12); and Group III, allograft plus ATS and bone marrow infusion (n = 15). Mean graft survival was compared using a one-way analysis of variance and a Student-Newman-Keuls post hoc test. There was no statistical difference in animal mortality among any of the three groups, and there was no evidence of infectious morbidity. Mean skin allograft survival was as follows: Group I, 9 days; Group II, 29 days; and Group III, 66 days (P less than 0.05 vs Group I and II). Nine animals in Group III had intact hair bearing grafts at 90 days when the study was terminated. This study suggests the potential use of induced specific unresponsiveness to skin allografts for wound coverage in thermal injury without use of chronic immunosuppression. In our animal study this was accomplished without increased mortality or apparent infectious morbidity.

  17. Computational Biology: Modeling Chronic Renal Allograft Injury.

    Science.gov (United States)

    Stegall, Mark D; Borrows, Richard

    2015-01-01

    New approaches are needed to develop more effective interventions to prevent long-term rejection of organ allografts. Computational biology provides a powerful tool to assess the large amount of complex data that is generated in longitudinal studies in this area. This manuscript outlines how our two groups are using mathematical modeling to analyze predictors of graft loss using both clinical and experimental data and how we plan to expand this approach to investigate specific mechanisms of chronic renal allograft injury.

  18. Novel stable isotope analyses demonstrate significant rates of glucose cycling in mouse pancreatic islets.

    Science.gov (United States)

    Wall, Martha L; Pound, Lynley D; Trenary, Irina; O'Brien, Richard M; Young, Jamey D

    2015-06-01

    A polymorphism located in the G6PC2 gene, which encodes an islet-specific glucose-6-phosphatase catalytic subunit, is the most important common determinant of variations in fasting blood glucose (FBG) levels in humans. Studies of G6pc2 knockout (KO) mice suggest that G6pc2 represents a negative regulator of basal glucose-stimulated insulin secretion (GSIS) that acts by hydrolyzing glucose-6-phosphate (G6P), thereby reducing glycolytic flux. However, this conclusion conflicts with the very low estimates for the rate of glucose cycling in pancreatic islets, as assessed using radioisotopes. We have reassessed the rate of glucose cycling in pancreatic islets using a novel stable isotope method. The data show much higher levels of glucose cycling than previously reported. In 5 mmol/L glucose, islets from C57BL/6J chow-fed mice cycled ∼16% of net glucose uptake. The cycling rate was further increased at 11 mmol/L glucose. Similar cycling rates were observed using islets from high fat-fed mice. Importantly, glucose cycling was abolished in G6pc2 KO mouse islets, confirming that G6pc2 opposes the action of the glucose sensor glucokinase by hydrolyzing G6P. The demonstration of high rates of glucose cycling in pancreatic islets explains why G6pc2 deletion enhances GSIS and why variants in G6PC2 affect FBG in humans. © 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

  19. Stem cell sources for clinical islet transplantation in type 1 diabetes: embryonic and adult stem cells.

    Science.gov (United States)

    Miszta-Lane, Helena; Mirbolooki, Mohammadreza; James Shapiro, A M; Lakey, Jonathan R T

    2006-01-01

    Lifelong immunosuppressive therapy and inadequate sources of transplantable islets have led the islet transplantation benefits to less than 0.5% of type 1 diabetics. Whereas the potential risk of infection by animal endogenous viruses limits the uses of islet xeno-transplantation, deriving islets from stem cells seems to be able to overcome the current problems of islet shortages and immune compatibility. Both embryonic (derived from the inner cell mass of blastocysts) and adult stem cells (derived from adult tissues) have shown controversial results in secreting insulin in vitro and normalizing hyperglycemia in vivo. ESCs research is thought to have much greater developmental potential than adult stem cells; however it is still in the basic research phase. Existing ESC lines are not believed to be identical or ideal for generating islets or beta-cells and additional ESC lines have to be established. Research with ESCs derived from humans is controversial because it requires the destruction of a human embryo and/or therapeutic cloning, which some believe is a slippery slope to reproductive cloning. On the other hand, adult stem cells are already in some degree specialized, recipients may receive their own stem cells. They are flexible but they have shown mixed degree of availability. Adult stem cells are not pluripotent. They may not exist for all organs. They are difficult to purify and they cannot be maintained well outside the body. In order to draw the future avenues in this field, existent discrepancies between the results need to be clarified. In this study, we will review the different aspects and challenges of using embryonic or adult stem cells in clinical islet transplantation for the treatment of type 1 diabetes.

  20. Insulin release from islets of Langerhans entrapped in a poly(N-isopropylacrylamide-co-acrylic acid) polymer gel.

    Science.gov (United States)

    Vernon, B; Kim, S W; Bae, Y H

    1999-01-01

    A copolymer of N-isopropylacrylamide (98 mol% in feed) and acrylic acid, poly(N-isopropylacrylamide-co-acrylic acid) (P(NIPAAm-co-AAc)), was prepared by free radical polymerization for development of a thermally reversible polymer to entrap islets of Langerhans for a refillable biohybrid artificial pancreas. A 5 wt% solution of the polymer in Hanks' balanced salt solution forms a gel at 37 degrees C that exhibits no syneresis. Diffusion of fluorescein isothiocyanate (FITC) dextrans having molecular weights of 4400 and 70000 were used to evaluate mass transport in the gel at 37 degrees C. Insulin secretion from islets in the polymer gel was also investigated in both static and dynamic systems. The polymer gel exhibited excellent diffusion of FITC dextran 4400 and FITC dextran 70000 with diffusion ratios, D/D0 (ratio of diffusion in the gel to diffusion in water), of 0.20+/-0.04 and 0.35+/-0.17, respectively. Human islets entrapped in the polymer gel showed prolonged insulin secretion in response to basal (5.5 mM) glucose concentration compared to free human islets. Rat islets showed prolonged insulin secretion in response to high (16.5 mM) glucose concentrations compared to free rat islets. Rat islets in the polymer gel maintained insulin secretion in response to the higher glucose concentration for over 26 days. Rat islets entrapped by the polymer also released higher quantities of insulin more rapidly in response to changes in concentrations of glucose and other stimulants than rat islets entrapped in an alginate control. These results suggest that this material would provide adequate diffusion for rapid insulin release in an application as a synthetic extracellular matrix for a biohybrid artificial pancreas.

  1. IDDM: an islet or an immune disease?

    Science.gov (United States)

    Boitard, C; Larger, E; Timsit, J; Sempe, P; Bach, J F

    1994-09-01

    Insulin-dependent diabetes develops as a consequence of the selective destruction of insulin-producing cells by an autoimmune reaction. However, the precise series of events which trigger anti-islet autoreactive T cells is still being investigated. Major issues will need to be raised before a comprehensive view of the anti-islet autoimmune reaction can be delineated. These include defining the primary site of activation of autoreactive lymphocytes and exploring hypotheses to explain the chronicity of the diabetes process. These issues all relate with the more general dilemma of the actual role of the islets of Langerhans in breaking self tolerance to beta-cell antigens. By studying non-obese diabetic mice deprived of beta cells following a single injection of a high dose of alloxan at 3 weeks of age, we recently obtained evidence that the activation of autoreactive T cells requires the presence of target islet cells in order to develop.

  2. Effects of mature Sertoli cells on allogeneic islets cocultured in vitro

    Institute of Scientific and Technical Information of China (English)

    Heli Xiang; Wujun Xue; Yan Teng; Xinshun Feng; Puxun Tian; Xiaoming Ding

    2006-01-01

    Objective: To set up a method for isolation and culture of mature Sertoli cells and to estimate their effects on allogeneic islets cocultured in vitro. Methods: Adult SD rat testicular Sertoli cells were prepared successfully by three-step enzyme digestion. Then they were cocultured respectively with allogeneic islets and activated Wistar rat splenocytes. 24-hour cumulative insulin release and glucose-stimulated insulin secretion test were performed to detect islet function between pure islets culture group and coculture group. Splenocyte proliferation activity was determined by MTT colorimetry assay to observe the inhibition effect of Sertoli cells in different densities. Result: Firstly, in pure islet culture group, the 24-hour cumulative insulin release was gradually decreased in 21-day culture time. Compared to day 3, this change was significant on day 7 (P < 0.05) and on day 10,14,21 (P < 0.01). In contrast, in coculture group, compared to day 3, the 24-hour cumulative insulin release was increased significantly on day 7 (P < 0.01 ), and then gradually decreased on day 10 and 14, but still higher than that of day 3. It was on day 21 that it began to decrease compared to day 3 (P < 0.05). During the culture time in vitro, the 24-hour cumulative insulin release of islet coculture group was significantly higher than that of pure islets culture group (P < 0.01). In the case of stimulation index(SI), there was a similar tendency as insulin release in the two groups. Secondly, mature Sertoli cells(1×106/mL)pretreated by 15 grays irradiation could decrease proliferation activity of activated splenocytes compared to that of control group (P < 0.01 ). This inhibition effect was dose-dependent. Conclusion: Mature Sertoli cells can improve the function and prolong the survival of islet cells cultured in vitro. They can also provide an immune protection to islet cells. The approach described above might be applicable to human islet transplantation as soon as

  3. Percutaneous transhepatic portal catheterization guided by ultrasound technology for islet transplantation in rhesus monkey

    Institute of Scientific and Technical Information of China (English)

    FengGao; Shao-DongAi; ShengLiu; Wen-BinZeng; WeiWang

    2012-01-01

    BACKGROUND: Pig islet xenotransplantation has the potential to overcome the shortage of donated human islets for islet cell transplantation in type 1 diabetes. Testing in non-human primate models is necessary before clinical application in humans. Intraportal islet transplantation in monkeys is usually performed by surgical infusion during laparotomy or laparoscopy. In this paper, we describe a new method of percutaneous transhepatic portal catheterization (PTPC) as an alternative to current methods of islet transplantation in rhesus monkeys. METHODS: We performed ultrasound-guided PTPC in five adult rhesus monkeys weighing 7-8 kg, with portal vein catheterization confirmed by digital subtraction angiography. We monitored for complications in the thoracic and abdominal cavity. To evaluate the safety of ultrasound-guided PTPC, we recorded the changes in portal pressure throughout the microbead transplantation procedure. RESULTS:  Ultrasound-guided PTPC and infusion of 16 000 microbeads/kg body weight into the portal vein was successful in all five monkeys. Differences in the hepatobiliary anatomy of rhesus monkeys compared to humans led to a higher initial complication rate. The first monkey died of abdominal hemorrhage 10 hours post-transplantation. The second suffered from a mild pneumothorax but recovered fully after taking only conservative measures. After gaining experience with the first two monkeys, we decreased both the hepatic puncture time and the number of puncture attempts required, with the remaining three monkeys experiencing no complications. Portal pressures initially increased proportional to the number of transplanted microbeads but returned to pre-infusion levels at 30 minutes post-transplantation. The changes in portal pressures occurring during the procedure were not significantly different. CONCLUSIONS: Ultrasound-guided PTPC is an effective, convenient, and minimally invasive method suitable for use in non-human primate models of

  4. Zebularine induces long-term survival of pancreatic islet allotransplants in streptozotocin treated diabetic rats.

    Directory of Open Access Journals (Sweden)

    Henrietta Nittby

    Full Text Available BACKGROUND: Coping with the immune rejection of allotransplants or autologous cells in patients with an active sensitization towards their autoantigens and autoimmunity presently necessitates life-long immune suppressive therapy acting on the immune system as a whole, which makes the patients vulnerable to infections and increases their risk of developing cancer. New technologies to induce antigen selective long-lasting immunosuppression or immune tolerance are therefore much needed. METHODOLOGY/PRINCIPAL FINDINGS: The DNA demethylating agent Zebularine, previously demonstrated to induce expression of the genes for the immunosuppressive enzymes indolamine-2,3-deoxygenase-1 (IDO1 and kynureninase of the kynurenine pathway, is tested for capacity to suppress rejection of allotransplants. Allogeneic pancreatic islets from Lewis rats were transplanted under the kidney capsule of Fischer rats previously made diabetic by a streptozotocin injection (40 mg/kg. One group was treated with Zebularine (225 mg/kg daily for 14 days from day 6 or 8 after transplantation, and a control group received no further treatment. Survival of the transplants was monitored by blood sugar measurements. Rats, normoglycemic for 90 days after allografting, were subjected to transplant removal by nephrectomy to confirm whether normoglycemia was indeed due to a surviving insulin producing transplant, or alternatively was a result of recovery of pancreatic insulin production in some toxin-treated rats. Of 9 Zebularine treated rats, 4 were still normoglycemic after 90 days and became hyperglycemic after nephrectomy. The mean length of normoglycemia in the Zebularine group was 67±8 days as compared to 14±3 days in 9 controls. Seven rats (2 controls and 5 Zebularine treated were normoglycemic at 90 days due to pancreatic recovery as demonstrated by failure of nephrectomy to induce hyperglycemia. CONCLUSIONS/SIGNIFICANCE: Zebularine treatment in vivo induces a long

  5. Justifying clinical trials for porcine islet xenotransplantation.

    Science.gov (United States)

    Ellis, Cara E; Korbutt, Gregory S

    2015-01-01

    The development of the Edmonton Protocol encouraged a great deal of optimism that a cell-based cure for type I diabetes could be achieved. However, donor organ shortages prevent islet transplantation from being a widespread solution as the supply cannot possibly equal the demand. Porcine islet xenotransplantation has the potential to address these shortages, and recent preclinical and clinical trials show promising scientific support. Consequently, it is important to consider whether the current science meets the ethical requirements for moving toward clinical trials. Despite the potential risks and the scientific unknowns that remain to be investigated, there is optimism regarding the xenotransplantation of some types of tissue, and enough evidence has been gathered to ethically justify clinical trials for the most safe and advanced area of research, porcine islet transplantation. Researchers must make a concerted effort to maintain a positive image for xenotransplantation, as a few well-publicized failed trials could irrevocably damage public perception of xenotransplantation. Because all of society carries the burden of risk, it is important that the public be involved in the decision to proceed. As new information from preclinical and clinical trials develops, policy decisions should be frequently updated. If at any point evidence shows that islet xenotransplantation is unsafe, then clinical trials will no longer be justified and they should be halted. However, as of now, the expected benefit of an unlimited supply of islets, combined with adequate informed consent, justifies clinical trials for islet xenotransplantation.

  6. Cytokines cause functional and structural damage to isolated islets of Langerhans

    DEFF Research Database (Denmark)

    Mandrup-Poulsen, T; Bendtzen, K; Bendixen, G

    1985-01-01

    -dependent diabetes mellitus, isolated human or rat islets of Langerhans were incubated for 7 days with cytokine-rich, cell-free supernatants of blood mononuclear cells from healthy human donors stimulated with or without purified protein derivative of tuberculin or phytohaemagglutinin. Glucose stimulated insulin......Cytokines are soluble, antigen non-specific, non-immunoglobulin mediators produced and secreted by blood mononuclear cells interacting in the cellular immune-response. To test the possibility that cytokines participate in the autoimmune destruction of the pancreatic beta-cells leading to insulin......-release, and contents of insulin and glucagon in islets incubated with cytokine-rich supernatants were markedly reduced. This impairment of islet function was due to a cytotoxic effect of cytokine-rich supernatants as judged by disintegration of normal light-microscopic morphology....

  7. Effect of two cleaning processes for bone allografts on gentamicin impregnation and in vitro antibiotic release.

    Science.gov (United States)

    Coraça-Huber, D C; Hausdorfer, J; Fille, M; Steidl, M; Nogler, M

    2013-06-01

    Bone allografts are a useful and sometimes indispensable tool for the surgeon to repair bone defects. Microbial contamination is a major reason for discarding allografts from bone banks. To improve the number of safe allografts, we suggest chemical cleaning of the grafts followed by antibiotic impregnation. Comparison of two chemical cleaning processes for bone allografts aiming for antibiotic impregnation and consequently delivery rates in vitro. Bone chips of 5-10 mm were prepared from human femoral heads. Two cleaning methods (cleaning A and cleaning B) based on solutions containing hydrogen peroxide, paracetic acid, ethanol and biological detergent were carried out and compared. After the cleaning processes, the bone chips were impregnated with gentamicin. Bacillus subtilis bioassay was used to determine the gentamicin release after intervals of 1-7 days. Differences were compared with non-parametric Mann-Whitney U tests. The zones of inhibition obtained from the bone grafts cleaned with both cleaning processes were similar between the groups. The concentration of the released antibiotic was decreasing gradually over time, following a similar pattern for both groups. The cleaning procedure A as well as the cleaning procedure B for bone allografts allowed the impregnation with gentamicin powder in the same concentrations in both groups. The delivery of gentamicin was similar for both groups. Both cleaning procedures were easy to be carried out, making them suitable for routine use at the bone banks.

  8. Histomorphology of the bottlenose dolphin (Tursiops truncatus) pancreas and association of increasing islet β-cell size with chronic hypercholesterolemia.

    Science.gov (United States)

    Colegrove, Kathleen M; Venn-Watson, Stephanie

    2015-04-01

    Bottlenose dolphins (Tursiops truncatus) can develop metabolic states mimicking prediabetes, including hyperinsulinemia, hyperlipidemia, elevated glucose, and fatty liver disease. Little is known, however, about dolphin pancreatic histomorphology. Distribution and area of islets, α, β, and δ cells were evaluated in pancreatic tissue from 22 dolphins (mean age 25.7years, range 0-51). Associations of these measurements were evaluated by sex, age, percent high glucose and lipids during the last year of life, and presence or absence of fatty liver disease and islet cell vacuolation. The most common pancreatic lesions identified were exocrine pancreas fibrosis (63.6%) and mild islet cell vacuolation (47.4%); there was no evidence of insulitis or amyloid deposition, changes commonly associated with type 2 diabetes. Dolphin islet architecture appears to be most similar to the pig, where α and β cells are localized to the central or periphery of the islet, respectively, or are well dispersed throughout the islet. Unlike pigs, large islets (greater than 10,000μm(2)) were common in dolphins, similar to that found in humans. A positive linear association was identified between dolphin age and islet area average, supporting a compensatory response similar to other species. The strongest finding in this study was a positive linear association between islet size, specifically β-cells, and percent blood samples with high cholesterol (greater than 280mg/dl, R(2)=0.57). This study is the most comprehensive assessment of the dolphin pancreas to date and may help direct future studies, including associations between chronic hypercholesterolemia and β-cell size.

  9. In vivo islet protection by a nuclear import inhibitor in a mouse model of type 1 diabetes.

    Directory of Open Access Journals (Sweden)

    Daniel J Moore

    Full Text Available BACKGROUND: Insulin-dependent Type 1 diabetes (T1D is a devastating autoimmune disease that destroys beta cells within the pancreatic islets and afflicts over 10 million people worldwide. These patients face life-long risks for blindness, cardiovascular and renal diseases, and complications of insulin treatment. New therapies that protect islets from autoimmune destruction and allow continuing insulin production are needed. Increasing evidence regarding the pathomechanism of T1D indicates that islets are destroyed by the relentless attack by autoreactive immune cells evolving from an aberrant action of the innate, in addition to adaptive, immune system that produces islet-toxic cytokines, chemokines, and other effectors of islet inflammation. We tested the hypothesis that targeting nuclear import of stress-responsive transcription factors evoked by agonist-stimulated innate and adaptive immunity receptors would protect islets from autoimmune destruction. PRINCIPAL FINDINGS: Here we show that a first-in-class inhibitor of nuclear import, cSN50 peptide, affords in vivo islet protection following a 2-day course of intense treatment in NOD mice, which resulted in a diabetes-free state for one year without apparent toxicity. This nuclear import inhibitor precipitously reduces the accumulation of islet-destructive autoreactive lymphocytes while enhancing activation-induced cell death of T and B lymphocytes derived from autoimmune diabetes-prone, non-obese diabetic (NOD mice that develop T1D. Moreover, in this widely used model of human T1D we noted attenuation of pro-inflammatory cytokine and chemokine production in immune cells. CONCLUSIONS: These results indicate that a novel form of immunotherapy that targets nuclear import can arrest inflammation-driven destruction of insulin-producing beta cells at the site of autoimmune attack within pancreatic islets during the progression of T1D.

  10. Activin B regulates islet composition and islet mass but not whole body glucose homeostasis or insulin sensitivity

    Science.gov (United States)

    Bonomi, Lara; Brown, Melissa; Ungerleider, Nathan; Muse, Meghan; Matzuk, Martin M.

    2012-01-01

    Based on the phenotype of the activin-like kinase-7 (ALK7)-null mouse, activins A and B have been proposed to play distinct roles in regulating pancreatic islet function and glucose homeostasis, with activin A acting to enhance islet function and insulin release while activin B antagonizes these actions. We therefore hypothesized that islets from activin B-null (BBKO) mice would have enhanced glucose-stimulated insulin secretion. In addition, we hypothesized that this enhanced islet function would translate into increased whole body glucose tolerance. We tested these hypotheses by analyzing glucose homeostasis, insulin secretion, and islet function in BBKO mice. No differences were observed in fasting glucose or insulin levels, glucose tolerance, or insulin sensitivity compared with weight-matched young or older males. Similarly, there were no significant differences in insulin secretion comparing islets from WT or BBKO males at either age. However, BBKO islets were more sensitive to activin A, myostatin (MSTN), and follistatin (FST) treatments, so that activin A and FST inhibited and MSTN enhanced glucose stimulated insulin secretion. While mean islet area and the distribution of islet areas were not different between the genotypes, islet mass, islet number, and the proportion of α-cells/islet were significantly reduced in BBKO islets. These results indicate that activin B does not antagonize activin A to influence whole body glucose homeostasis or β-cell function but does influence islet mass and proportion of α-cells/islet. Therefore, loss of activin B signaling alone does not account for the ALK7-null phenotype, but activin B may have important roles in modulating islet mass, islet number, and the cellular composition of islets. PMID:22739106

  11. Successful treatment of verruca vulgaris with Thuja occidentalis in a renal allograft recipient

    Directory of Open Access Journals (Sweden)

    R Joseph

    2013-01-01

    Full Text Available Human papillomavirus-driven verruca vulgaris infection is common in solid organ transplant recipients and increases the risk for squamous cell carcinoma. The available treatment modalities have limited response. We report a renal allograft recipient who presented with multiple warts not responding to cryotherapy and radiosurgery with one turning malignant, needing amputation of the finger. An extract from Thuja occidentalis (White cedar tree cured the resistant warts on the other fingers, leaving only superficial scars and without affecting allograft function. We have reviewed the pharmacological and clinical properties of T. occidentalis.

  12. Total lymphoid irradiation assessed for possible enhancement of immunosuppression in hyperimmunized dogs receiving renal allografts

    Energy Technology Data Exchange (ETDEWEB)

    Sonoda, Kazuhiko (Yamato Seiwa Hospital, Kanagawa (Japan)); Rapaport, F.T.

    1992-12-01

    With performed antibodies to human leukocyte antigens (HLA) appearing in an increasing number of patients today, hyperimmunization constitutes a major problem in clinical transplantation. In adult beagle dogs hyperimmunized with skin allografts and buffy coat injection, we performed renal allograft transplantation to assess the efficacy of total lymphoid irradiation (TLI) employed as a preoperative measure in combination with cyclosporine (CyA) and methyl-prednisolone (MPL) in effecting immunosuppression. The mean survival period were 6.5 days in dogs withheld preliminary treatment, 9.0 days in the dogs receiving CyA and MPL, 26.7 days in those administered one-stage TLI, and 68 days (terminated by euthanasia) of the dogs given two-stage TLI. TLI administered two stages is considered an effective method of enhancing immunosuppression sufficiently to enable the attenuation of adverse reaction to renal allograft in hyperimmunized recipients. (author).

  13. Scintigraphy of lower extremity cadaveric bone allografts in osteosarcoma patients.

    Science.gov (United States)

    Bar-Sever, Z; Connolly, L P; Gebhardt, M C; Treves, S T

    1997-08-01

    To describe scintigraphic characteristics of bone allografts used in limb salvage reconstruction after resection of lower extremity osteosarcoma. The authors reviewed 85 skeletal scintigrams of 20 pediatric patients followed up for 0.5-5.7 years after resection of lower extremity osteosarcoma and allograft reconstruction. Uptake in the allograft and adjacent host tissues was assessed visually. Lack of tracer uptake in the allografts was seen in 99% of the studies and a faint rim of tracer localization outlining the allograft's periphery was seen in 95% of the studies. Increased uptake was noted at the allograft-host bone junction in 78% of the studies. Uptake was increased in the joint surfaces of native bones articulating with allografts (97% of studies), including the patella (93% of studies) when the knee was involved. These findings were stabilized as time passed. Cadaveric bone allografts have a characteristic scintigraphic appearance in this selected patient group that reflects the physiology of their incorporation process.

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

    Directory of Open Access Journals (Sweden)

    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.

  15. Long-term biocompatibility, chemistry, and function of microencapsulated pancreatic islets.

    Science.gov (United States)

    de Vos, Paul; van Hoogmoed, Chris G; van Zanten, Jacoba; Netter, S; Strubbe, Jan H; Busscher, Henk J

    2003-01-01

    Transplantation of encapsulated living cells is a promising approach for the treatment of a wide variety of diseases. Large-scale application of the technique, however, is hampered by insufficient biocompatibility of the capsules. In the present study, we have implemented new as well as previously reported technologies to test biocompatibility issues of immunoisolating microcapsules on the long term (i.e. 2 years) instead of usually reported short time periods. When transplanted empty, the capsules proved to be highly biocompatible not only for short periods (i.e. 1 month) but also on the long term as evidenced by the absence of any significant biological response up to 2 years after implantation in rats. The immunoprotective properties of the capsules were confirmed by prolonged survival of encapsulated islet allografts up to 200 days. The surface of the applied capsule was analyzed and provides new insight in the chemical structure of true biocompatible and immunoprotective capsules applicable for transplantation of encapsulated islets in type I diabetes.

  16. Genetics and Epigenetics of Chronic Allograft Dysfunction in Kidney Transplants.

    Science.gov (United States)

    Zununi Vahed, Sepideh; Samadi, Nasser; Mostafidi, Elmira; Ardalan, Mohammad Reza; Omidi, Yadollah

    2016-01-01

    Chronic allograft dysfunction is the most common cause of allograft lost. Chronic allograft dysfunction happens as a result of complex interactions at the molecular and cellular levels. Genetic and environmental factors both influence the evolution and progression of the chronic allograft dysfunction. Epigenetic modification could be considered as a therapeutically modifiable element to pause the fibrosis process through novel strategies. In this review, the PubMed database was searched for English-language articles on these new areas.

  17. Re-exposure to beta cell autoantigens in pancreatic allograft recipients with preexisting beta cell autoantibodies.

    Science.gov (United States)

    Mujtaba, Muhammad Ahmad; Fridell, Jonathan; Book, Benita; Faiz, Sara; Sharfuddin, Asif; Wiebke, Eric; Rigby, Mark; Taber, Tim

    2015-11-01

    Re-exposure to beta cell autoantigens and its relevance in the presence of donor-specific antibodies (DSA) in pancreatic allograft recipients is not well known. Thirty-three patients requiring a pancreas transplant were enrolled in an IRB approved study. They underwent prospective monitoring for DSA and beta cell autoantibody (BCAA) levels to GAD65, insulinoma-associated antigen 2 (IA-2), insulin (micro-IAA [mIAA]), and islet-specific zinc transporter isoform-8 (ZnT8). Twenty-five (75.7%) had pre-transplant BCAA. Twenty had a single antibody (mIAA n = 15, GAD65 n = 5); five had two or more BCAA (GAD65 + mIAA n = 2, GAD65 + mIAA+IA-2 n = 2, GA65 + mIAA+IA-2 + ZnT8 = 1). No changes in GAD65 (p > 0.29), IA-2 (>0.16), and ZnT8 (p > 0.07) were observed between pre-transplant and post-transplant at 6 or 12 months. A decrease in mIAA from pre- to post-6 months (p BCAA was observed at one yr. Seven (21.0%) developed de novo DSA. The incidence of DSA was 24% in patients with BCAA vs. 25% in patients without BCAA (p = 0.69). Pancreatic allograft function of patients with vs. without BCAA, and with and without BCAA + DSA was comparable until last follow-up (three yr). Re-exposure to beta cell autoantigens by pancreas transplant may not lead to increased levels or development of new BCAA or pancreatic allograft dysfunction.

  18. Immediate retransplantation for pancreas allograft thrombosis.

    Science.gov (United States)

    Hollinger, E F; Powelson, J A; Mangus, R S; Kazimi, M M; Taber, T E; Goble, M L; Fridell, J A

    2009-04-01

    Early pancreas allograft failure most commonly results from thrombosis and requires immediate allograft pancreatectomy. Optimal timing for retransplantation remains undefined. Immediate retransplantation facilitates reuse of the same anatomic site before extensive adhesions have formed. Some studies suggest that early retransplantation is associated with a higher incidence of graft loss. This study is a retrospective review of immediate pancreas retransplants performed at a single center. All cases of pancreas allograft loss within 2 weeks were examined. Of 228 pancreas transplants, 12 grafts were lost within 2 weeks of surgery. Eleven of these underwent allograft pancreatectomy for thrombosis. One suffered anoxic brain injury and was not a retransplantation candidate, one was retransplanted at 3.5 months and nine patients underwent retransplantation 1-16 days following the original transplant. Of the nine early retransplants, one pancreas was lost to heparin-induced thrombocytopenia, one recipient died with function at 2.9 years and the other grafts continue to function at 76-1137 days (mean 572 days). One-year graft survival for early retransplantation was 89% compared to 91% for all pancreas transplants at our center. Immediate retransplantation following pancreatic graft thrombosis restores durable allograft function with outcomes comparable to first-time pancreas transplantation.

  19. Radionuclide surveillance of the allografted pancreas

    Energy Technology Data Exchange (ETDEWEB)

    George, E.A.; Salimi, Z.; Carney, K.; Castaneda, M.; Garvin, P.J.

    1988-04-01

    To determine the value of scintigraphy to detect posttransplantation complications of the allografted pancreas, we retrospectively reviewed 209 scintigrams obtained with /sup 99m/Tc-sulfur colloid (/sup 99m/Tc-SC) and /sup 99m/Tc-glucoheptonate (/sup 99m/Tc-GH). The scintigraphic studies were performed in 37 recipients of simultaneous renal and pancreatic allografts harvested from the same donor. /sup 99m/Tc-SC was used as an indicator of thrombotic vasculitis; pancreatic perfusion and blood-pool parameters were monitored with /sup 99m/Tc-GH. In 11 of the 37 recipients, scintigraphic abnormalities suggested posttransplantation infarction. Recurrent episodes of acute rejection of the pancreatic allograft, which always coincided with acute rejection of the renal allograft, were monitored in 24 recipients. Rejection-induced ischemic pancreatitis was suggested in 12 of the 24 recipients and persisted in 10 recipients for several weeks after improvement of renal allograft rejection. Pancreatic atrophy was suggested scintigraphically in 16 of the 24 recipients with recurrent episodes of rejection. Spontaneous pancreatic-duct obstruction and obstructive pancreatitis were associated with a scintigraphic pattern similar to that of rejection-induced ischemic pancreatitis. We concluded that the specific radionuclides used in this series are useful for the surveillance and assessment of posttransplantation pancreatic infarction, acute rejection, pancreatitis, and atrophy

  20. Assessment of Toxicological Perturbations and Variants of Pancreatic Islet Development in the Zebrafish Model

    Directory of Open Access Journals (Sweden)

    Karilyn E. Sant

    2016-09-01

    Full Text Available The pancreatic islets, largely comprised of insulin-producing beta cells, play a critical role in endocrine signaling and glucose homeostasis. Because they have low levels of antioxidant defenses and a high perfusion rate, the endocrine islets may be a highly susceptible target tissue of chemical exposures. However, this endpoint, as well as the integrity of the surrounding exocrine pancreas, is often overlooked in studies of developmental toxicology. Disruption of development by toxicants can alter cell fate and migration, resulting in structural alterations that are difficult to detect in mammalian embryo systems, but that are easily observed in the zebrafish embryo model (Danio rerio. Using endogenously expressed fluorescent protein markers for developing zebrafish beta cells and exocrine pancreas tissue, we documented differences in islet area and incidence rates of islet morphological variants in zebrafish embryos between 48 and 96 h post fertilization (hpf, raised under control conditions commonly used in embryotoxicity assays. We identified critical windows for chemical exposures during which increased incidences of endocrine pancreas abnormalities were observed following exposure to cyclopamine (2–12 hpf, Mono-2-ethylhexyl phthalate (MEHP (3–48 hpf, and Perfluorooctanesulfonic acid (PFOS (3–48 hpf. Both islet area and length of the exocrine pancreas were sensitive to oxidative stress from exposure to the oxidant tert-butyl hydroperoxide during a highly proliferative critical window (72 hpf. Finally, pancreatic dysmorphogenesis following developmental exposures is discussed with respect to human disease.

  1. Protein phosphatase 1 (PP-1)-dependent inhibition of insulin secretion by leptin in INS-1 pancreatic β-cells and human pancreatic islets.

    Science.gov (United States)

    Kuehnen, Peter; Laubner, Katharina; Raile, Klemens; Schöfl, Christof; Jakob, Franz; Pilz, Ingo; Päth, Günter; Seufert, Jochen

    2011-05-01

    Leptin inhibits insulin secretion from pancreatic β-cells, and in turn, insulin stimulates leptin biosynthesis and secretion from adipose tissue. Dysfunction of this adipoinsular feedback loop has been proposed to be involved in the development of hyperinsulinemia and type 2 diabetes mellitus. At the molecular level, leptin acts through various pathways, which in combination confer inhibitory effects on insulin biosynthesis and secretion. The aim of this study was to identify molecular mechanisms of leptin action on insulin secretion in pancreatic β-cells. To identify novel leptin-regulated genes, we performed subtraction PCR in INS-1 β-cells. Regulated expression of identified genes was confirmed by RT-PCR and Northern and Western blotting. Furthermore, functional impact on β-cell function was characterized by insulin-secretion assays, intracellular Ca²(+) concentration measurements, and enzyme activity assays. PP-1α, the catalytic subunit of protein phosphatase 1 (PP-1), was identified as a novel gene down-regulated by leptin in INS-1 pancreatic β-cells. Expression of PP-1α was verified in human pancreatic sections. PP-1α mRNA and protein expression is down-regulated by leptin, which culminates in reduction of PP-1 enzyme activity in β-cells. In addition, glucose-induced insulin secretion was inhibited by nuclear inhibitor of PP-1 and calyculin A, which was in part mediated by a reduction of PP-1-dependent calcium influx into INS-1 β-cells. These results identify a novel molecular pathway by which leptin confers inhibitory action on insulin secretion, and impaired PP-1 inhibition by leptin may be involved in dysfunction of the adipoinsular axis during the development of hyperinsulinemia and type 2 diabetes mellitus.

  2. Active haemorrhage of a renal allograft detected on portable ultrasound

    OpenAIRE

    Ricketts, James; Pang, Chun Lap; Dissanayake, Prageeth; Hutchinson, Rachel; Gutteridge, Catherine

    2013-01-01

    Function of a renal allograft relies on the integrity of its vascular anatomy. Renal biochemistry, ultrasound and percutaneous biopsy are used in combination to determine allograft function. Biopsy is not without risk, and in this case study we demonstrate a rare but a potentially life-threatening complication of renal allograft biopsy.

  3. Calprotectin - A novel noninvasive marker for intestinal allograft monitoring

    NARCIS (Netherlands)

    Sudan, Debra; Vargas, Luciano; Sun, Yimin; Bok, Lisette; Dijkstra, Gerard; Langnas, Alan

    2007-01-01

    Objective: To identify a noninvasive screening test for intestinal allograft monitoring. Summary Background Data: Intestinal allograft rejection is difficult to distinguish from other causes of diarrhea and can rapidly lead to severe exfoliation or death. Protocol biopsies are standard for allograft

  4. Islet Amyloid Polypeptide: Structure, Function, and Pathophysiology

    Directory of Open Access Journals (Sweden)

    Rehana Akter

    2016-01-01

    Full Text Available The hormone islet amyloid polypeptide (IAPP, or amylin plays a role in glucose homeostasis but aggregates to form islet amyloid in type-2 diabetes. Islet amyloid formation contributes to β-cell dysfunction and death in the disease and to the failure of islet transplants. Recent work suggests a role for IAPP aggregation in cardiovascular complications of type-2 diabetes and hints at a possible role in type-1 diabetes. The mechanisms of IAPP amyloid formation in vivo or in vitro are not understood and the mechanisms of IAPP induced β-cell death are not fully defined. Activation of the inflammasome, defects in autophagy, ER stress, generation of reactive oxygen species, membrane disruption, and receptor mediated mechanisms have all been proposed to play a role. Open questions in the field include the relative importance of the various mechanisms of β-cell death, the relevance of reductionist biophysical studies to the situation in vivo, the molecular mechanism of amyloid formation in vitro and in vivo, the factors which trigger amyloid formation in type-2 diabetes, the potential role of IAPP in type-1 diabetes, the development of clinically relevant inhibitors of islet amyloidosis toxicity, and the design of soluble, bioactive variants of IAPP for use as adjuncts to insulin therapy.

  5. Donor-specific Regulatory T Cells Acquired from Tolerant Mice Bearing Cardiac allograft Promote Mixed Chimerism and Prolong Intestinal Allograft Survival

    Directory of Open Access Journals (Sweden)

    Xiaofei Shen

    2016-11-01

    Full Text Available The induction of donor-specific transplant tolerance has always been a central problem for small bowel transplantation, which is thought to be the best therapy for end-stage bowel failure. With the development of new tolerance-inducing strategies, mixed chimerism induced by co-stimulation blockade has become most potent for tolerance of allografts such as skin, kidney and heart. However, a lack of clinically available co-stimulation blockers has hindered efficient application in humans. Furthermore, unlike those for other types of solid organ transplantation, strategies to induce robust mixed chimerism for intestinal allografts have not been fully developed. To improve current mixed chimerism induction protocols for future clinical application, we developed a new protocol using donor-specific regulatory T (Treg cells from mice with heart allograft tolerance, clinically available immunosuppressive drugs, and low doses of irradiation. Our results demonstrated that donor-specific Treg cells acquired from tolerant mice after in vitro expansion generate stable chimerism and lead to acceptance of intestinal allograft. Increased intragraft Treg cells and clonal deletion both contribute to the development of small bowel transplantation tolerance.

  6. Chronic Kidney Isograft and Allograft Rejection

    Institute of Scientific and Technical Information of China (English)

    严群; 张鹏; 杨传永

    2002-01-01

    Summary: In this study antigen-independent factor in the pathogenesis of chronic rejection of organ transplants was examined. Kidney isografts and allografts were transplanted orthotopically into bilaterally nephroectomized rat recipients and studied functionally, morphologically and immunohistologically, at serial intervals up to 52 weeks after transplantation. Allograft recipients developed progressive proteinuria after 12 weeks, with gradual renal failure ultimately leading to death. At the same time, morphological changes, including progressive arteriosclerosis and glomerulosclerosis, tubular atrophy and interstitial fibrosis, developed. Immunohistologically, macrophages infiltrated glomeruli during this period and cytokines became unregulated. Our resuits showed that antigen-independent functional and morphological changes occurred in long-term kidney isografts and mimicked those appearing much earlier in allografts that reject chronically.Initial injury and extent of functioning renal mass is suggested to be important factor for such late changes.

  7. Urinary calprotectin and posttransplant renal allograft injury

    DEFF Research Database (Denmark)

    Tepel, Martin; Borst, Christoffer; Bistrup, Claus

    2014-01-01

    OBJECTIVE: Current methods do not predict the acute renal allograft injury immediately after kidney transplantation. We evaluated the diagnostic performance of urinary calprotectin for predicting immediate posttransplant allograft injury. METHODS: In a multicenter, prospective-cohort study of 144...... incipient renal transplant recipients, we postoperatively measured urinary calprotectin using an enzyme-linked immunosorbent assay and estimated glomerular filtration rate (eGFR) after 4 weeks, 6 months, and 12 months. RESULTS: We observed a significant inverse association of urinary calprotectin...... regression showed that higher urinary calprotectin concentrations and older donor age predicted lower eGFR four weeks, 6 months, and 12 months after transplantation. CONCLUSIONS: Urinary calprotectin is an early, noninvasive predictor of immediate renal allograft injury after kidney transplantation....

  8. Risk factors of cardiac allograft vasculopathy.

    Science.gov (United States)

    Szyguła-Jurkiewicz, Bożena; Szczurek, Wioletta; Gąsior, Mariusz; Zembala, Marian

    2015-12-01

    Despite advances in prevention and treatment of heart transplant rejection, development of cardiac allograft vasculopathy (CAV) remains the leading factor limiting long-term survival of the graft. Cardiac allograft vasculopathy etiopathogenesis is not fully understood, but a significant role is attributed to endothelial cell damage, caused by immunological and non-immunological mechanisms. Immunological factors include the differences between the recipient's and the donor's HLA systems, the presence of alloreactive antibodies and episodes of acute rejection. Among the non-immunological factors the most important are the age of the donor, ischemia-reperfusion injury and cytomegalovirus infection. The classical cardiovascular risk factors (diabetes, hypertension, obesity and hyperlipidemia) are also important. This study presents an up-to-date overview of current knowledge on the vasculopathy etiopathogenesis and the role played by endothelium and inflammatory processes in CAV, and it also investigates the factors which may serve as risk markers of cardiac allograft vasculopathy.

  9. Mesenchymal stem cells as feeder cells for pancreatic islet transplants.

    OpenAIRE

    2010-01-01

    Allogeneic islet transplantation serves as a source of insulin-secreting beta-cells for the maintenance of normal glucose levels and treatment of diabetes. However, limited availability of islets, high rates of islet graft failure, and the need for life-long non-specific immunosuppressive therapy are major obstacles to the widespread application of this therapeutic approach. To overcome these problems, pancreatic islet transplantation was recently suggested as a potential target of the "thera...

  10. MicroRNA-10b downregulation mediates acute rejection of renal allografts by derepressing BCL2L11

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xiaoyou [Department of Organ Transplantation, Zhujiang Hospital, Guangzhou 510282 (China); Dong, Changgui [Institute of Molecular Ecology and Evolution, East China Normal University, Shanghai 200062 (China); Jiang, Zhengyao [Department of Organ Transplantation, Zhujiang Hospital, Guangzhou 510282 (China); Wu, William K.K. [Department of Anaesthesia and Intensive Care, The Chinese University of Hong Kong, Shatin, NT, Hong Kong (China); State Key Laboratory of Digestive Diseases, LKS Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong (China); Chan, Matthew T.V. [Department of Anaesthesia and Intensive Care, The Chinese University of Hong Kong, Shatin, NT, Hong Kong (China); Zhang, Jie [Department of Organ Transplantation, Zhujiang Hospital, Guangzhou 510282 (China); Li, Haibin; Qin, Ke [Guangxi Key Laboratory for Transplantation Medicine Department of Organ Transplantation in Guangzhou Military Region, Institute of Transplant Medicine, 303 Hospital of People' s Liberation Army, Nanning, Guangxi 530021 (China); Sun, Xuyong, E-mail: sunxuyong0528@163.com [Guangxi Key Laboratory for Transplantation Medicine Department of Organ Transplantation in Guangzhou Military Region, Institute of Transplant Medicine, 303 Hospital of People' s Liberation Army, Nanning, Guangxi 530021 (China)

    2015-04-10

    Kidney transplantation is the major therapeutic option for end-stage kidney diseases. However, acute rejection could cause allograft loss in some of these patients. Emerging evidence supports that microRNA (miRNA) dysregulation is implicated in acute allograft rejection. In this study, we used next-generation sequencing to profile miRNA expression in normal and acutely rejected kidney allografts. Among 75 identified dysregulated miRNAs, miR-10b was the most significantly downregulated miRNAs in rejected allografts. Transfecting miR-10b inhibitor into human renal glomerular endothelial cells recapitulated key features of acute allograft rejection, including endothelial cell apoptosis, release of pro-inflammatory cytokines (interleukin-6, tumor necrosis factor α, interferon-γ, and chemokine (C–C motif) ligand 2) and chemotaxis of macrophages whereas transfection of miR-10b mimics had opposite effects. Downregulation of miR-10b directly derepressed the expression of BCL2L11 (an apoptosis inducer) as revealed by luciferase reporter assay. Taken together, miR-10b downregulation mediates many aspects of disease pathogenicity of acute kidney allograft rejection. Restoring miR-10b expression in glomerular endothelial cells could be a novel therapeutic approach to reduce acute renal allograft loss. - Highlights: • miR-10b was the most downregulated microRNAs in acutely rejected renal allografts. • miR-10b downregulation triggered glomerular endothelial cell apoptosis. • miR-10b downregulation induced release of pro-inflammatory cytokines. • miR-10b downregulation derepressed its pro-apoptotic target BCL2L11.

  11. 小鼠异体Sertoli细胞诱导胰岛移植免疫耐受的作用%Iso-Sertoli cell in mice induced immunotolerance in islet transplantation

    Institute of Scientific and Technical Information of China (English)

    郑宗珩; 朱晓峰; 马毅

    2009-01-01

    目的 观察小鼠Sertoli细胞是否能在异体内起到诱导局部免疫耐受、保护共移植异体胰岛的作用.方法 以糖尿病C57小鼠作移植受体,随机分4组,每组6只;以正常BALB/C小鼠为胰岛供体,正常C57小鼠和正常BALB/C小鼠各作为Serloli细胞供体.A组:单纯移植异体胰岛;B组:移植来源于C57小鼠的Sertoli细胞+BALB/C小鼠来源的胰岛;C组:移植均来源于BALB/C小鼠的Sertoli细胞及胰岛;D组:假手术组.监测各组移植受体的血糖尿糖变化,观察移植物的存活时间.结果 A组移植物平均存活时间为(6.50±2.35)d;B组为(55.67±4.84)d;C组为(51.33±5.05)d;D组未观察到血糖正常.B组及C组的移植方式均可逆转糖尿病小鼠的高血糖状态,移植物存活期均较A组有明显延长,其差异有统计学意义(P0.05).结论 同种异体来源的睾丸Sertoli细胞在异体内可起到诱导局部免疫耐受的效果,对共移植同种异体胰岛起到保护作用,其效果与自体睾丸Sertoli细胞相当.%Objective To study whether the sertoli cell allograft can achieve the immunotolerance and protect the co-transplant islet allograft on the heterotopic situation. Methods The diabetic C57 mice were used as recipients, and healthy BALB/C mice as islet donors,respectively. Healthy BALB/C and C57 mice were used as testis sertoli cell donors. The recipients were randomly divided into 4 groups,6 mice in each group : group A: only transplant with islet allograft;group B: co-transplant with islet allograft and serto-li isograft;group C:co-transplant with islet allograft and sertoli allograft;group D:sham-operated group. The blood and urine glucose levels in the models, and the survival time of the graft were observed. Results The mean survive time of graft in groups A, B, and C was (6.50±2.35 ), (55.67±4.84), and (51.33± 5.05 ) days respectively. In group D, blood glucose level was abnormal. The hyperglycemia of the diabetic C57 mice could be reversed by the

  12. Renal adenocarcinoma, hepatocellular carcinoma, and pancreatic islet cell carcinoma in a binturong (Arctictis binturong).

    Science.gov (United States)

    Klaphake, Eric; Shoieb, Ahmed; Ramsay, Ed; Schumacher, Juergen; Craig, Linden

    2005-03-01

    A 19-yr-old binturong (Arctictis binturong) with acute upper respiratory disease was euthanized. Postmortem findings included hepatocellular carcinoma, pancreatic islet cell carcinoma, and renal adenocarcinoma with metastasis to the spleen, pleura, and pericardium. A link between primary hepatic and renal neoplasms has been noted in older humans.

  13. Exploration of α1-antitrypsin treatment protocol for islet transplantation: dosing plan and route of administration.

    Science.gov (United States)

    Baranovski, Boris M; Ozeri, Eyal; Shahaf, Galit; Ochayon, David E; Schuster, Ronen; Bahar, Nofar; Kalay, Noa; Cal, Pablo; Mizrahi, Mark I; Nisim, Omer; Strauss, Pnina; Schenker, Eran; Lewis, Eli C

    2016-11-07

    Life-long weekly infusions of human α1-antitrypsin (hAAT) are currently administered as augmentation therapy for patients with genetic AAT deficiency (AATD). Several recent clinical trials attempt to extend hAAT therapy to conditions outside AATD, including type 1 diabetes. Since the endpoint for AATD is primarily the reduction of risk for pulmonary emphysema, the present study explores hAAT dose protocols and routes of administration in attempt to optimize hAAT therapy for islet-related injury. Islet-grafted mice were treated with hAAT (Glassia™; i.p. or s.c.) under an array of clinically relevant dosing plans. Serum hAAT and immunocyte cell membrane association were examined, as well as parameters of islet survival. Results indicate that dividing the commonly prescribed 60 mg/kg i.p. dose to three 20 mg/kg injections is superior in affording islet graft survival; in addition, a short dynamic descending dose protocol (240→120→60→60 mg/kg i.p.) is comparable in outcomes to indefinite 60 mg/kg injections. While hAAT pharmacokinetics after i.p. administration in mice resembles exogenous hAAT treatment in humans, s.c. administration better imitated the physiological progressive rise of hAAT during acute phase responses; nonetheless, only the 60 mg/kg dose depicted an advantage using the s.c. route. Taken together, this study provides a platform for extrapolating an islet-relevant clinical protocol from animal models that use hAAT to protect islets. In addition, the study places emphasis on outcome-oriented analyses of drug efficacy, particularly important when considering that hAAT is presently at an era of drug-repurposing towards an extended list of clinical indications outside genetic AATD.

  14. Microwell scaffolds for the extrahepatic transplantation of islets of langerhans

    NARCIS (Netherlands)

    Buitinga, M.; Truckenmuller, R.; Engelse, M.A.; Moroni, L.; Ten Hoopen, H.W.; van Blitterswijk, C.A.; de Koning, E.J.; van Apeldoorn, A.A.; Karperien, M.

    2013-01-01

    Allogeneic islet transplantation into the liver has the potential to restore normoglycemia in patients with type 1 diabetes. However, the suboptimal microenvironment for islets in the liver is likely to be involved in the progressive islet dysfunction that is often observed post-transplantation. Thi

  15. [Kidney allograft: a target for systemic disease].

    Science.gov (United States)

    Canaud, Guillaume; Legendre, Christophe

    2012-03-01

    Recurrence of disease after transplantation is frequent and represents the third cause of allograft loss. Recurrence of lupus nephritis after transplantation is rare. Kidney transplantation in patients with antiphospholipid syndrome or lupus anticoagulant is challenging due to the high risk of immediate post-transplant thrombosis and bleeding risk associated to the subsequent anticoagulation. Moreover, vascular changes associated to the presence of antiphospholipid antibodies negatively impact allograft rate survival. Recurrence of pauci immune glomerulonephritis or Goodpasture syndrome is exceptional. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  16. Effects of growth hormone, prolactin, and placental lactogen on insulin content and release, and deoxyribonucleic acid synthesis in cultured pancreatic islets

    DEFF Research Database (Denmark)

    Nielsen, Jens Høiriis

    1982-01-01

    The direct effects of human GH (hGH), ovine pituitary PRL (oPRL), and human chorionic somatomammotropin [placental lactogen (hPL)] on the endocrine pancreas were studied in isolated pancreatic islets maintained in tissue culture. Islets of Langerhans were isolated by collagenase treatment of panc...... and related hormones have a direct stimulatory effect on both the insulin production and DNA synthesis in isolated islets of Langerhans. Whether the effect is directly on the beta-cell or mediated via locally produced growth factors remains to be determined....

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

    Science.gov (United States)

    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

  18. Xenotransplantation of piscine islets into hyperglycemic rats

    Energy Technology Data Exchange (ETDEWEB)

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

    1975-02-01

    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.

  19. Inhibition of Insulin-Degrading Enzyme Does Not Increase Islet Amyloid Deposition in Vitro.

    Science.gov (United States)

    Hogan, Meghan F; Meier, Daniel T; Zraika, Sakeneh; Templin, Andrew T; Mellati, Mahnaz; Hull, Rebecca L; Leissring, Malcolm A; Kahn, Steven E

    2016-09-01

    Islet amyloid deposition in human type 2 diabetes results in β-cell loss. These amyloid deposits contain the unique amyloidogenic peptide human islet amyloid polypeptide (hIAPP), which is also a known substrate of the protease insulin-degrading enzyme (IDE). Whereas IDE inhibition has recently been demonstrated to improve glucose metabolism in mice, inhibiting it has also been shown to increase cell death when synthetic hIAPP is applied exogenously to a β-cell line. Thus, we wanted to determine whether a similar deleterious effect is observed when hIAPP is endogenously produced and secreted from islets. To address this issue, we cultured hIAPP transgenic mouse islets that have the propensity to form amyloid for 48 and 144 hours in 16.7 mM glucose in the presence and absence of the IDE inhibitor 1. At neither time interval did IDE inhibition increase amyloid formation or β-cell loss. Thus, the inhibition of IDE may represent an approach to improve glucose metabolism in human type 2 diabetes, without inducing amyloid deposition and its deleterious effects.

  20. Islet-intrinsic effects of CFTR mutation.

    Science.gov (United States)

    Koivula, Fiona N Manderson; McClenaghan, Neville H; Harper, Alan G S; Kelly, Catriona

    2016-07-01

    Cystic fibrosis-related diabetes (CFRD) is the most significant extra-pulmonary comorbidity in cystic fibrosis (CF) patients, and accelerates lung decline. In addition to the traditional view that CFRD is a consequence of fibrotic destruction of the pancreas as a whole, emerging evidence may implicate a role for cystic fibrosis transmembrane-conductance regulator (CFTR) in the regulation of insulin secretion from the pancreatic islet. Impaired first-phase insulin responses and glucose homeostasis have also been reported in CF patients. CFTR expression in both human and mouse beta cells has been confirmed, and recent studies have shown differences in endocrine pancreatic morphology from birth in CF. Recent experimental evidence suggests that functional CFTR channels are required for insulin exocytosis and the regulation of membrane potential in the pancreatic beta cell, which may account for the impairments in insulin secretion observed in many CF patients. These novel insights suggest that the pathogenesis of CFRD is more complicated than originally thought, with implications for diabetes treatment and screening in the CF population. This review summarises recent emerging evidence in support of a primary role for endocrine pancreatic dysfunction in the development of CFRD. Summary • CF is an autosomal recessive disorder caused by mutations in the CFTR gene • The vast majority of morbidity and mortality in CF results from lung disease. However CFRD is the largest extra-pulmonary co-morbidity and rapidly accelerates lung decline • Recent experimental evidence shows that functional CFTR channels are required for normal patterns of first phase insulin secretion from the pancreatic beta cell • Current clinical recommendations suggest that insulin is more effective than oral glucose-lowering drugs for the treatment of CFRD. However, the emergence of CFTR corrector and potentiator drugs may offer a personalised approach to treating diabetes in the CF population.

  1. Isolation and culture of adult Sertoli cells and their effects on the function of co-cultured allogeneic islets in vitro

    Institute of Scientific and Technical Information of China (English)

    TENG Yan; XUE Wu-jun; DING Xiao-ming; FENG Xin-shun; XIANG He-li; JIANG Ya-zhuo; TIAN Pu-xun

    2005-01-01

    Background Globally, 180 million people suffer from diabetes mellitus. Islet transplantation is believed to be an almost ideal therapy for insulin-dependent patients. How to maintain the viability and the function of isolated human islets is a challenge in clinical practice. Sertoli cells are considered ‘nurse cells'in the seminiferous tubules and have been used in cell graft protocols for neurodegenerative diseases and diabetes in many studies. Many researchers have used immature murine testes as the primarily source of Sertoli cells in islet transplantation because they are easily purified. Mature human Sertoli cells have been seldom investigated. In the present study, we developed a method for the isolation and culture of Sertoli cells derived from adult human testes, and investigated their effects on the function of allogeneic islets when they were cultured together in vitro. Methods Adult Sertoli cells were prepared successfully by two-step enzyme digestion with trypsin, collagenase and hyaluronidase. They were identified by morphological characteristics and their activity was determined by MTT colorimetry over a 28-day culture time in vitro. A glucose-stimulated insulin secretion test was performed to detect the effects of Sertoli cells on allogeneic islets' function when they were co-cultured for 21 days in vitro. Results In cultured cells, mature human Sertoli cells accounted for more than 90% of total cells. The activity of Sertoli cells reached 95% and they remained highly cytoactive for a long time in vitro (P>0.05). Compared with the islets cultured alone, the co-cultured islets with allogeneic Sertoli cells maintained higher sensitivity to glucose stimulation for the duration of the experiment (P<0.01). Conclusions A method of isolation and culture of Sertoli cells from adult testes has been established. Sertoli cells could enhance allogeneic islets' function when they were co-cultured in vitro. They could be a helper cell in islet transplantation.

  2. Renal Allograft in a Professional Boxer

    Directory of Open Access Journals (Sweden)

    Einollahi Behzad

    2008-01-01

    Full Text Available Significant health benefits result from regular physical activity for kidney transplant recipients. Nevertheless, some adverse effects also have been shown to be associated with highly intensive exercises. We report a kidney transplant professional boxer whose kidney allograft has remained in good health, despite his violent sport activities.

  3. Ultrastructural basis of acute renal allograft rejection

    NARCIS (Netherlands)

    V.D. Vuzevski (Vojislav)

    1976-01-01

    textabstractAn attempt was made: I. to demonstrate the evolution and the time of onset of the ultrastructural morphological changes in the renal parenchyma and blood vessels, as well as the ultrastructural feature of the interstitial cellular infiltration in acute rejection of kidney allografts; 2.

  4. Renal allograft rejection. Unusual scintigraphic findings

    Energy Technology Data Exchange (ETDEWEB)

    Desai, A.G.; Park, C.H.

    1986-11-01

    During sequential renal imagining for evaluation of clinically suspected rejection, focal areas of functioning renal tissue were seen in two cases of renal transplant in the midst of severe and irreversible renal allograft rejection. A probable explanation for this histopathologically confirmed and previously unreported finding is discussed.

  5. Differentiation of mesenchymal stem cells derived from pancreatic islets and bone marrow into islet-like cell phenotype.

    Directory of Open Access Journals (Sweden)

    Cristina Zanini

    Full Text Available BACKGROUND: Regarding regenerative medicine for diabetes, accessible sources of Mesenchymal Stem Cells (MSCs for induction of insular beta cell differentiation may be as important as mastering the differentiation process itself. METHODOLOGY/PRINCIPAL FINDINGS: In the present work, stem cells from pancreatic islets (human islet-mesenchymal stem cells, HI-MSCs and from human bone marrow (bone marrow mesenchymal stem cells, BM-MSCs were cultured in custom-made serum-free medium, using suitable conditions in order to induce differentiation into Islet-like Cells (ILCs. HI-MSCs and BM-MSCs were positive for the MSC markers CD105, CD73, CD90, CD29. Following this induction, HI-MSC and BM-MSC formed evident islet-like structures in the culture flasks. To investigate functional modifications after induction to ILCs, ultrastructural analysis and immunofluorescence were performed. PDX1 (pancreatic duodenal homeobox gene-1, insulin, C peptide and Glut-2 were detected in HI-ILCs whereas BM-ILCs only expressed Glut-2 and insulin. Insulin was also detected in the culture medium following glucose stimulation, confirming an initial differentiation that resulted in glucose-sensitive endocrine secretion. In order to identify proteins that were modified following differentiation from basal MSC (HI-MSCs and BM-MSCs to their HI-ILCs and BM-ILCs counterparts, proteomic analysis was performed. Three new proteins (APOA1, ATL2 and SODM were present in both ILC types, while other detected proteins were verified to be unique to the single individual differentiated cells lines. Hierarchical analysis underscored the limited similarities between HI-MSCs and BM-MSCs after induction of differentiation, and the persistence of relevant differences related to cells of different origin. CONCLUSIONS/SIGNIFICANCE: Proteomic analysis highlighted differences in the MSCs according to site of origin, reflecting spontaneous differentiation and commitment. A more detailed understanding of

  6. Comparative clinical and radiographic evaluation of mineralized cancellous bone allograft (puros®) and autogenous bone in the treatment of human periodontal intraosseous defects: 6-months follow-up study

    Science.gov (United States)

    Reddy, B. Ravinder; Sudhakar, J.; Rajesh, Nichenametla; Sandeep, V.; Reddy, Y. Muralidhar; Gnana Sagar, W. R.

    2016-01-01

    Aims: Several materials have been introduced as bone grafts, i.e., autografts, allograft, xenografts, and alloplastic grafts, and studies have shown them to produce greater clinical bone defect fill than open flap debridement alone. The aim of this clinical and radiological 6-month study was to compare and evaluate the clinical outcome of deep intraosseous defects following reconstructive surgery with the use of mineralized cancellous bone allograft (Puros®) or autogenous bone. Materials and Methods: Ten patients with 12 sites exhibiting signs of moderate generalized chronic periodontitis were enrolled in the study. The investigations were confined to two and three-walled intra bony defects with a preoperative probing depth of ≥5 mm. Six of these defects were treated with Puros® (group A) the remaining six were treated with autogenous bone graft (group B). Allocation to the two groups was randomized. The clinical parameters, plaque index (PI), gingival index (GI), probing pocket depth (PPD), clinical attachment level (CAL), and bone fill, were recorded at different time intervals at the baseline, 1 month, 3 months, and 6 months. Intraoral radiographs were taken using standardized paralleling cone technique at baseline, 1, 3, and 6 months. Statistical analysis was done by using the one-way analysis of variance (ANOVA) followed by Tukey highly significant difference. Results: Both groups resulted in decrease in probing depth (group A, 3.0 mm; group B, 2.83 mm) and gain in clinical attachment level (group A, 3.33 mm; group B, 3.0 mm) over a period of 6 months, which was statistically insignificant. Conclusion: Within the limitations of the present study, it can be concluded that both mineralized cancellous bone allograft (Puros®) or autogenous bone result in significant clinical improvements. PMID:28217545

  7. 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.

  8. Is islet transplantation ready for widespread use in diabetes?

    Institute of Scientific and Technical Information of China (English)

    WANG Jin; WANG Qing-hua; XIA Tian; TAN Jian-ming

    2011-01-01

    Up till 2000 when Edmonton group introduced islet transplant procedure in conjunction with a novel glucocorticoid-free immunosuppressive regimen rendering 100% (n=7) of patients with type 1 diabetes insulin-independent for at least 1 year, islet transplant was taken into the clinic. Although significant progress in clinical islet transplant has occurred during recent years, challenges remain, including shortage of available donor organs, technical aspects of islet preparation and transplantation, immunological rejection post-transplant, unclear long-term outcomes of islet transplantation. Special attention is given to current limitation in islet transplantation together with new possible strategies that raise expectations for the widespread use of islet transplantation in the future.

  9. Decreased humoral antibody episodes of acute renal allograft rejection in recipients expressing the HLA-DQβ1*0202 allele.

    Science.gov (United States)

    Mannam, Venkat K R; Santos, Mark; Lewis, Robert E; Cruse, Julius M

    2012-10-01

    The present investigation was designed to show the effect of human leukocyte antigen (HLA) class II molecular allelic specificities in the recipient on the induction of humoral antibody rejection, identified by C4d peritubular capillary staining, as well as specific antibody identified by Luminex technology. Major histocompatibility complex (MHC) class II molecules are expressed on dendritic cells, macrophages, and B lymphocytes and they present antigenic peptides to CD4 positive T lymphocytes. Human renal peritubular and glomerular capillaries express class II MHC molecules upon activation. Expression of class II molecules on renal microvascular endothelial cells exposes them to possible interaction with specific circulating antibodies. We hypothesize that HLA-DQβ1*0202 expression in recipients decreases the likelihood of antibody-mediated renal allograft rejection. We found that 80% (=25) of DQ2 positive haplotype recipients failed to induce humoral antibody renal allograft rejection and 20% (n=25) of DQ2 positive haplotype recipients induced humoral antibody renal allograft rejection (p=0.008). By contrast, 48% (n=46) of DQ2 negative haplotype recipients failed to induce a humoral antibody component of renal allograft rejection and 52% (n=46) of DQ2 negative haplotype recipients induced humoral antibody-mediated renal allograft rejection. Our results suggest that recipients who express the DQβ1*0202 allele are less likely to induce a humoral antibody component of acute renal allograft rejection than are those expressing DQ1, DQ3, or DQ4 alleles. DQβ1*0202 allele expression in recipients could possibly be protective against acute humoral allograft rejection and might serve as a future criterion in recipient selection and in appropriate therapy for acute renal rejection episodes.

  10. Arthroscopic meniscal allograft transplantation without bone plugs.

    Science.gov (United States)

    Alentorn-Geli, Eduard; Seijas Vázquez, Roberto; García Balletbó, Montserrat; Álvarez Díaz, Pedro; Steinbacher, Gilbert; Cuscó Segarra, Xavier; Rius Vilarrubia, Marta; Cugat Bertomeu, Ramón

    2011-02-01

    Partial or total meniscectomy are common procedures performed at Orthopedic Surgery departments. Despite providing a great relief of pain, it has been related to early onset knee osteoarthritis. Meniscal allograft transplantation has been proposed as an alternative to meniscectomy. The purposes of this study were to describe an arthroscopic meniscal allograft transplantation without bone plugs technique and to report the preliminary results. All meniscal allograft transplantations performed between 2001 and 2006 were approached for eligibility, and a total of 35 patients (involving 37 menisci) were finally engaged in the study. Patients were excluded if they had ipsilateral knee ligament reconstruction or cartilage repair surgery before meniscal transplantation or other knee surgeries after the meniscal transplantation. Scores on Lysholm, Subjective IKDC Form, and Visual Analogue Scale (VAS) scale for pain were obtained at a mean follow-up of 38.6 months and compared to pre-operative data. Data on chondral lesions were obtained during the arthroscopic procedure and through imaging (radiographs and MRI) studies pre-operatively. Two graft failures out of 59 transplants (3.4%) were found. Daily life accidents were responsible for all graft failures. Significant improvements for Lysholm, Subjective IKDC Form, and VAS for pain scores following the meniscal allograft transplantation were found (P lesion, there was no significant interactions for Lysholm (n.s.), Subjective IKDC Form (n.s.), and VAS for pain scores (n.s.). This study demonstrated that an arthroscopic meniscal allograft transplantation without bone plugs improved knee function and symptoms after a total meniscectomy. Improvements were observed independently of the degree of chondral lesion.

  11. The influence of porcine pancreas digestion parameters and islet histomorphology on islet isolation outcome.

    Science.gov (United States)

    Kinasiewicz, J; Sabat, M; Antosiak-Iwańska, M; Godlewska, E; Sitarek, E; Orłowski, T

    2011-01-01

    Transplantation of the pig islets of Langerhans is considered as the future treatment for patients suffering from type I diabetes mellitus. Despite the adaptation of modified Ricordi method and highly purified collagenase, the results of pancreas digestions are precarious. Selection of proper donor and optimal digestion procedure are fundamental. The aim of this study was to assess the impact of pancreas procuring parameters on pig islets yield. The pancreata were harvested from 69 market sows weighting over 150 kg. After intraductal injection of cold collagenase solution pancreata were transported in UW solution or under conditions of two layer method (TLM). In laboratory pancreata were digested at 37 degrees C according to Ricordi isolation method or stationary in the bottle. The particular parameters of isolation procedure were considered as substantial. Pig weight, volume of infused collagenase solution, TLM application and pancreas dividing before digestion positively affected islet yield. Additionally, the influence of pancreatic islet tissue histomorphology on isolation outcome was studied. Proper donor selection as well as adequate digestion parameters could improve pig islet recovery during islet isolation.

  12. Repair of radial and digital nerve defect with human acellular nerve allograft:6 cases report%去细胞同种异体神经移植修复桡神经和指神经缺损六例

    Institute of Scientific and Technical Information of China (English)

    唐举玉; 俞芳; 吴攀峰; 黄臻; 梁捷予; 何波; 刘小林

    2014-01-01

    Objective To explore the safety and clinical effect of the human acellular nerve allograft (hANG) for repairing peripheral nerve defects.Methods During November,2009 to October,2010,6 patients with 3 digital nerve defects and 3 radial nerve defects were repaired with hANG.During postoperation period,safety was evaluated by local wound response and laboratory testing,while the efficacy was evaluated by British Medical Research Council sensory function assessment standards,static 2-point discrimination (2PD) and Semmes-Weinstein monofilament testing.Results Three patients with 6 digital nerve defects received hANG transplant.The length of nerve graft was 20-50 mm(mean 30.8 mm).After followed up for 31-40 months,the excellent rate of 2PD was 66.7%.Two of 3 patients rahabilited as well as the normal.Three patients with radial nerve defects,whose length of nerve graft was 35-60 mm(mean 48.3 mm).The strength of extensor carpiradialis longus muscle had restored Ⅲ in 1 case,and other 2 cases had no restoration.Conclusion hANG is safe and effective for repairing peripheral nerve defects,especially for digital nerve defects.%目的 探讨去细胞同种异体神经(hANG)移植修复周围神经缺损的安全性和有效性. 方法 2009年9月-2010年10月,应用hANG移植修复周围神经缺损6例,其中指神经缺损3例、桡神经缺损3例,术后观察伤口愈合情况及生化、免疫学检查,采用英国医学研究会感觉功能评定标准、Semmes-Weinstein单丝触觉和静态两点辨别觉(2PD)评价hANG的临床效果. 结果 所有病例切口术后无红肿及渗出、愈合良好.3例指神经损伤患者共有6条指神经缺损,神经移植长度20~ 50 mm(平均30.8 mm),随访31~40个月,静态2PD优良率66.7%,其中2例4条指神经缺损患者术后感觉基本恢复正常;3例桡神经损伤患者,神经移植长度35 ~ 60 mm(平均48.3 mm),随访18 ~ 36个月,其中l例桡侧腕伸肌肌力恢复至Ⅲ级,术后

  13. Transplantation of human embryonic stem cells derived pancreatic progenitors and islets corrects diabetes in NOD/SCID mice%人胚胎干细胞体外定向诱导分化胰腺前体细胞及胰岛细胞移植治疗NOD/SCID糖尿病小鼠

    Institute of Scientific and Technical Information of China (English)

    华秀峰; 孙强; 李华峰; 孟晓梅; 王延伟; 于胜强; 丛晋; 刘芙君; 靳少华

    2014-01-01

    Objective To investigate whether pancreatic progenitors and islets differentiated from human embryonic stem cells(hESCs) could correct hyperglycemia in non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice.Methods We obtained pancreatic progenitors and islets derived from hES cells line YT1 according to the optimized four-stage differentiation protocol.Stage 1:definitive endoderm formation; Stage 2:pan creatic specialization; Stage 3:amplification of pancreatic progenitors,Stage 4:maturation of pancreatic islets.To observe the morphological changes of each stage and immunofluorescent expression of pancreatic and duodenal homeobox gene(PDX-1),glucagon,insulin,C-peptide,glucose transporter-2(Glut-2).The differentiated cells from stage 3 and stage 4 were then transplanted into one of the epididymal fat pads(EFP) of NOD/SCID mice.The survival and function of the graft were measured by immunohistochemistry and blood glucose monitor.Results The stage 4-differentiated pancreatic islets expressed mature β cell-specific markers such as glucagon,insulin and Glut 2,and even PDX-1 and C-peptide-double-positive.The stage 4-pancreatic islets had nearly 17.1% insulin-positive cells as assayed by flow cytometry analysis.Differentiated pancreatic islets released insulin/C-peptide in response to glucose stimulation.After implantation into EFP of NOD/SCID mice,hES cell-derived human pancreatic progenitors and islets corrected hyperglycemia for at least 12 weeks.Conclusions Pancreatic progenitors and islets differentiated from hESCs can correct hyperglycemia in NOD/SCID mice.%目的 探讨人胚胎干细胞(human embryonic stem cells,hESCs)体外定向诱导分化胰腺前体细胞及胰岛细胞移植治疗非肥胖糖尿病/严重联合免疫缺陷(non-obese diabetic/severe combined immunodeficient,NOD/SCID)小鼠的可行性.方法 体外分4阶段诱导hESCs定向分化为胰岛细胞:①诱导分化形成定型内胚层;②诱导胰腺细胞定向分化;③扩增

  14. Mitis group streptococci express variable pilus islet 2 pili.

    Directory of Open Access Journals (Sweden)

    Dorothea Zähner

    Full Text Available BACKGROUND: Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis are members of the Mitis group of streptococci and agents of oral biofilm, dental plaque and infective endocarditis, disease processes that involve bacteria-bacteria and bacteria-host interactions. Their close relative, the human pathogen S. pneumoniae uses pilus-islet 2 (PI-2-encoded pili to facilitate adhesion to eukaryotic cells. METHODOLOGY/PRINCIPAL FINDINGS: PI-2 pilus-encoding genetic islets were identified in S. oralis, S. mitis, and S. sanguinis, but were absent from other isolates of these species. The PI-2 islets resembled the genetic organization of the PI-2 islet of S. pneumoniae, but differed in the genes encoding the structural pilus proteins PitA and PitB. Two and three variants of pitA (a pseudogene in S. pneumoniae and pitB, respectively, were identified that showed ≈20% difference in nucleotide as well as corresponding protein sequence. Species-independent combinations of pitA and pitB variants indicated prior intra- and interspecies horizontal gene transfer events. Polyclonal antisera developed against PitA and PitB of S. oralis type strain ATCC35037 revealed that PI-2 pili in oral streptococci were composed of PitA and PitB. Electronmicrographs showed pilus structures radiating >700 nm from the bacterial surface in the wild type strain, but not in an isogenic PI-2 deletion mutant. Anti-PitB-antiserum only reacted with pili containing the same PitB variant, whereas anti-PitA antiserum was cross-reactive with the other PitA variant. Electronic multilocus sequence analysis revealed that all PI-2-encoding oral streptococci were closely-related and cluster with non-PI-2-encoding S. oralis strains. CONCLUSIONS/SIGNIFICANCE: This is the first identification of PI-2 pili in Mitis group oral streptococci. The findings provide a striking example of intra- and interspecies horizontal gene transfer. The PI-2 pilus diversity provides a possible key to

  15. Hearing Benefit in Allograft Tympanoplasty Using Tutoplast Processed Malleus

    Directory of Open Access Journals (Sweden)

    Anja Lieder

    2014-01-01

    Full Text Available Objectives. Tutoplast processed human cadaveric ossicular allografts are a safe alternative for ossicular reconstruction where there is insufficient material suitable for autograft ossiculoplasty. We present a series of 7 consecutive cases showing excellent air-bone gap closure following canal-wall-down mastoidectomy for cholesteatoma and reconstruction of the middle ear using Tutoplast processed malleus. Patients and Methods. Tympanoplasty with Tutoplast processed malleus was performed in seven patients to reconstruct the middle ear following canal-wall-down mastoidectomy in a tertiary ENT centre. Main Outcome Measures. Hearing improvement and recurrence-free period were assessed. Pre-and postoperative audiograms were performed. Results. The average pre operative hearing loss was 50 ± 13 dB, with an air-bone gap of 33 ± 7 dB. Post operative audiograms at 25 months demonstrated hearing thresholds of 29 ± 10 dB, with an air-bone gap of 14 ± 6 dB. No prosthesis extrusion was observed, which compares favourably to other commercially available prostheses. Conclusions. Tutoplast processed allografts restore conductive hearing loss in patients undergoing mastoidectomy and provide an excellent alternative when there is insufficient material suitable for autograft ossiculoplasty.

  16. Islet transplantation in type 1 diabetes

    NARCIS (Netherlands)

    de Kort, H.; de Koning, E.; Rabelink, T.; Bruijn, J.A.; Bajema, I.

    2011-01-01

    Hanneke de Kort, research fellow1, Eelco J de Koning, associate professor, head of clinical islet transplantation programme234, Ton J Rabelink, professor of medicine, chair of department of nephrology2, Jan A Bruijn, professor immunopathology1, Ingeborg M Bajema, renal and transplantation pathologis

  17. Immune tolerance in pancreatic islet xenotransplantation

    Institute of Scientific and Technical Information of China (English)

    Tian-Hua Tang; Chun-Lin Li; Xin Li; Feng-Qin Jiang; Yu-Kun Zhang; Hai-Quan Ren; Shan-Shan Su; Guo-Sheng Jiang

    2004-01-01

    AIM: To observe the effect of tail vein injection with donor hepatocytes and/or splenocytes on the islet xenotransplantation rejection.METHODS: New-bom male pigs and BALB/C mice were selected as donors and recipients respectively. Islet xenotransplantation was performed in recipients just after the third time of tail vein injection with donor hepatocytes and/or splenocytes.Macrophage phagocytosis, NK(natural killing cell) killing activity, T lymphocyte transforming function of spleen cells,antibody forming function of B lymphocytes, and T lymphocyte subsets were taken to monitor transplantation rejection. The effects of this kind of transplantation were indicated as variation of blood glucose and survival days of recipients.RESULTS: The results showed that streptozotocin (STZ) could induce diabetes mellitus models of mice. The preinjection of donor hepatocytes, splenocytes or their mixture by tail vein injection was effective in preventing donor islet transplantation from rejection, which was demonstrated by the above-mentioned immunological marks. Each group of transplantation could decrease blood glucose in recipients and increase survival days. Pre-injection of mixture of donor hepatocytes and splenocytes was more effective in preventing rejection as compared with that of donor hepatocyte or splenocyte pre-injection respectively.CONCLUSION: Pre-injection of donor hepatocytes, splenocytes or their mixture before donor islet transplantation is a good way in preventing rejection.

  18. Limitations in the Use of Fluorescein Diacetate/Propidium Iodide (FDA/PI) and Cell Permeable Nucleic Acid Stains for Viability Measurements of Isolated Islets of Langerhans.

    Science.gov (United States)

    Boyd, Vinc; Cholewa, Olivia Maria; Papas, Klearchos K

    2008-03-01

    BACKGROUND: A review of current literature shows that the combined use of the cell permeable esterase-substrate fluorescein diacetate (FDA) and the cell impermeant nucleic acid stain propidium iodide (PI) to be one of the most common fluorescence-based methods to assess the viability of isolated islets of Langerhans, and it is currently used for islet product release prior to transplantation in humans. However, results from this assay do not correlate with islet viability and function or islet transplantation success in animals or humans (Eckhard et al. 2004; Ricordi et al. 2001). This may be in part attributed to considerable differences as well as discrepancies in the use of these reagents on islets. We critically surveyed the literature and evaluated the impact of a number of variables associated with the use of FDA/PI to determine their reliability in assessing islet cell viability. In addition, we evaluated other fluorescent stains, such as SYTO(R)13, SYTO(R)24 and SYBR(R)14 as possible alternatives to FDA. RESULTS: We found that the stability of stains in storage and stock solutions, the number of islets stained, concentration of stains, staining incubation time, the buffer/media used, and the method of examining islets were significant in the final scoring of viability. For archival file photos, the exposure time and camera/software settings can also impact interpretation of viability. Although our results show that FDA does detect intracellular esterase activity and staining with PI does assess cell membrane integrity, the results obtained from using these stains did not correlate directly with expected islet function and viability per transplantation into diabetic athymic nude mice (Papas et al. 2007). In addition, the use of two nucleic acid stains, such as SYTO(R)13 and PI, for live/dead scoring exhibited staining anomalies which limit their accuracy in assessing islet viability. CONCLUSIONS: From a review of the literature and from our observations on

  19. Noninvasive Imaging of Acute Renal Allograft Rejection by Ultrasound Detection of Microbubbles Targeted to T-lymphocytes in Rats.

    NARCIS (Netherlands)

    Grabner, A.; Kentrup, D.; Mühlmeister, M.; Pawelski, H.; Biermann, C.; Bettinger, T.; Pavenstadt, H.; Schlatter, E.; Tiemann, K.; Reuter, S.

    2016-01-01

    PURPOSE: We propose CD3-antibody-mediated contrast-enhanced ultrasonography using human T-lymphocytes for image-based diagnosis of acute allograft rejection (AR) established in a rat renal transplantation model. MATERIALS AND METHODS: 15 minutes after tail vein injection of 30 x 10(6) human T-lympho

  20. Lagoon islets as indicators of recent environmental changes in the South Pacific - The New Caledonian example

    Science.gov (United States)

    Garcin, Manuel; Vendé-Leclerc, Myriam; Maurizot, Pierre; Le Cozannet, Gonéri; Robineau, Bernard; Nicolae-Lerma, Alexandre

    2016-07-01

    The question of the impacts of climate change and sea level rise on small islands is currently much discussed. The many thousands of Pacific islands in their different contexts (geodynamic, climatic, etc.) and the insufficient data available explain why it is difficult to clearly discern the specific role of climate change in the recent evolution of these islands. To address this question, we investigated the recent changes affecting 21 islets in New Caledonia's lagoon. These islets are either located on small patch-reefs inside the New Caledonia Island lagoon or lie directly on the barrier reef. Based on the studies we conducted (field surveys, reconstruction of changes in the islets over the last decades, shoreline changes) we were able to define a typology of the islets that includes 6 stages and a life expectancy index. Using the life expectancy index, we found that of the 21 islets studied, 19% are in a highly critical situation, meaning they are very likely to be endangered in the short term (within the next few years), 9.5% are in a critical situation, i.e., likely to disappear in the near future and very likely to disappear in the medium term (next few decades), 19% are evolving rapidly, which could lead to their disappearance in the medium term but not in the short term, 9.5% are not endangered in the short and medium term and 43% are not endangered at all (stable or accreting, large area, relatively higher altitude). In this context, the rise in sea level induced by climate change is an adverse factor which is likely to lower the resilience of the islets to erosion processes. Other factors like the degradation of the reef ecosystem due to variations in ocean salinity, temperature and acidity, lower sediment stocks on the beaches and foreshores, human visitors, coastal development and so on are other adverse factors that could modify the capacity for resilience of these islets. Due to their variety and sensitivity, New Caledonia's islets could thus serve

  1. Management of nonfunctioning islet cell tumors

    Institute of Scientific and Technical Information of China (English)

    Han Liang; Pu Wang; Xiao-Na Wang; Jia-Cang Wang; Xi-Shan Hao

    2004-01-01

    AIM: To more clearly define the clinical and pathological characteristics and appropriate diagnosis and treatment of nonfunctioning (NFICTs) islet cell tumors, and to review our institutional experience over the last 30 years.METHODS: The records of 43 patients confirmed to have nonfunctioning islet cell tumors of pancreas were retrospectively reviewed. Survival was estimated by the Kaplan-Meier methods and potential risk factors for survival were compared with the log-rank tests.RESULTS: The mean age was 31.63 years (range, 8 to 67 years). There were 7 men and 36 women. Twentyeight patients had a confirmed diagnosis of nonfunctioning islet cell carcinoma (NFICC) and benign islet cell tumors were found in 15 patients. The most common symptoms in patients with NFICTs were abdominal pain (55.8%),nausea and/or vomiting (32.6%), fatigue (25.6%) and abdominal mass (23.3%). Preoperative ultrasonic and computed tomography localized the tumors in all patients.Forty-three NFICTs were distributed throughout the pancreas, with 21 located to the right of the superior mesenteric vessels, 10 in the body of the pancreas, 6 in the tail of the pancreas, and multiple tumors were found in one patient. Thirty-nine of 43 patients (91%) underwent surgical resection. Surgical treatment was curative in 30patients (70%) and palliative in 9(21%). The resectability and curative resection rate in patients with NFICC of pancreas were 89% and 61%, respectively. The overall cumulative 5- and 10-year survival rates for patients with NFICC were 58.05% and 29.03%, respectively. Radical operation and diameter of cancer small than :10 cm were positive prognostic factors in females younger than 30years old. Multivariate Cox regression analysis indicated that radical operation was the only independent prognostic factor, P=0.007.CONCLUSION: Nonfunctioning islet cell tumors of pancreas are found mainly in young women. The long-term results for patients undergone surgery, especially curative resection are

  2. The Langerhans islet cells of female rabbits are differentially affected by hypothyroidism depending on the islet size.

    Science.gov (United States)

    Rodríguez-Castelán, J; Nicolás, L; Morimoto, S; Cuevas, E

    2015-04-01

    Effects of hypothyroidism on the glucose and insulin levels are controversial, and its impact on the Langerhans islet morphology of adult subjects has been poorly addressed. In spite of hypothyroidism and diabetes mellitus are more frequent in females than in males, most studies using animal models have been done in males. The effect of hypothyroidism on the immunolabeling of thyroid hormone receptors (TRs) and thyrotropin receptor (TSHR) of islet cells is unknown. The aim of this study was to determine the effect of hypothyroidism on the glucose and insulin concentrations, morphometry of islets, and immunostaining of TRs α1-2 and β1 and TSHR of islet cells in female rabbits. Control and hypothyroid (0.02% of methimazole for 30 days) animals were used to quantify blood levels of glucose and insulin, density of islets, cross-sectional area (CSA) of islets, number of cells per islet, cell proliferation, and the immunolabeling of TRs α1-2, TRβ1, and TSHR. Student's t or Mann-Whitney-U tests, two-way ANOVAs, and Fischer's tests were applied. Concentrations of glucose and insulin, as well as the insulin resistance were similar between groups. Hypothyroidism did not affect the density or the CSA of islets. The analysis of islets by size showed that hypothyroidism reduced the cell number in large and medium islets, but not in small ones. In small islets, cell proliferation was increased. The immunoreactivity of TRα1-2, TRβ1, and TSHR was increased by hypothyroidism in all islet sizes. Our results show that hypothyroidism affects differentially the islet cells depending on the size of islets.

  3. Mechanoreceptor Reinnervation of Autografts Versus Allografts After Anterior Cruciate Ligament Reconstruction

    Science.gov (United States)

    Young, Simon W.; Valladares, Roberto D.; Loi, Florence; Dragoo, Jason L.

    2016-01-01

    Background: Loss of proprioceptive function occurs after anterior cruciate ligament (ACL) rupture. Clinical, motor, and proprioceptive function is known to improve after ACL reconstruction but does not return to normal. While histological studies of human ACL allografts have been unable to demonstrate mechanoreceptor reinnervation, animal data suggest that reinnervation may occur when an autograft is used. Purpose: To compare the presence or absence of mechanoreceptors between allograft versus autograft after ACL reconstruction in humans. Study Design: Cohort study; Level of evidence, 3. Methods: Ten patients with previous ACL reconstruction presenting for either revision ACL surgery or knee arthroscopy for other reasons were enrolled in a prospective, comparative study. Five patients had a previous autograft ACL and 5 patients had an allograft. Biopsies, either from intact or ruptured grafts, were taken from identical locations as close to the femoral and tibial insertions as possible. Specimens were stained with hematoxylin-eosin (H-E) and monoclonal antibodies against neurofilament protein (NFP), known to be present in mechanoreceptor tissue. Immunohistochemical examination was carried out, and the number of NFP+ neural tissue analogs was counted and compared with that of native ACL tissue. Results: The mean time between original graft and biopsy was 6.9 years (range, 0.5-15 years). Histological examination showed significantly less NFP+ neural analogs in allograft and autograft patients than control tissue (mean number of NFP+ analogs per high-power field, 0.7 ± 0.9 [allograft] and 0.5 ± 0.8 [autograft] vs 4.7 ± 0.9 [controls]; P proprioceptive deficits seen clinically after ACL reconstruction. PMID:27803939

  4. Nonmyeloablative allografting for newly diagnosed multiple myeloma: the experience of the Gruppo Italiano Trapianti di Midollo

    Science.gov (United States)

    Rotta, Marcello; Patriarca, Francesca; Mattei, Daniele; Allione, Bernardino; Carnevale-Schianca, Fabrizio; Sorasio, Roberto; Rambaldi, Alessandro; Casini, Marco; Parma, Matteo; Bavaro, Pasqua; Onida, Francesco; Busca, Alessandro; Castagna, Luca; Benedetti, Edoardo; Iori, Anna Paola; Giaccone, Luisa; Palumbo, Antonio; Corradini, Paolo; Fanin, Renato; Maloney, David; Storb, Rainer; Baldi, Ileana; Ricardi, Umberto; Boccadoro, Mario

    2009-01-01

    Despite recent advances, allografting remains the only potential cure for myeloma. From July 1999 to June 2005, 100 newly diagnosed patients younger than 65 years were enrolled in a prospective multicenter study. First-line treatment included vincristin, adriamycin, and dexamethasone (VAD)–based induction chemotherapy, a cytoreductive autograft (melphalan 200 mg/m2) followed by a single dose of nonmyeloablative total body irradiation and allografting from an human leukocyte antigen (HLA)–identical sibling. Primary end points were the overall survival (OS) and event-free survival (EFS) from diagnosis. After a median follow-up of 5 years, OS was not reached, and EFS was 37 months. Incidences of acute and chronic graft-versus-host disease (GVHD) were 38% and 50%, respectively. Complete remission (CR) was achieved in 53% of patients. Profound cytoreduction (CR or very good partial remission) before allografting was associated with achievement of posttransplantation CR (hazard ratio [HR] 2.20, P = .03) and longer EFS (HR 0.33, P < .01). Conversely, development of chronic GVHD was not correlated with CR or response duration. This tandem transplantation approach allows prolonged survival and long-term disease control in patients with reduced tumor burden at the time of allografting. We are currently investigating the role of “new drugs” in intensifying pretransplantation cytoreduction and posttransplantation graft-versus-myeloma effects to further improve clinical outcomes. (http://ClinicalTrials.gov; NCT-00702247.) PMID:19064724

  5. Optimized total body irradiation for induction of renal allograft tolerance through mixed chimerism in cynomolgus monkeys

    Energy Technology Data Exchange (ETDEWEB)

    Kimikawa, Masaaki; Kawai, Tatsuo; Ota, Kazuo [Tokyo Women`s Medical Coll. (Japan)

    1996-12-01

    We previously demonstrated that a nonmyeloablative preparative regimen can induce mixed chimerism and renal allograft tolerance between MHC-disparate non-human primates. The basic regimen includes anti-thymocyte globulin (ATG), total body irradiation (TBI, 300 cGy), thymic irradiation (TI, 700 cGy), splenectomy, donor bone marrow (DBM) infusion, and posttransplant cyclosporine therapy (CYA, discontinued after 4 weeks). To evaluate the importance and to minimize the toxicity of irradiation, kidney allografts were transplanted with various manipulations of the irradiation protocol. Monkeys treated with the basic protocol without TBI and TI did not develop chimerism or long-term allograft survival. In monkeys treated with the full protocol, all six monkeys treated with two fractionated dose of 150 cGy developed chimerism and five monkeys appeared tolerant. In contrast, only two of the four monkeys treated with fractionated doses of 125 cGy developed chimerism and only one monkey survived long term. The degree of lymphocyte depletion in all recipients was proportional to the TBI dose. The fractionated TBI regimen of 150 cGy appears to be the most consistently effective regimen for establishing donor bone marrow cell engraftment and allograft tolerance. (author)

  6. Characterization of acute renal allograft rejection by proteomic analysis of renal tissue in rat.

    Science.gov (United States)

    Chen, Gang; Huang, Jing-Bin; Mi, Jie; He, Yun-Feng; Wu, Xiao-Hou; Luo, Chun-Li; Liang, Si-Min; Li, Jia-Bing; Tang, Ya-Xiong; Li, Jie

    2012-02-01

    Rapid and reliable biomarkers of renal allograft rejection have not been available. This study aimed to investigate biomarkers in renal allograft tissue using proteomic analysis. Orthotopic kidney transplantations were performed using Fisher (F344) or Lewis rats as donors and Lewis rats as recipients. Syngenic control group (Group I) constituted F344-to-F344 orthotopic kidney allo-transplantations (n = 8); and allogenic group (Group II) consisted of F344-to-Lewis orthotopic kidney allo-transplantations (n = 8). Renal tissues were harvested 7 days after transplantation. Samples were analyzed using 2-D electrophoresis and matrix assisted laser desorption ionization-time of flight mass spectrometry. 6 differentially expressed proteins were identified between allogenic group and syngenic control group. A rat model of acute renal allograft rejection was successfully set up. Differentially expressed proteins in renal allograft tissue of rat were detected using proteomic analysis and might serve as novel diagnostic and therapeutic targets in human. Quantitative proteomics, using MALDL-TOF-MS methodology has the potential to provide a profiling and a deeper understanding of acute renal rejection.

  7. Evaluation of kidney allograft status using novel ultrasonic technologies

    Directory of Open Access Journals (Sweden)

    Cheng Yang

    2015-07-01

    Full Text Available Early diagnosis of kidney allograft injury contributes to proper decisions regarding treatment strategy and promotes the long-term survival of both the recipients and the allografts. Although biopsy remains the gold standard, non-invasive methods of kidney allograft evaluation are required for clinical practice. Recently, novel ultrasonic technologies have been applied in the evaluation and diagnosis of kidney allograft status, including tissue elasticity quantification using acoustic radiation force impulse (ARFI and contrast-enhanced ultrasonography (CEUS. In this review, we discuss current opinions on the application of ARFI and CEUS for evaluating kidney allograft function and their possible influencing factors, advantages and limitations. We also compare these two technologies with other non-invasive diagnostic methods, including nuclear medicine and radiology. While the role of novel non-invasive ultrasonic technologies in the assessment of kidney allografts requires further investigation, the use of such technologies remains highly promising.

  8. Characterization of a novel functional protein in the pancreatic islet: islet homeostasis protein regulation of glucagon synthesis in α cells.

    Science.gov (United States)

    Oh, Seh-Hoon; Darwiche, Houda; Cho, Jae-Hyoung; Shupe, Thomas; Petersen, Bryon E

    2012-01-01

    We have identified a novel protein in bone marrow-derived insulin-producing cells. Here we characterize this protein, hereby named islet homeostasis protein (IHoP), in the pancreatic islet. Detection of IHoP mRNA and protein was performed using reverse transcriptase-polymerase chain reaction, immunocytochemistry, and in situ hybridization. Islet homeostasis protein functions were utilizing proliferation, insulin secretion by in vitro assays, and following small interfering RNA protocols for suppression of IHoP. We found that IHoP did not homolog with known pancreatic hormones. Islet homeostasis protein expression was seen in both bone marrow-derived insulin-producing cells and isolated pancreatic islets. Immunohistochemistry on pancreatic islet revealed that IHoP localized to the glucagon-synthesizing α cells. Inhibition of IHoP by small interfering RNA resulted in the loss of glucagon expression, which induced low blood glucose levels (63-85 mg/dL). Subsequently, cellular apoptosis was observed throughout the islet, including the insulin-producing β cells. Islets of preonset diabetic patients showed normal expression of IHoP and glucagon; however, IHoP was lost upon onset of the disease. These data suggest that IHoP could be a new functional protein in the islet and may play a role in islet homeostasis.

  9. Islet β-cell ghrelin signaling for inhibition of insulin secretion.

    Science.gov (United States)

    Dezaki, Katsuya; Yada, Toshihiko

    2012-01-01

    Ghrelin, an acylated 28-amino acid peptide, was isolated from the stomach, where circulating ghrelin is produced predominantly. In addition to its unique role in regulating growth-hormone release, mealtime hunger, lipid metabolism, and the cardiovascular system, ghrelin is involved in the regulation of glucose metabolism. Ghrelin is expressed in pancreatic islets and released into pancreatic microcirculations. Ghrelin inhibits insulin release in mice, rats, and humans. Pharmacological and genetic blockades of islet-derived ghrelin markedly augment glucose-induced insulin release. The signal transduction mechanisms of ghrelin in islet β-cells are very unique, being distinct from those utilized for growth-hormone release. Ghrelin attenuates the glucose-induced cAMP production and PKA activation, which drives activation of Kv channels and suppression of the glucose-induced [Ca(2+)](i) increase and insulin release in β-cells. Insulinostatic function of the ghrelin-GHS-R system in islets is a potential therapeutic target for type 2 diabetes. Copyright © 2012 Elsevier Inc. All rights reserved.

  10. Reversal of Early Diabetic Nephropathy by Islet Transplantation under the Kidney Capsule in a Rat Model

    Directory of Open Access Journals (Sweden)

    Yunqiang He

    2016-01-01

    Full Text Available Objective. Diabetic nephropathy (DN is a common microvascular complication of diabetes mellitus, and insulin therapy has many side effects in the treatment of DN. Islet transplantation has emerged as a promising therapy for diabetic patients. This study was established to investigate its advantageous effects in a rat model of early DN. Methods. Streptozotocin was administered to the rats to induce diabetes. Twelve weeks later, the diabetic rats were divided into 3 groups: the islet-transplanted group (IT group, the insulin-treated group (IN group, and the untreated group (DN group. Renal injury and kidney structure were assessed by urinalysis and transmission electron microscopy (TEM detection. Immunohistochemical staining and western blotting were performed to assess renal fibrosis levels. Results. The early DN features were reversed and the glomerular filtration barrier and basement membrane structures were improved at 4 weeks after islet transplantation. The urine microalbumin-to-creatinine ratio (ACR, protein-to-creatinine ratio, and mean thickness of the glomerular basement membrane (GBM were significantly decreased in the IT group. The expression of renal fibrotic factors was also significantly decreased. Conclusions. These data suggest that early DN can be reversed after islet transplantation, and they may facilitate the development of a clinical therapeutic strategy for human diabetes mellitus.

  11. Collagen V Is a Potential Substrate for Clostridial Collagenase G in Pancreatic Islet Isolation

    Directory of Open Access Journals (Sweden)

    Hiroki Shima

    2016-01-01

    Full Text Available The clostridial collagenases, H and G, play key roles in pancreatic islet isolation. Collagenases digest the peptide bond between Yaa and the subsequent Gly in Gly-Xaa-Yaa repeats. To fully understand the pancreatic islet isolation process, identification of the collagenase substrates in the tissue is very important. Although collagen types I and III were reported as possible substrates for collagenase H, the substrate for collagenase G remains unknown. In this study, collagen type V was focused upon as the target for collagenases. In vitro digestion experiments for collagen type V were performed and analyzed by SDS-PAGE and mass spectrometry. Porcine pancreatic tissues were digested in vitro under three conditions and observed during digestion. The results revealed that collagen type V was only digested by collagenase G and that the digestion was initiated from the N-terminal part. Tissue degradation during porcine islet isolation was only observed in the presence of both collagenases H and G. These findings suggest that collagen type V is one of the substrates for collagenase G. The enzymatic activity of collagenase G appears to be more important for pancreatic islet isolation in large mammals such as pigs and humans.

  12. Dexamethasone counteracts the effect of prolactin on islet function: implications for islet regulation in late pregnancy.

    Science.gov (United States)

    Weinhaus, A J; Bhagroo, N V; Brelje, T C; Sorenson, R L

    2000-04-01

    Islets undergo a number of up-regulatory changes to meet the increased demand for insulin during pregnancy, including increased insulin secretion and beta-cell proliferation. It has been shown that elevated lactogenic hormone is directly responsible for these changes, which occur in a phasic pattern, peaking on day 15 of pregnancy and returning to control levels by day 20 (term). As placental lactogen levels remain elevated through late gestation, it was of interest to determine whether glucocorticoids (which increase during late gestation) could counteract the effects of lactogens on insulin secretion, beta-cell proliferation, and apoptosis. We found that insulin secretion measured over 24 h in culture and acute secretion measured over 1 h in response to high glucose were increased at least 2-fold by PRL treatment after 6 days in culture. Dexamethasone (DEX) treatment had a significant inhibitory effect on secretion in a dose-dependent manner at concentrations greater than 1 nM. At 100 nM, a concentration equivalent to the plasma corticosteroid level during late pregnancy, DEX inhibited secretion to below control levels. The addition of DEX (>1 nM) inhibited secretion from PRL-treated islets to levels similar to those produced by DEX treatment alone. Bromodeoxyuridine (10 microM) staining for the final 24 h of a 6-day culture showed that PRL treatment increased cell proliferation 6-fold over the control level. DEX treatment alone (1-1000 nM) did not reduce cell division below the control level, but significantly inhibited the rate of division in PRL-treated islets. YoYo-1, an ultrasensitive fluorescent nucleic acid stain, was added (1 microM; 8 h) to the medium after 1-3 days of culture to examine cell death. Islets examined under confocal microscopy showed that DEX treatment (100 nM) increased the number of cells with apoptotic nuclear morphologies. This was quantified by counting the number of YoYo-labeled nuclei per islet under conventional epifluorescence

  13. [Tubulointerstitial rejection of renal allografts].

    Science.gov (United States)

    Malušková, Jana; Honsová, Eva

    2015-01-01

    Tubulo-intersticial rejection represents T-cell mediated rejection of kidney allografts with the morphology of immune-mediated interstitial nephritis. Diagnosis is dependent on the histopathological evaluation of a graft biopsy sample. The key morphological features are interstitial inflammatory infiltrate and damage to tubular epithelial cell which in severe cases can result in the ruptures of the tubular basement membranes. The differential diagnosis of tubulo-interstitial rejection includes acute interstitial nephritis and viral inflammatory kidney diseases, mainly polyomavirus nephropathy.

  14. Urinary calprotectin and posttransplant renal allograft injury.

    Directory of Open Access Journals (Sweden)

    Martin Tepel

    Full Text Available OBJECTIVE: Current methods do not predict the acute renal allograft injury immediately after kidney transplantation. We evaluated the diagnostic performance of urinary calprotectin for predicting immediate posttransplant allograft injury. METHODS: In a multicenter, prospective-cohort study of 144 incipient renal transplant recipients, we postoperatively measured urinary calprotectin using an enzyme-linked immunosorbent assay and estimated glomerular filtration rate (eGFR after 4 weeks, 6 months, and 12 months. RESULTS: We observed a significant inverse association of urinary calprotectin concentrations and eGFR 4 weeks after transplantation (Spearman r =  -0.33; P<0.001. Compared to the lowest quartile, patients in the highest quartile of urinary calprotectin had an increased risk for an eGFR less than 30 mL/min/1.73 m(2 four weeks after transplantation (relative risk, 4.3; P<0.001; sensitivity, 0.92; 95% CI, 0.77 to 0.98; specificity, 0.48; 95% CI, 0.31 to 0.66. Higher urinary calprotectin concentrations predicted impaired kidney function 4 weeks after transplantation, as well as 6 months and 12 months after transplantation. When data were analyzed using the urinary calprotectin/creatinine-ratio similar results were obtained. Urinary calprotectin was superior to current use of absolute change of plasma creatinine to predict allograft function 12 months after transplantation. Urinary calprotectin predicted an increased risk both in transplants from living and deceased donors. Multivariate linear regression showed that higher urinary calprotectin concentrations and older donor age predicted lower eGFR four weeks, 6 months, and 12 months after transplantation. CONCLUSIONS: Urinary calprotectin is an early, noninvasive predictor of immediate renal allograft injury after kidney transplantation.

  15. Late de novo minimal change disease in a renal allograft

    Directory of Open Access Journals (Sweden)

    Madhan Krishan

    2009-01-01

    Full Text Available Among the causes of the nephrotic syndrome in renal allografts, minimal change disease is a rarity with only few cases described in the medical literature. Most cases described have occurred early in the post-transplant course. There is no established treatment for the condition but prognosis is favorable. We describe a case of minimal change disease that developed 8 years after a successful transplantation of a renal allograft in a middle-aged woman. The nephrotic syndrome was accompanied by deterioration of allograft function. Treatment with mycophenolate mofetil was successful in inducing remission and stabilizing allograft function.

  16. Late de novo minimal change disease in a renal allograft.

    Science.gov (United States)

    Madhan, Krishan K; Temple-Camp, Cynric R E

    2009-03-01

    Among the causes of the nephrotic syndrome in renal allografts, minimal change disease is a rarity with only few cases described in the medical literature. Most cases described have occurred early in the post-transplant course. There is no established treatment for the condition but prognosis is favorable. We describe a case of minimal change disease that developed 8 years after a successful transplantation of a renal allograft in a middle-aged woman. The nephrotic syndrome was accompanied by deterioration of allograft function. Treatment with mycophenolate mofetil was successful in inducing remission and stabilizing allograft function.

  17. Cytomegalovirus and chronic allograft rejection in liver transplantation

    Institute of Scientific and Technical Information of China (English)

    Liang-Hui Gao; Shu-Sen Zheng

    2004-01-01

    Cytomegalovirus (CMV) remains one of the most frequent viral infections and the most common cause of death after liver transplantation (LT). Chronic allograft liver rejection remains the major obstacle to long-term allograft survival and CMV infection is one of the suggested risk factors for chronic allograft rejection. The precise relationship between cytomegalovirus and chronic rejection remains uncertain.This review addresses the morbidity of cytomegalovirus infection and the risk factors associated with it, the relationship between cytomegalovirus and chronic allograft liver rejection and the potential mechanisms of it.

  18. Differentiation of fetal pancreatic stem cells into neuron-like and islet-like cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Xiufeng Hua; Yanwei Wang; Peiwen Lian; Shouxin Zhang; Jianyuan Li; Haiyan Wang; Shulin Chen; Wei Gao

    2012-01-01

    Pancreatic stem cells were isolated and cultured from aborted human fetal pancreases of gestational age 14-20 weeks.They were seeded at a density of 1 × 104 in serum-free media for differentiation into neuron-like cells, expressing β-tubulin III and glial fibrillary acidic protein.These neuron-like cells displayed a synapse-like morphology and appeared to form a neuronal network.Pancreatic stem cells were also seeded at a density of 1 × 105 for differentiation into islet-like cells, expressing insulin and glucagon, with an islet-like morphology.These cells had glucose-stimulated secretion of human insulin and C-peptide.Results suggest that pancreatic stem cells can be differentiated into neuron-like and islet-like cells.

  19. Glycerol-Preserved Arterial Allografts Evaluated in the Infrarenal Rat Aorta

    National Research Council Canada - National Science Library

    Fahner, P.J; Idu, M.M; van Gulik, T.M; van Wijk, B; van der Wal, A.C; Legemate, D.A

    2009-01-01

    .... Since glycerol preservation proved effective for the storage of skin allografts, this preservation method was investigated for vascular allografts using a rat aortic transplantation model. Methods...

  20. Rabbit trochlear model of osteochondral allograft transplantation.

    Science.gov (United States)

    To, Nhat; Curtiss, Shane; Neu, Corey P; Salgado, Christopher J; Jamali, Amir A

    2011-10-01

    Allografting and autografting of osteochondral tissues is a promising strategy to treat articular cartilage lesions in damaged joints. We developed a new model of fresh osteochondral allografting using the entire rabbit trochlea. The objective of the current study was to demonstrate that this model would achieve reproducible graft-host healing and maintain normal articular cartilage histologic, immunolocalization, and biochemical characteristics after transplantation under diverse storage and transplantation conditions. New Zealand white (n = 8) and Dutch belted (n = 8) rabbits underwent a 2-stage transplantation operation using osteochondral grafts that had been stored for 2 or 4 wk. Trochlear grafts harvested from the left knee were transplanted to the right knee as either autografts or allografts. Grafts were fixed with 22-gauge steel wire or 3-0 nylon suture. Rabbits were euthanized for evaluation at 1, 2, 4, 6, and 12 wk after transplantation. All grafts that remained in vivo for at least 4 wk demonstrated 100% interface healing by microCT. Trabecular bridging was present at the host-graft interface starting at 2 wk after transplantation, with no significant difference in cartilage histology between the various groups. The combined histology scores indicated minimal evidence of osteoarthritis. Immunostaining revealed that superficial zone protein was localized at the surface of all transplants. The rabbit trochlear model met our criteria for a successful model in regard to the ease of the procedure, low rate of surgical complications, relatively large articular cartilage surface area, and amount of host-graft bone interface available for analysis.

  1. Extensor mechanism allograft in total knee arthroplasty

    Science.gov (United States)

    Helito, Camilo Partezani; Gobbi, Riccardo Gomes; Tozi, Mateus Ramos; Félix, Alessandro Monterroso; Angelini, Fábio Janson; Pécora, José Ricardo

    2013-01-01

    Objective To analyze the experience with allograft transplantation of the extensor mechanism in total knee arthroplasty and compare results with the international experience. Methods We retrospectively evaluated three cases of extensor mechanism allograft after total knee arthroplasty performed in our hospital with the aid of one of the few tissue banks in Brazil and attempt to establish whether our experiences were similar to others reported in the world literature regarding patient indication, techniques, and outcomes. Results Two cases went well with the adopted procedure, and one case showed bad results and progressed to amputation. As shown in the literature, the adequate tension of the graft, appropriate tibial fixation and especially the adequate patient selection are the better predictors of good outcomes. Previous chronic infection can be an unfavorable predictor. Conclusion This surgical procedure has precise indication, albeit uncommon, either because of the rarity of the problem or because of the low availability of allografts, due to the scarcity of tissue banks in Brazil. Level of Evidence IV, Case Series. PMID:24453688

  2. Uremic escape of renal allograft rejection

    Energy Technology Data Exchange (ETDEWEB)

    van Schilfgaarde, R. (Rijksuniversiteit Leiden (Netherlands). Academisch Ziekenhuis); van Breda Vriesman, P.J.C. (Rijksuniversiteit Limburg Maastricht (Netherlands). Dept. of Immunopathology)

    1981-10-01

    It is demonstrated in rats that, in the presence of early postoperative severe but transient uremia, the survival of first set Brown-Norway (BN) renal allografts in Lewis (LEW) recipients is at least three times prolonged when compared to non-uremic controls. This phenomenon is called 'uremic escape of renal allograft rejection'. By means of lethal X-irradiation of donors of BN kidneys transplanted into transiently uremic and non-uremic LEW recipients, the presence of passenger lymphocyte immunocompetence is demonstrated to be obilgatory for this phenomenon to occur. As a result of mobile passenger lymphocyte immunocompetence, a graft-versus-host (GVH) reaction is elicited in the spleens of LEW recipients of BN kidneys which amplifies the host response. The splenomegaly observed in LEW recipients of BN kidneys is caused not only by this GVH reaction, which is shown to be exquisitely sensitive to even mild uremia. It is also contributed to by a proliferative response of the host against the graft (which latter response is equated with an in vivo equivalent of a unilateral mixed lymphocyte reaction (MLR)), since the reduction in spleen weights caused by abrogation of mobile passenger lymphocyte immunocompetence brought about by lethal donor X-irradiation is increased significantly by early postoperative severe but transient uremia. It is concluded that in uremic escape of renal allograft rejection both reactions are suppressed by uremia during the early post-operative period.

  3. Possible modulatory effect of endogenous islet catecholamines on insulin secretion

    Directory of Open Access Journals (Sweden)

    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.

  4. File list: Pol.Pan.50.AllAg.Pancreatic_islets [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Pan.50.AllAg.Pancreatic_islets hg19 RNA polymerase Pancreas Pancreatic islets h...ttp://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.Pan.50.AllAg.Pancreatic_islets.bed ...

  5. File list: Pol.Pan.10.AllAg.Pancreatic_islets [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Pan.10.AllAg.Pancreatic_islets hg19 RNA polymerase Pancreas Pancreatic islets h...ttp://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.Pan.10.AllAg.Pancreatic_islets.bed ...

  6. File list: Pol.Pan.20.AllAg.Pancreatic_islets [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Pan.20.AllAg.Pancreatic_islets hg19 RNA polymerase Pancreas Pancreatic islets h...ttp://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.Pan.20.AllAg.Pancreatic_islets.bed ...

  7. File list: Pol.Pan.05.AllAg.Pancreatic_islets [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Pol.Pan.05.AllAg.Pancreatic_islets hg19 RNA polymerase Pancreas Pancreatic islets h...ttp://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/Pol.Pan.05.AllAg.Pancreatic_islets.bed ...

  8. Expression of GSK-3β in renal allograft tissue and its significance in pathogenesis of chronic allograft dysfunction

    Directory of Open Access Journals (Sweden)

    Yan Qiang

    2012-01-01

    Full Text Available Abstract Objective To explore the expression of Glycogen synthase kinase 3 beta (GSK-3β in renal allograft tissue and its significance in the pathogenesis of chronic allograft dysfunction. Methods Renal allograft biopsy was performed in all of the renal allograft recipients with proteinuria or increased serum creatinine level who came into our hospital from January 2007 to December 2009. Among them 28 cases was diagnosed as chronic allograft dysfunction based on pahtological observation, including 21 males with a mean age of 45 ± 10 years old and 7 females with a mean age of 42 ± 9 years old. The time from kidney transplantation to biopsy were 1-9 (3.5 years. Their serum creatinine level were 206 ± 122 umol/L. Immunohistochemical assay and computer-assisted genuine color image analysis system (imagepro-plus 6.0 were used to detect the expression of GSK-3β in the renal allografts of 28 cases of recipients with chronic allograft dysfunction. Mean area and mean integrated optical density of GSK-3β expression were calculated. The relationship between expression level of GSK-3β and either the grade of inflammatory cell infiltration or interstitial fibrosis/tubular atrophy in renal allograft was analyzed. Five specimens of healthy renal tissue were used as controls. Results The expression level of the GSK-3β was significantly increased in the renal allograft tissue of recipients with chronic allograft dysfunction, compared to normal renal tissues, and GSK-3β expression became stronger along with the increasing of the grade of either inflammatory cell infiltration or interstitial fibrosis/tubular atrophy in renal allograft tissue. Conclusion There might be a positive correlation between either inflammatory cell infiltration or interstitial fibrosis/tubular atrophy and high GSK-3β expression in renal allograft tissue. Virtual slides The virtual slide(s for this article can be found here: http

  9. Computer simulations in comparison with in vivo measurements of nifedipine-induced changes in renal allograft hemodynamics.

    Science.gov (United States)

    Merkus, J W; van Asten, W N; Hilbrands, L B; Hoitsma, A J; Koene, R A; Skotnicki, S H

    1993-09-01

    Analysis of Doppler spectrum waveforms is increasingly used in the differential diagnosis of human renal allograft dysfunction. The physiologic interpretation of changes in Doppler spectra obtained from renal allografts, however, remains a major problem. Computer simulation models of the renal circulation may provide insight into the physiologic mechanisms responsible for changes in Doppler spectrum characteristics. The results of measurements of renal allograft hemodynamics with both determinations of PAH clearance and Doppler spectrum analysis in 11 kidney allograft recipients were explained physiologically using a computer simulation model of kidney allograft hemodynamics. Using PAH clearance and blood pressure measurements a significant decrease in RVR was found (from 0.32 +/- 0.17 to 0.20 +/- 0.07 mm Hg x min/ml, P nifedipine. The Doppler spectrum waveform obtained from interlobar renal arteries showed a decrease in the RI (from 0.60 +/- 0.04 to 0.56 +/- 0.06; P < 0.05) and Tmax (from 133 +/- 32 to 98 +/- 32 ms; P < 0.05). The user-designed simulation model of renal hemodynamics showed comparable changes of the waveform when, in the model, the analogs of blood pressure, impedance of the artery, and the impedance of the peripheral vascular bed were altered proportionally.

  10. Cytokines inducing bone marrow SCA+ cells migration into pancreatic islet and conversion into insulin-positive cells in vivo.

    Directory of Open Access Journals (Sweden)

    LuGuang Luo

    Full Text Available We hypothesize that specific bone marrow lineages and cytokine treatment may facilitate bone marrow migration into islets, leading to a conversion into insulin producing cells in vivo. In this study we focused on identifying which bone marrow subpopulations and cytokine treatments play a role in bone marrow supporting islet function in vivo by evaluating whether bone marrow is capable of migrating into islets as well as converting into insulin positive cells. We approached this aim by utilizing several bone marrow lineages and cytokine-treated bone marrow from green fluorescent protein (GFP positive bone marrow donors. Sorted lineages of Mac-1(+, Mac-1(-, Sca(+, Sca(-, Sca(-/Mac-1(+ and Sca(+/Mac-1(- from GFP positive mice were transplanted to irradiated C57BL6 GFP negative mice. Bone marrow from transgenic human ubiquitin C promoter GFP (uGFP, with strong signal C57BL6 mice was transplanted into GFP negative C57BL6 recipients. After eight weeks, migration of GFP positive donor' bone marrow to the recipient's pancreatic islets was evaluated as the percentage of positive GFP islets/total islets. The results show that the most effective migration comes from the Sca(+/Mac(- lineage and these cells, treated with cytokines for 48 hours, were found to have converted into insulin positive cells in pancreatic islets in vivo. This study suggests that bone marrow lineage positive cells and cytokine treatments are critical factors in determining whether bone marrow is able to migrate and form insulin producing cells in vivo. The mechanisms causing this facilitation as well as bone marrow converting to pancreatic beta cells still need to be investigated.

  11. Tacrolimus inhibits the revascularization of isolated pancreatic islets.

    Directory of Open Access Journals (Sweden)

    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.

  12. Lack of evidence for a role of islet autoimmunity in the aetiology of canine diabetes mellitus.

    Directory of Open Access Journals (Sweden)

    Kerstin M Ahlgren

    Full Text Available AIMS/HYPOTHESIS: Diabetes mellitus is one of the most common endocrine disorders in dogs and is commonly proposed to be of autoimmune origin. Although the clinical presentation of human type 1 diabetes (T1D and canine diabetes are similar, the aetiologies may differ. The aim of this study was to investigate if autoimmune aetiology resembling human T1D is as prevalent in dogs as previously reported. METHODS: Sera from 121 diabetic dogs representing 40 different breeds were tested for islet cell antibodies (ICA and GAD65 autoantibodies (GADA and compared with sera from 133 healthy dogs. ICA was detected by indirect immunofluorescence using both canine and human frozen sections. GADA was detected by in vitro transcription and translation (ITT of human and canine GAD65, followed by immune precipitation. Sections of pancreata from five diabetic dogs and two control dogs were examined histopathologically including immunostaining for insulin, glucagon, somatostatin and pancreas polypeptide. RESULTS: None of the canine sera analysed tested positive for ICA on sections of frozen canine or human ICA pancreas. However, serum from one diabetic dog was weakly positive in the canine GADA assay and serum from one healthy dog was weakly positive in the human GADA assay. Histopathology showed marked degenerative changes in endocrine islets, including vacuolisation and variable loss of immune-staining for insulin. No sign of inflammation was noted. CONCLUSIONS/INTERPRETATIONS: Contrary to previous observations, based on results from tests for humoral autoreactivity towards islet proteins using four different assays, and histopathological examinations, we do not find any support for an islet autoimmune aetiology in canine diabetes mellitus.

  13. CT features of nonfunctioning islet cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    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.

  14. Interest of Polyelectrolyte Multilayer thin Films in Tissue Engineering:Application to Vascular Allograft

    Institute of Scientific and Technical Information of China (English)

    Halima; KERDJOUDJ; Cedric; BOURA; Vanessa; MOBY; Dominique; DUMAS; Luc; MARCHAL; Jean-Claude; VOEGEL; Jean-Franois; STOLTZ; Patrick; MENU

    2005-01-01

    1 Introduction Obstructive atherosclerosis vascular disease remains one of the greatest public health threats in the world. Surgical treatment to replace diseased blood vessels is usually done using major human allografts (veins or arteries) or synthetic prosthesis (PTFE, Dacron). However, these substitutes have not a good patency, because of the lack of endothelial cells (ECs) layer, which prevents thrombus formation. The challenge of tissue engineering vessels is to build-up blood/substitute interface nea...

  15. Altered islet morphology but normal islet secretory function in vitro in a mouse model with microvascular alterations in the pancreas.

    Directory of Open Access Journals (Sweden)

    Elena Kostromina

    Full Text Available BACKGROUND: Our previous studies have shown that signal transducer and activator of transcription 3 (STAT3 signaling is important for the development of pancreatic microvasculature via its regulation of vascular endothelial growth factor-A (VEGF-A. Pancreas-specific STAT3-KO mice exhibit glucose intolerance and impaired insulin secretion in vivo, along with microvascular alterations in the pancreas. However, the specific role of STAT3 signaling in the regulation of pancreatic islet development and function is not entirely understood. METHODOLOGY/PRINCIPAL FINDINGS: To investigate the role of STAT3 signaling in the formation and maintenance of pancreatic islets, we studied pancreas-specific STAT3-KO mice. Histological analysis showed that STAT3 deficiency affected pancreatic islet morphology. We found an increased proportion of small-sized islets and a reduced fraction of medium-sized islets, indicating abnormal islet development in STAT3-KO mice. Interestingly, the islet area relative to the whole pancreas area in transgenic and control mice was not significantly different. Immunohistochemical analysis on pancreatic cryosections revealed abnormalities in islet architecture in STAT3-KO mice: the pattern of peripheral distribution of glucagon-positive α-cells was altered. At the same time, islets belonging to different size categories isolated from STAT3-KO mice exhibited normal glucose-stimulated insulin secretion in perifusion experiments in vitro when compared to control mice. CONCLUSIONS: Our data demonstrate that STAT3 signaling in the pancreas is required for normal islet formation and/or maintenance. Altered islet size distribution in the KO mice does not result in an impaired islet secretory function in vitro. Therefore, our current study supports that the glucose intolerance and in vivo insulin secretion defect in pancreas-specific STAT3-KO mice is due to altered microvasculature in the pancreas, and not intrinsic beta-cell function.

  16. Effect of polyethylene glycol grafted onto islet capsules on prevention of splenocyte and cytokine attacks.

    Science.gov (United States)

    Lee, Dong Yun; Nam, Jong Hee; Byun, Youngro

    2004-01-01

    In the graft rejection of transplanted islets, the host's immune cells recognize the islets as antigens, which then stimulate the immune cells to begin the cytokine secretion and also the proliferation of immune cells. To prevent the recognition of islets by the immune cells, we grafted biocompatible polyethylene glycol (PEG) onto the collagen capsule of islets without incurring any changes in the morphology and function of islets. To evaluate the efficiency of PEG grafting, PEG-grafted islets were cultured with splenocytes consisting mainly of lymphocytes and macrophages. A splenocyte proliferation assessment using a BrdU incorporation assay showed that the PEG-grafted islets did not stimulate the splenocytes. In addition, the viability and microorganisms in islet cells of co-cultured PEG-grafted islets were not altered. However, in the co-culture of free islets (control) splenocytes were stimulated; they mainly secreted TNF-alpha and strongly affected the viability and structure of free islets. Furthermore, when islets were treated with the rat recombinant TNF-alpha for 7 days, the viabilities of PEG-grafted and free islets were significantly damaged, although the viability of PEG-grafted islets was higher than that of free islets by nearly three times. These results demonstrate that PEG grafted on the surface of islets could prevent the recognition of islets by splenocytes, but could not completely protect islets from cytokines.

  17. Expanded Hematopoietic Progenitor Cells Reselected for High Aldehyde Dehydrogenase Activity Demonstrate Islet Regenerative Functions.

    Science.gov (United States)

    Seneviratne, Ayesh K; Bell, Gillian I; Sherman, Stephen E; Cooper, Tyler T; Putman, David M; Hess, David A

    2016-04-01

    Human umbilical cord blood (UCB) hematopoietic progenitor cells (HPC) purified for high aldehyde dehydrogenase activity (ALDH(hi) ) stimulate islet regeneration after transplantation into mice with streptozotocin-induced β cell deletion. However, ALDH(hi) cells represent a rare progenitor subset and widespread use of UCB ALDH(hi) cells to stimulate islet regeneration will require progenitor cell expansion without loss of islet regenerative functions. Here we demonstrate that prospectively purified UCB ALDH(hi) cells expand efficiently under serum-free, xeno-free conditions with minimal growth factor supplementation. Consistent with the concept that ALDH-activity is decreased as progenitor cells differentiate, kinetic analyses over 9 days revealed the frequency of ALDH(hi) cells diminished as culture time progressed such that total ALDH(hi) cell number was maximal (increased 3-fold) at day 6. Subsequently, day 6 expanded cells (bulk cells) were sorted after culture to reselect differentiated progeny with low ALDH-activity (ALDH(lo) subset) from less differentiated progeny with high ALDH-activity (ALDH(hi) subset). The ALDH(hi) subset retained primitive cell surface marker coexpression (32.0% ± 7.0% CD34(+) /CD38(-) cells, 37.0% ± 6.9% CD34(+) /CD133(+) cells), and demonstrated increased hematopoietic colony forming cell function compared with the ALDH(lo) subset. Notably, bulk cells or ALDH(lo) cells did not possess the functional capacity to lower hyperglycemia after transplantation into streptozotocin-treated NOD/SCID mice. However, transplantation of the repurified ALDH(hi) subset significantly reduced hyperglycemia, improved glucose tolerance, and increased islet-associated cell proliferation and capillary formation. Thus, expansion and delivery of reselected UCB cells that retain high ALDH-activity after short-term culture represents an improved strategy for the development of cellular therapies to enhance islet regeneration in situ.

  18. The dual role of Fas-ligand as an injury effector and defense strategy in diabetes and islet transplantation.

    Science.gov (United States)

    Pearl-Yafe, Michal; Yolcu, Esma S; Yaniv, Isaac; Stein, Jerry; Shirwan, Haval; Askenasy, Nadir

    2006-02-01

    The exact process that leads to the eruption of autoimmune reactions against beta cells and the evolution of diabetes is not fully understood. Macrophages and T cells may launch an initial immune reaction against the pancreatic islets of Langerhans, provoking inflammation and destructive insulitis. The information on the molecular mechanisms of the emergence of beta cell injury is controversial and points to possibly important roles for the perforin-granzyme, Fas-Fas-ligand (FasL) and tumor-necrosis-factor-mediated apoptotic pathways. FasL has several unique features that make it a potentially ideal immunomodulatory tool. Most important, FasL is selectively toxic to cytotoxic T cells and less harmful to regulatory T cells. This review discusses the intrinsic sensitivity of beta cells to FasL-mediated apoptosis, the conditions that underlie this beta cell sensitivity, and the feasibility of using FasL to arrest autoimmunity and prevent islet allograft rejection. In both the autoimmune and transplant settings, it is imperative to progress from the administration of nonspecific immunosuppressive therapy to the concept of beta-cell-specific immunomodulation. FasL evolves as a prime candidate for antigen-specific immunomodulation.

  19. Correction of diabetic pattern of insulin release from islets of the spiny mouse (Acomys cahirinus) by glucose priming in vitro.

    Science.gov (United States)

    Nesher, R; Abramovitch, E; Cerasi, E

    1985-04-01

    Insulin release kinetics were studied in perifused islets of Langerhans, isolated from mildly hyperglycaemic and from normoglycaemic spiny mice (Acomys cahirinus), a rodent predisposed to develop spontaneously non-ketotic diabetes. In both groups, insulin response to glucose (16.7 mmol/l) was delayed in comparison with that of rat islets, the release kinetics being analogous to that of human Type 2 (non-insulin-dependent) diabetes. Thirty min priming of the isolated Acomys islets with glucose (16.7 mmol/l) resulted in potentiation of the insulin release to a second stimulation. The degree of potentiation decreased exponentially with the time interval between stimulations, showing a t1/2 of 18 min. Induction of potentiation by glucose was time-dependent, giving a maximal effect after 20 min of priming. In addition to overall amplification of the insulin response, priming with glucose accelerated markedly the initial release rates, correcting the dynamics of the response. We conclude that: (1) decreased and delayed insulin secretion is found in Acomys cahirinus before the development of hyperglycaemia; (2) induction of time-dependent potentiation in the islet by priming with glucose corrects the diabetic-type dynamics of insulin release; (3) therefore the deficient insulin release of Acomys is of a functional nature, the mechanism of potentiation bypassing the defect; (4) since insulin release in Acomys resembles that in prediabetic and diabetic man, similar conclusions might apply to the islet dysfunction in Type 2 diabetes.

  20. Allograft Arthrodesis of the Knee in High-grade Osteosarcoma

    Directory of Open Access Journals (Sweden)

    Teng-Le Huang

    2005-09-01

    Conclusion: Due to the high rate of complications in this study, we conclude that allograft arthrodesis should be left as a salvage or “back-up” reconstructive procedure after resection of osteosarcoma around the knee, unless there are special indications for this procedure. We found allograft fracture to be the most common complication.

  1. Veto cell suppression mechanisms in the prevention of allograft rejection

    DEFF Research Database (Denmark)

    Jacobsen, I M; Claesson, Mogens Helweg

    1998-01-01

    Substantial evidence has accumulated to suggest that in the near future implementation of the veto-cell-suppressor concept in the treatment of kidney allograft recipients might lead to the establishment of life-long specific allograft tolerance in the absence of further immunosuppressive therapy....

  2. Use of local allograft irradiation following renal transplantation

    Energy Technology Data Exchange (ETDEWEB)

    Halperin, E.C.; Delmonico, F.L.; Nelson, P.W.; Shipley, W.U.; Cosimi, A.B.

    1984-07-01

    Over a 10 year period, 67 recipients of 71 renal allografts received graft irradiation following the diagnosis of rejection. The majority of kidneys were treated with a total dose of 600 rad, 150 rad per fraction, in 4 daily fractions. Fifty-three kidneys were irradiated following the failure of standard systemic immunosuppression and maximally tolerated antirejection measures to reverse an episode of acute rejection. Twenty-two (42%) of these allografts were noted to have stable (i.e. no deterioration) or improved function 1 month following the treatment with irradiation. Eleven (21%) of these allografts maintained function 1 year following transplantation. Biopsies were obtained of 41 allografts. Of the 24 renal allografts with predominantly cellular rejection, 10 (42%) had the process reversed or stabilized at 1 month following irradiation. Five (21%) of these allografts were functioning at 1 year following irradiation. Rejection was reversed or stabilized in 6 of 17 (35%) allografts at 1 month when the histologic features of renal biopsy suggested predominantly vascular rejection. Local graft irradiation has helped maintain a limited number of allografts in patients whose rejection has failed to respond to systemic immunosuppression. Irradiation may also benefit patients with ongoing rejection in whom further systemic immunosuppression is contra-indicated.

  3. Asymptomatic bacteriuria and urinary tract infections among renal allograft recipients.

    Science.gov (United States)

    Singh, Ramandeep; Geerlings, Suzanne E; Bemelman, Frederike J

    2015-02-01

    Bacteriuria is common among renal allograft recipients. It can be categorized into asymptomatic bacteriuria (ASB) and urinary tract infection (UTI). However, in medical literature, the classifications of bacteriuria are often not clear or ASB is also classified as a UTI. This contributes to difficulties in interpretation of the incidence and risk factors of these two entities. In this review, we describe the epidemiology, risk factors, management and the impact on renal allograft function of these two entities separately according to the recent literature. Risk factors for ASB are not completely comparable to the risk factors of UTIs. Persistent ASB has been associated with development of acute rejection and allograft pyelonephritis. The available data suggest that treatment of ASB is not very effective. Prophylaxis with trimethoprim-sulfamethoxazole does not prevent UTIs such as allograft pyelonephritis. Blood stream infections and emphysematous allograft pyelonephritis are associated with renal allograft loss. ASB is the most common manifestation of bacteriuria after renal transplantation. More effective interventions are needed to prevent bacteriuria. Renal allograft recipients with persistent ASB should be closely monitored since they could be at risk for developing not only UTIs, such as allograft pyelonephritis, but also acute rejection.

  4. Acellular Nerve Allografts in Peripheral Nerve Regeneration: A Comparative Study

    Science.gov (United States)

    Moore, Amy M.; MacEwan, Matthew; Santosa, Katherine B.; Chenard, Kristofer E.; Ray, Wilson Z.; Hunter, Daniel A.; Mackinnon, Susan E.; Johnson, Philip J.

    2011-01-01

    Background Processed nerve allografts offer a promising alternative to nerve autografts in the surgical management of peripheral nerve injuries where short deficits exist. Methods Three established models of acellular nerve allograft (cold-preserved, detergent-processed, and AxoGen® -processed nerve allografts) were compared to nerve isografts and silicone nerve guidance conduits in a 14 mm rat sciatic nerve defect. Results All acellular nerve grafts were superior to silicone nerve conduits in support of nerve regeneration. Detergent-processed allografts were similar to isografts at 6 weeks post-operatively, while AxoGen®-processed and cold-preserved allografts supported significantly fewer regenerating nerve fibers. Measurement of muscle force confirmed that detergent-processed allografts promoted isograft-equivalent levels of motor recovery 16 weeks post-operatively. All acellular allografts promoted greater amounts of motor recovery compared to silicone conduits. Conclusions These findings provide evidence that differential processing for removal of cellular constituents in preparing acellular nerve allografts affects recovery in vivo. PMID:21660979

  5. Incidence, risk factors, and the impact of allograft pyelonephritis on renal allograft function.

    Science.gov (United States)

    Singh, R; Geerlings, S E; Peters-Sengers, H; Idu, M M; Hodiamont, C J; Ten Berge, I J M; Bemelman, F J

    2016-10-01

    The impact of allograft pyelonephritis (AGPN) on renal allograft function is controversial. In this study, we evaluated the incidence, risk factors, and the impact of AGPN on renal allograft function. Retrospective cohort study in adult renal allograft recipients with 1-year follow-up after transplantation (Tx). Renal allograft function was evaluated by estimated glomerular filtration rate (eGFR) (by Modification of Diet in Renal Disease formula) and 24-h urine protein excretion. A total of 431 renal allograft recipients were analyzed; 57 (13.2%) developed AGPN within 1 year after Tx. Median time between Tx and AGPN was 50 days. Risk factors for AGPN were the presence of a urological catheter (odds ratio [OR] = 18.93, 95% confidence interval [CI] = 8.00-44.81, P < 0.001) and preceding asymptomatic bacteriuria (ASB) (OR = 2.16, 95% CI = 1.20-3.90, P = 0.009). In 72.7%, the causative microorganism of ASB was identical to that of the succeeding AGPN episode. Multivariable linear regression analysis showed that experiencing AGPN did not decrease the eGFR (P = 0.61) nor did increased proteinuria (P = 0.29) 1 year after Tx. For the eGFR, an interaction was found between AGPN/bacteriuria (BU) and acute rejection (AR): the group experiencing BU preceding AR had significantly (P < 0.001) lower eGFR compared with the group that experienced only AR (21 mL/min/1.73 m(2) vs. 48 mL/min/1.73 m(2) ), as a result of increased prevalence of combined rejections within the BU group. Indwelling urological catheters and preceding ASB are associated with developing AGPN. An incident of AGPN itself does not impair renal allograft function 1 year after Tx. However, a relevant interaction occurs between BU and AR, in which the sequence of occurrence of these 2 events synergistically impairs the eGFR. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  6. The voltage-gated proton channel Hv1 is expressed in pancreatic islet β-cells and regulates insulin secretion

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Qing [Department of Biophysics, School of Physics Science, The Key Laboratory of Bioactive Materials, Ministry of Education, Nankai University, Tianjin 300071 (China); Che, Yongzhe [School of Medicine, Nankai University, Tianjin 300071 (China); Li, Qiang; Zhang, Shangrong [Department of Biophysics, School of Physics Science, The Key Laboratory of Bioactive Materials, Ministry of Education, Nankai University, Tianjin 300071 (China); Gao, Ying-Tang [Key Laboratory of Artificial Cell, Third Central Clinical College of Tianjin Medical University, Tianjin 300170 (China); Wang, Yifan; Wang, Xudong; Xi, Wang; Zuo, Weiyan [Department of Biophysics, School of Physics Science, The Key Laboratory of Bioactive Materials, Ministry of Education, Nankai University, Tianjin 300071 (China); Li, Shu Jie, E-mail: shujieli@nankai.edu.cn [Department of Biophysics, School of Physics Science, The Key Laboratory of Bioactive Materials, Ministry of Education, Nankai University, Tianjin 300071 (China)

    2015-12-25

    The voltage-gated proton channel Hv1 is a potent acid extruder that participates in the extrusion of the intracellular acid. Here, we showed for the first time, Hv1 is highly expressed in mouse and human pancreatic islet β-cells, as well as β-cell lines. Imaging studies demonstrated that Hv1 resides in insulin-containing granules in β-cells. Knockdown of Hv1 with RNA interference significantly reduces glucose- and K{sup +}-induced insulin secretion in isolated islets and INS-1 (832/13) β-cells and has an impairment on glucose- and K{sup +}-induced intracellular Ca{sup 2+} homeostasis. Our data demonstrated that the expression of Hv1 in pancreatic islet β-cells regulates insulin secretion through regulating Ca{sup 2+} homeostasis.

  7. Exploration of α1-Antitrypsin Treatment Protocol for Islet Transplantation: Dosing Plan and Route of Administration

    OpenAIRE

    Baranovski, Boris M.; Ozeri, Eyal; Shahaf, Galit; Ochayon, David E.; Schuster, Ronen; Bahar, Nofar; Kalay, Noa; Cal, Pablo; Mizrahi, Mark I.; Nisim, Omer; Strauss, Pnina; Schenker, Eran; Eli C Lewis

    2016-01-01

    Lifelong weekly infusions of human α1-antitrypsin (hAAT) are currently administered as augmentation therapy for patients with genetic AAT deficiency (AATD). Several recent clinical trials attempt to extend hAAT therapy to conditions outside AATD, including type 1 diabetes. Because the endpoint for AATD is primarily the reduction of risk for pulmonary emphysema, the present study explores hAAT dose protocols and routes of administration in attempt to optimize hAAT therapy for islet-related inj...

  8. Pancreatic islet amyloidosis, β-cell apoptosis, and α-cell proliferation are determinants of islet remodeling in type-2 diabetic baboons

    Science.gov (United States)

    Guardado-Mendoza, Rodolfo; Davalli, Alberto M.; Chavez, Alberto O.; Hubbard, Gene B.; Dick, Edward J.; Majluf-Cruz, Abraham; Tene-Perez, Carlos E.; Goldschmidt, Lukasz; Hart, John; Perego, Carla; Comuzzie, Anthony G.; Tejero, Maria Elizabeth; Finzi, Giovanna; Placidi, Claudia; La Rosa, Stefano; Capella, Carlo; Halff, Glenn; Gastaldelli, Amalia; DeFronzo, Ralph A.; Folli, Franco

    2009-01-01

    β-Cell dysfunction is an important factor in the development of hyperglycemia of type-2 diabetes mellitus, and pancreatic islet amyloidosis (IA) has been postulated to be one of the main contributors to impaired insulin secretion. The aim of this study was to evaluate the correlation of IA with metabolic parameters and its effect on islets of Langerhans remodeling and relative endocrine-cell volume in baboons. We sequenced the amylin peptide, determined the fibrillogenic propensities, and evaluated pancreatic histology, clinical and biochemical characteristics, and endocrine cell proliferation and apoptosis in 150 baboons with different metabolic status. Amylin sequence in the baboon was 92% similar to humans and showed superimposable fibrillogenic propensities. IA severity correlated with fasting plasma glucose (FPG) (r = 0.662, P < 0.001) and HbA1c (r = 0.726, P < 0.001), as well as with free fatty acid, glucagon values, decreased homeostasis model assessment (HOMA) insulin resistance, and HOMA-B. IA severity was associated with a decreased relative β-cell volume, and increased relative α-cell volume and hyperglucagonemia. These results strongly support the concept that IA and β-cell apoptosis in concert with α-cell proliferation and hypertrophy are key determinants of islets of Langerhans “dysfunctional remodeling” and hyperglycemia in the baboon, a nonhuman primate model of type-2 diabetes mellitus. The most important determinants of IA were age and FPG (R2 = 0.519, P < 0.0001), and different FPG levels were sensitive and specific to predict IA severity. Finally, a predictive model for islet amyloid severity was generated with age and FPG as required variables. PMID:19666551

  9. Sterilization with electron beam irradiation influences the biomechanical properties and the early remodeling of tendon allografts for reconstruction of the anterior cruciate ligament (ACL).

    Science.gov (United States)

    Schmidt, Tanja; Hoburg, Arnd; Broziat, Christine; Smith, Mark D; Gohs, Uwe; Pruss, Axel; Scheffler, Sven

    2012-08-01

    Although allografts for anterior cruciate ligament (ACL) replacement have shown advantages compared to autografts, their use is limited due to the risk of disease transmission and the limitations of available sterilization methods. Gamma sterilization has shown detrimental effects on graft properties at the high doses required for sufficient pathogen inactivation. In our previous in vitro study on human patellar tendon allografts, Electron beam (Ebeam) irradiation showed less detrimental effects compared to gamma sterilization (Hoburg et al. in Am J Sports Med 38(6):1134-1140, 2010). To investigate the biological healing and restoration of the mechanical properties of a 34 kGy Ebeam treated tendon allograft twenty-four sheep underwent ACL replacement with either a 34 kGy Ebeam treated allograft or a non-sterilized fresh frozen allograft. Biomechanical testing of stiffness, ultimate failure load and AP-laxity as well as histological analysis to investigate cell, vessel and myofibroblast-density were performed after 6 and 12 weeks. Native sheep ACL and hamstring tendons (HAT, each n = 9) served as controls. The results of a previous study analyzing the remodeling of fresh frozen allografts (n = 12) and autografts (Auto, n = 18) with the same study design were also included in the analysis. Statistics were performed using Mann-Whitney U test followed by Bonferroni-Holm correction. Results showed significantly decreased biomechanical properties during the early remodeling period in Ebeam treated grafts and this was accompanied with an increased remodeling activity. There was no recovery of biomechanical function from 6 to 12 weeks in this group in contrast to the results observed in fresh frozen allografts and autografts. Therefore, high dose Ebeam irradiation investigated in this paper cannot be recommended for soft tissue allograft sterilization.

  10. OSTEOARTICULAR ALLOGRAFTS IN PAEDIATRIC BONE TUMOR RECONSTRUCTION OF THE KNEE.

    Science.gov (United States)

    Campanacci, D A; Dursky, S; Totti, F; Frenos, F; Scoccianti, G; Beltrami, G; Capanna, R

    2015-01-01

    Osteoarticular allografts represent a reconstructive option after bone tumor resection around the knee in growing children. The major advantage is the chance to preserve the growth plate of the remaining bone, but the disadvantage is the high failure rate eventually requiring definitive prosthetic replacement at skeletal maturity. We retrospectively reviewed 22 patients who underwent osteoarticular allograft reconstructions of the distal femur (16) or proximal tibia (6). There were 12 females and 10 males with an average age at surgery of 11 years (7-15). The diagnosis was osteosarcoma in 19 cases and Ewing sarcoma in 3. All patients underwent pre- and post-operative chemotherapy. At an average follow-up of 103 months (12-167), 18 patients (82%) were alive and 4 had died (18%). We observed 10 allograft failures requiring prosthetic replacement, 6 in distal femur and 4 in proximal tibia reconstructions. At last follow-up 8 allografts (36%) were still in place. Overall allograft survival was 79.6% at five and 45.8% at ten years. In distal femur, allograft survival was 86.2% at five and 59.1% at ten years. In proximal tibia, allograft survival was 62.5% at 5 years and 31.2% at 67 months. Average limb shortening was 3 cm (0- 5) in 8 patients with the allograft still in situ and 2 cm (0-4) in 10 patients after prosthetic replacement. Average MSTS functional score of the whole series was 25 (83.7%). The MSTS score of patients after revision with prosthetic replacement was 24 (80%) while patients who still had the allograft retained had an average MSTS scores of 26.8 (89.3%). In conclusion, osteoarticular allograft reconstruction of the knee after bone tumor resection in pediatric age can be considered a temporary solution with the aim to limit limb length discrepancy before definitive prosthetic replacement after skeletal maturity.

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

    DEFF Research Database (Denmark)

    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...

  12. Chemical sterilization of allograft dermal tissues.

    Science.gov (United States)

    Phipps, Abigail; Vaynshteyn, Edward; Kowalski, John B; Ngo, Manh-Dan; Merritt, Karen; Osborne, Joel; Chnari, Evangelia

    2017-08-10

    Common terminal sterilization methods are known to alter the natural structure and properties of soft tissues. One approach to providing safe grafts with preserved biological properties is the combination of a validated chemical sterilization process followed by an aseptic packaging process. This combination of processes is an accepted method for production of sterile healthcare products as described in ANSI/AAMI ST67:2011. This article describes the validation of the peracetic acid and ethanol-based (PAAE) chemical sterilization process for allograft dermal tissues at the Musculoskeletal Transplant Foundation (MTF, Edison, NJ). The sterilization capability of the PAAE solution used during routine production of aseptically processed dermal tissue forms was determined based on requirements of relevant ISO standards, ISO 14161:2009 and ISO 14937:2009. The resistance of spores of Bacillus subtilis, Clostridium sporogenes, Mycobacterium terrae, Pseudomonas aeruginosa, Enterococcus faecium, and Staphylococcus aureus to the chemical sterilization process employed by MTF was determined. Using a worst-case scenario testing strategy, the D value was calculated for the most resistant microorganism, Bacillus. The 12D time parameter determined the minimum time required to achieve a SAL of 10(-6). Microbiological performance qualification demonstrated a complete kill of 10(6) spores at just a quarter of the full cycle time. The validation demonstrated that the PAAE sterilization process is robust, achieves sterilization of allograft dermal tissue to a SAL 10(-6), and that in combination with aseptic processing secures the microbiological safety of allograft dermal tissue while avoiding structural and biochemical tissue damage previously observed with other sterilization methods such as ionizing irradiation.

  13. Teucrium polium complex with molybdate enhance cultured islets secretory function.

    Science.gov (United States)

    Mohseni Salehi Monfared, Seyed Sajad; Pournourmohammadi, Shirin

    2010-02-01

    Islet transplantation has become a promising treatment in the therapy of type 1 diabetes. Its function improvement, after isolation and before transplantation, is crucial because of their loss both in number and function of islets after isolation procedures. Trace elements sodium orthovanadate (SOV) and sodium molybdate (SM), as well as medicinal plant Teucrium polium L. (TP), showed and possessed high beneficial antioxidative potential and even hypoglycemic properties via their effect on islets. We evaluated the effect of these components in combination on cultured islet function in order to improve pancreatic islet transplantation. Rat pancreatic islets were cultured for 24 h then incubated with different concentrations of TP (0.01 and 0.1 mg/mL) alone and in combination with SOV (1 mM) or SM (1 mM). Insulin concentration in buffer media was measured as islet secretory function. Administration of TP (0.01 mg/mL), SM, and SOV alone or in combination with each other significantly increased insulin secretion at high glucose concentration (16.7 mM); insulin secretion was significantly greater in the group containing both TP and SM than other treated groups (p molybdate with TP could improve islet cells function before transplantation.

  14. Modifying cyclosporine associated renal allograft dysfunction

    Directory of Open Access Journals (Sweden)

    Mohapatra N

    2009-01-01

    Full Text Available Transplantation is accepted therapy for chronic kidney disease. However the essential immuno-suppressive agents for graft survival have their own side-effects. Renal biopsy is a reliable tool for diagnosing cyclosporine (CsA nephrotoxicity. To present our observations on CsA toxicity in renal allograft biopsies, we studied prospectively 207 renal allograft biopsies performed for graft dysfunction as per Ahmedabad Tole-rance Induction Protocol (ATIP and compared them to 50 controls from January to October 2007. The ATIP comprised donor specific leucocyte infusions, low dose target specific irradiation; non-myeloablative condi-tioning with Anti-T ± B cell antibodies followed by intraportal administration of cultured donor bone marrow (BM ± adipose tissue derived mesenchymal stem cells. Renal transplantation was performed following nega-tive lymphocytotoxicity cross-matching. The post-transplant immunosuppressive agents included CsA 2.5 ± 0.5 mg/kg BW/day and prednisone 0.2 mg/kg BW/day. The controls were transplanted using standard triple immunosuppressive agents including CsA 5 ± 1 mg/Kg BW/day, prednisone 0.6 mg/kg BW/day, and MMF/ Azathioprine. The Institutional Review Board approved the ATIP. The biopsies were categorized into 2 groups; group A (N=97: performed < 6 months, group B (N= 160, > 6 months posttransplant. Acute CsA toxicity was observed in group A: 2.5% ATIP and 11.1% controls; group B: 16.2% ATIP and 8.8% controls. Chronic CsA toxicity was observed in group B: 10.8 % ATIP and 17.6 % controls. Acute toxicity was more in the ATIP, while chronic toxicity was more in the controls. CsA doses were reduced post-biopsy and resulted in improved graft function evaluated by serum creatinine. We conclude that CsA nephrotoxicity evaluated by allograft biopsy resulted in allograft function recovery by decreasing the cyclosporine dose, and the ATIP decreased the incidence of CsA nephrotoxicity.

  15. Evaluation of new antibiotic cocktails against contaminating bacteria found in allograft tissues.

    Science.gov (United States)

    Serafini, Agnese; Riello, Erika; Trojan, Diletta; Cogliati, Elisa; Palù, Giorgio; Manganelli, Riccardo; Paolin, Adolfo

    2016-12-01

    Contamination of retrieved tissues is a major problem for allograft safety. Consequently, tissue banks have implemented decontamination protocols to eliminate microorganisms from tissues. Despite the widespread adoption of these protocols, few comprehensive studies validating such methods have been published. In this manuscript we compare the bactericidal activity of different antibiotic cocktails at different temperatures against a panel of bacterial species frequently isolated in allograft tissues collected at the Treviso Tissue Bank Foundation, a reference organization of the Veneto Region in Italy that was instituted to select, recover, process, store and distribute human tissues. We were able to identify at least two different formulations capable of killing most of the bacteria during prolonged incubation at 4 °C.

  16. Interest of Polyelectrolyte Multilayer thin Films in Tissue Engineering:Application to Vascular Allograft

    Institute of Scientific and Technical Information of China (English)

    Halima KERDJOUDJ; Cedric BOURA; Vanessa MOBY; Dominique DUMAS; Luc MARCHAL; Jean-Claude VOEGEL; Jean-Fran(c)ois STOLTZ; Patrick MENU

    2005-01-01

    @@ 1 Introduction Obstructive atherosclerosis vascular disease remains one of the greatest public health threats in the world. Surgical treatment to replace diseased blood vessels is usually done using major human allografts (veins or arteries) or synthetic prosthesis ( PTFE, Dacron). However, these substitutes have not a good pateney, because of the lack of endothelial cells (ECs) layer, which prevents thrombus formation. The challenge of tissue engineering vessels is to build-up blood/substitute interface near native vessels.In order to improve ECs adhesion, it is necessary to precoat the intra-luminal vessel. Recently, a new surface modification technique arose, based on the alternate adsorption of oppositely charged polyelectrolytes. Our objective was to favour the endothelialization of the cryo-preserved allografts, treated with a thin polyelectrolyte multilayered film, made of PSS (poly (sodium-4-styrenesulfonate) ) or PAH (poly (allylamine hydrochloride) ).

  17. Treatment with insulin analog X10 and IGF-1 increases growth of colon cancer allografts.

    Directory of Open Access Journals (Sweden)

    Henning Hvid

    Full Text Available Obesity and type 2 diabetes are associated with an increased risk for development of certain forms of cancer, including colon cancer. The publication of highly controversial epidemiological studies in 2009 raised the possibility that use of the insulin analog glargine increases this risk further. However, it is not clear how mitogenic effects of insulin and insulin analogs measured in vitro correlate with tumor growth-promoting effects in vivo. The aim of this study was to examine possible growth-promoting effects of native human insulin, insulin X10 and IGF-1, which are considered positive controls in vitro, in a short-term animal model of an obesity- and diabetes-relevant cancer. We characterized insulin and IGF-1 receptor expression and the response to treatment with insulin, X10 and IGF-1 in the murine colon cancer cell line (MC38 cells in vitro and in vivo. Furthermore, we examined pharmacokinetics and pharmacodynamics and monitored growth of MC38 cell allografts in mice with diet-induced obesity treated with human insulin, X10 and IGF-1. Treatment with X10 and IGF-1 significantly increased growth of MC38 cell allografts in mice with diet-induced obesity and we can therefore conclude that supra-pharmacological doses of the insulin analog X10, which is super-mitogenic in vitro and increased the incidence of mammary tumors in female rats in a 12-month toxicity study, also increase growth of tumor allografts in a short-term animal model.

  18. ACL reconstruction with BPTB autograft and irradiated fresh frozen allograft

    Institute of Scientific and Technical Information of China (English)

    Kang SUN; Shao-qi TIAN; Ji-hua ZHANG; Chang-suo XIA; Cai-long ZHANG; Teng-bo YU

    2009-01-01

    Objective: To analyze the clinical outcomes of arthroscopic anterior cruciate ligament (ACL) reconstruction with irradiated bone-patellar tendon-bone (BPTB) allograft compared with non-irradiated allograft and autograft. Methods: All BPTB allografts were obtained from a single tissue bank and the irradiated allografts were sterilized with 2.5 mrad of irradiation prior to distribution. A total of 68 patients undergoing arthroscopic ACL reconstruction were prospectively randomized consecutively into one of the two groups (autograft and irradiated allograft groups). The same surgical technique was used in all operations done by the same senior surgeon. Before surgery and at the average of 31 months of follow-up (ranging from 24 to 47 months), patients were evaluated by the same observer according to objective and subjective clinical evaluations. Results: Of these patients, 65 (autograft 33, irradiated allograft 32) were available for full evaluation. When the irradiated allograft group was compared to the autografi group at the 31-month follow-up by the Lachman test, the anterior drawer test (ADT), the pivot shift test, and KT-2000 arthrometer test, statistically significant differences were found. Most importantly, 87.8% of patients in the autograft group and just only 31.3% in the irradiated allograft group had a side-to-side difference of less than 3 mm according to KT-2000. The failure rate of the ACL reconstruction with irradiated allograft (34.4%) was higher than that with autograft (6.1%). The anterior and rotational stabilities decreased significantly in the irradiated allograft group. According to the overall International Knee Docu-mentation Committee (IKDC), functional and subjective evaluations, and activity level testing, no statistically significant dif-ferences were found between the two groups. Besides, patients in the irradiated allograft group had a shorter operation time and a longer duration of postoperative fever. When the patients had a fever

  19. 人胚胎干细胞体外定向诱导分化胰岛细胞移植治疗非肥胖联合免疫缺陷糖尿病小鼠%Pancreatic islets differentiated from human embryonic stem cells correct hyperglycemia in non-obese diabetic /severe combined immunodeficient mice

    Institute of Scientific and Technical Information of China (English)

    华秀峰; 孙强; 王延伟; 丛晋; 刘芙君; 孟晓梅; 李华峰; 靳少华; 王海燕; 李建远

    2013-01-01

    目的 探讨人胚胎干细胞(hESs)体外定向诱导分化胰岛细胞移植治疗非肥胖联合免疫缺陷(NOD/SCID)糖尿病小鼠的可行性.方法 体外分四阶段诱导hESs定向分化为胰岛细胞:第一阶段,予以活化素A(activin A)、渥曼青霉素(wortmannin)诱导分化形成定型内胚层;第二阶段,予以全反式维甲酸(RA)、NOGGIN、碱性成纤维细胞生长因子(bFGF)诱导胰腺细胞定向分化;第三阶段,予以表皮生长因子(EGF)扩增胰腺祖细胞;第四阶段,予以尼克酰胺(nicotinamide)、唾液素4(exendin-4)、bFGF及骨形成蛋白(BMP4)促进胰岛细胞成熟;观察诱导各阶段细胞形态变化、免疫荧光鉴定胰十二指肠同源异型盒基因(PDX-1)、胰高糖素、胰岛素、C肽、葡萄糖转运子2(Glut-2)的表达;四阶段分化成熟的胰岛细胞体外检测胰岛素释放反应并植入链脲菌素(STZ)诱导形成的NOD/SCID糖尿病小鼠一侧附睾脂肪垫内,观察血糖变化.结果 诱导第四阶段14天时hESs出现胰高糖素荧光表达;20天时细胞出现PDX-1和C肽共表达;22天形成的成熟胰岛细胞出现Glut-2和胰岛素的阳性表达;流式鉴定胰岛素阳性细胞占17.1%,C肽阳性细胞占3.8%;体外检测有葡萄糖刺激的胰岛素释放反应.分化成熟胰岛细胞约(3~5)×106植入NOD/SCID糖尿病小鼠体内可以逆转其高血糖至少8周.结论 体外定向诱导hESs分化形成的胰岛细胞植入NOD/SCID糖尿病小鼠附睾脂肪垫内可以逆转其高血糖.%Objective To investigate whether pancreatic progenitors differentiated from human embryonic stem (hES) cells could correct hyperglycemia in non-obese diabetic(NOD)/severe combined immunodeficient(SCID) mice or not. Methods Pancreatic islets derived from hES cells line YT1 according to the optimized four-stage differentiation protocol in a chemical-defined culture system were observed. In the first stage.activin A and wortmannin were utilized to induce definitive endoderm

  20. The proapoptotic BH3-only proteins Bim and Puma are downstream of endoplasmic reticulum and mitochondrial oxidative stress in pancreatic islets in response to glucotoxicity.

    Science.gov (United States)

    Wali, J A; Rondas, D; McKenzie, M D; Zhao, Y; Elkerbout, L; Fynch, S; Gurzov, E N; Akira, S; Mathieu, C; Kay, T W H; Overbergh, L; Strasser, A; Thomas, H E

    2014-03-13

    Apoptosis of pancreatic beta cells is a feature of type 2 diabetes and its prevention may have therapeutic benefit. High glucose concentrations induce apoptosis of islet cells, and this requires the proapoptotic Bcl-2 homology domain 3 (BH3)-only proteins Bim and Puma. We studied the stress pathways induced by glucotoxicity in beta cells that result in apoptosis. High concentrations of glucose or ribose increased expression of the transcription factor CHOP (C/EBP homologous protein) but not endoplasmic reticulum (ER) chaperones, indicating activation of proapoptotic ER stress signaling. Inhibition of ER stress prevented ribose-induced upregulation of Chop and Puma mRNA, and partially protected islets from glucotoxicity. Loss of Bim or Puma partially protected islets from the canonical ER stressor thapsigargin. The antioxidant N-acetyl-cysteine also partially protected islets from glucotoxicity. Islets deficient in both Bim and Puma, but not Bim or Puma alone, were significantly protected from killing induced by the mitochondrial reactive oxygen species donor rotenone. Our data demonstrate that high concentrations of glucose induce ER and oxidative stress, which causes cell death mediated by Bim and Puma. We observed significantly higher Bim and Puma mRNA in islets of human donors with type 2 diabetes. This indicates that inhibition of Bim and Puma, or their inducers, may prevent beta-cell destruction in type 2 diabetes.

  1. The unique hypusine modification of eIF5A promotes islet beta cell inflammation and dysfunction in mice.

    Science.gov (United States)

    Maier, Bernhard; Ogihara, Takeshi; Trace, Anthony P; Tersey, Sarah A; Robbins, Reiesha D; Chakrabarti, Swarup K; Nunemaker, Craig S; Stull, Natalie D; Taylor, Catherine A; Thompson, John E; Dondero, Richard S; Lewis, Eli C; Dinarello, Charles A; Nadler, Jerry L; Mirmira, Raghavendra G

    2010-06-01

    In both type 1 and type 2 diabetes, pancreatic islet dysfunction results in part from cytokine-mediated inflammation. The ubiquitous eukaryotic translation initiation factor 5A (eIF5A), which is the only protein to contain the amino acid hypusine, contributes to the production of proinflammatory cytokines. We therefore investigated whether eIF5A participates in the inflammatory cascade leading to islet dysfunction during the development of diabetes. As described herein, we found that eIF5A regulates iNOS levels and that eIF5A depletion as well as the inhibition of hypusination protects against glucose intolerance in inflammatory mouse models of diabetes. We observed that following knockdown of eIF5A expression, mice were resistant to beta cell loss and the development of hyperglycemia in the low-dose streptozotocin model of diabetes. The depletion of eIF5A led to impaired translation of iNOS-encoding mRNA within the islet. A role for the hypusine residue of eIF5A in islet inflammatory responses was suggested by the observation that inhibition of hypusine synthesis reduced translation of iNOS-encoding mRNA in rodent beta cells and human islets and protected mice against the development of glucose intolerance the low-dose streptozotocin model of diabetes. Further analysis revealed that hypusine is required in part for nuclear export of iNOS-encoding mRNA, a process that involved the export protein exportin1. These observations identify the hypusine modification of eIF5A as a potential therapeutic target for preserving islet function under inflammatory conditions.

  2. The unique hypusine modification of eIF5A promotes islet β cell inflammation and dysfunction in mice

    Science.gov (United States)

    Maier, Bernhard; Ogihara, Takeshi; Trace, Anthony P.; Tersey, Sarah A.; Robbins, Reiesha D.; Chakrabarti, Swarup K.; Nunemaker, Craig S.; Stull, Natalie D.; Taylor, Catherine A.; Thompson, John E.; Dondero, Richard S.; Lewis, Eli C.; Dinarello, Charles A.; Nadler, Jerry L.; Mirmira, Raghavendra G.

    2010-01-01

    In both type 1 and type 2 diabetes, pancreatic islet dysfunction results in part from cytokine-mediated inflammation. The ubiquitous eukaryotic translation initiation factor 5A (eIF5A), which is the only protein to contain the amino acid hypusine, contributes to the production of proinflammatory cytokines. We therefore investigated whether eIF5A participates in the inflammatory cascade leading to islet dysfunction during the development of diabetes. As described herein, we found that eIF5A regulates iNOS levels and that eIF5A depletion as well as the inhibition of hypusination protects against glucose intolerance in inflammatory mouse models of diabetes. We observed that following knockdown of eIF5A expression, mice were resistant to β cell loss and the development of hyperglycemia in the low-dose streptozotocin model of diabetes. The depletion of eIF5A led to impaired translation of iNOS-encoding mRNA within the islet. A role for the hypusine residue of eIF5A in islet inflammatory responses was suggested by the observation that inhibition of hypusine synthesis reduced translation of iNOS-encoding mRNA in rodent β cells and human islets and protected mice against the development of glucose intolerance the low-dose streptozotocin model of diabetes. Further analysis revealed that hypusine is required in part for nuclear export of iNOS-encoding mRNA, a process that involved the export protein exportin1. These observations identify the hypusine modification of eIF5A as a potential therapeutic target for preserving islet function under inflammatory conditions. PMID:20501948

  3. Autograft versus allograft in anterior cruciate ligament reconstruction

    Science.gov (United States)

    Kan, Shun-Li; Yuan, Zhi-Fang; Ning, Guang-Zhi; Yang, Bo; Li, Hai-Liang; Sun, Jing-Cheng; Feng, Shi-Qing

    2016-01-01

    Abstract Background: Anterior cruciate ligament (ACL) reconstruction is considered as the standard surgical procedure for the treatment of ACL tear. However, there is a crucial controversy in terms of whether to use autograft or allograft in ACL reconstruction. The purpose of this meta-analysis is to compare autograft with allograft for patients undergoing ACL reconstruction. Methods: PubMed, EMBASE, and the Cochrane Library were searched for randomized controlled trials that compared autograft with allograft in ACL reconstruction up to January 31, 2016. The relative risk or mean difference with 95% confidence interval was calculated using either a fixed- or random-effects model. The risk of bias for individual studies according to the Cochrane Handbook. The trial sequential analysis was used to test the robustness of our findings and get more conservative estimates. Results: Thirteen trials were included, involving 1636 participants. The results of this meta-analysis indicated that autograft brought about lower clinical failure, better overall International Knee Documentation Committee (IKDC) level, better pivot-shift test, better Lachman test, greater Tegner score, and better instrumented laxity test (P allograft. Autograft was not statistically different from allograft in Lysholm score, subjective IKDC score, and Daniel 1-leg hop test (P > 0.05). Subgroup analyses demonstrated that autograft was superior to irradiated allograft for patients undergoing ACL reconstruction in clinical failure, Lysholm score, pivot-shift test, Lachman test, Tegner score, instrumented laxity test, and subjective IKDC score (P allograft. Conclusions: Autograft is superior to irradiated allograft for patients undergoing ACL reconstruction concerning knee function and laxity, but there are no significant differences between autograft and nonirradiated allograft. However, our results should be interpreted with caution, because the blinding methods were not well used. PMID

  4. The number of regulatory T cells in transbronchial lung allograft biopsies is related to FoxP3 mRNA levels in bronchoalveolar lavage fluid and to the degree of acute cellular rejection

    DEFF Research Database (Denmark)

    Krustrup, Dorrit; Madsen, Caroline B; Iversen, Martin;

    2013-01-01

    The transcription factor Forkhead Box P3 (FoxP3) is a marker of regulatory T cells (Tregs) - a subset of T cells known to suppress a wide range of immune responses. These cells are considered to be pivotal for the induction of tolerance to donor antigens in human allografts. We aimed to correlate...... the number of lymphocytes expressing FoxP3 in transbronchial biopsies from lung allografts with the FoxP3 expression in bronchoalveolar lavage fluid (BALF). In addition, we aimed to correlate the number of FoxP3+ cells in transbronchial biopsies with the degree of acute cellular rejection in lung allografts....

  5. 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...

  6. High-pressure saline washing of allografts reduces bacterial contamination.

    Science.gov (United States)

    Hirn, M Y; Salmela, P M; Vuento, R E

    2001-02-01

    60 fresh-frozen bone allografts were contaminated on the operating room floor. No bacterial growth was detected in 5 of them after contamination. The remaining 55 grafts had positive bacterial cultures and were processed with three methods: soaking in saline, soaking in antibiotic solution or washing by high-pressure saline. After high-pressure lavage, the cultures were negative in three fourths of the contaminated allografts. The corresponding figures after soaking grafts in saline and antibiotic solution were one tenth and two tenths, respectively. High-pressure saline cleansing of allografts can be recommended because it improves safety by reducing the superficial bacterial bioburden.

  7. Percutaneous fusion of lumbar facet with bone allograft

    Directory of Open Access Journals (Sweden)

    Félix Dolorit Verdecia

    2015-03-01

    Full Text Available OBJECTIVE: To assess the evolution of the cases treated with percutaneous facet fusion with bone allograft in lumbar facet disease. METHOD: Between 2010 and 2014, 100 patients (59 women and 41 men diagnosed with lumbar facet disease underwent surgery. RESULTS: The lumbar facet fusion with bone allograft shows good clinical results, is performed on an outpatient basis, and presents minimal complications and rapid incorporation of the patient to the activities of daily living. CONCLUSIONS: The lumbar facet fusion with bone allograft appears to be an effective treatment for lumbar facet disease.

  8. Significance of Anti-HLA Antibodies on Adult and Pediatric Heart Allograft Outcomes

    Science.gov (United States)

    Mangiola, Massimo; Marrari, Marilyn; Feingold, Brian; Zeevi, Adriana

    2017-01-01

    As methods for human leukocyte antigens (HLA) antibody detection have evolved and newer solid phase assays are much more sensitive, the last 15 years has seen a renewed focus on the importance of HLA antibodies in solid organ transplant rejection. However, there is still much controversy regarding the clinical significance of antibody level as depicted by the mean fluorescence intensity of a patient’s neat serum. Emerging techniques, including those that identify antibody level and function, show promise for the detection of individuals at risk of allograft rejection, determination of the effectiveness of desensitization prior to transplant, and for monitoring treatment of rejection. Here, we review current publications regarding the relevance of donor-specific HLA antibodies (DSA) in adult and pediatric heart transplantation (HT) with graft survival, development of antibody-mediated rejection and cardiac allograft vasculopathy (CAV). The negative impact of DSA on patient and allograft survival is evident in adult and pediatric HT recipients. Many questions remain regarding the most appropriate frequency of assessment of pre- and posttransplant DSA as well as the phenotype of DSA memory vs. true de novo antibody using large multicenter adult and pediatric cohorts and state-of-the-art methodologies for DSA detection and characterization. PMID:28191005

  9. Allograft inflammatory factor-1 in the pathogenesis of bleomycin-induced acute lung injury.

    Science.gov (United States)

    Nagahara, Hidetake; Yamamoto, Aihiro; Seno, Takahiro; Obayashi, Hiroshi; Kida, Takashi; Nakabayashi, Amane; Kukida, Yuji; Fujioka, Kazuki; Fujii, Wataru; Murakami, Ken; Kohno, Masataka; Kawahito, Yutaka

    2016-02-01

    Allograft inflammatory factor-1 (AIF-1) is a protein expressed by macrophages infiltrating the area around the coronary arteries of rats with an ectopic cardiac allograft. Some studies have shown that expression of AIF-1 increased in a mouse model of trinitrobenzene sulfonic acid-induced acute colitis and in acute cellular rejection of human cardiac allografts. These results suggest that AIF-1 is related to acute inflammation. The current study used bleomycin-induced acute lung injury to analyze the expression of AIF-1 and to examine its function in acute lung injury. Results showed that AIF-1 was significantly expressed in lung macrophages and increased in bronchoalveolar lavage fluid from mice with bleomycin-induced acute lung injury in comparison to control mice. Recombinant AIF-1 increased the production of IL-6 and TNF-α from RAW264.7 (a mouse macrophage cell line) and primary lung fibroblasts, and it also increased the production of KC (CXCL1) from lung fibroblasts. These results suggest that AIF-1 plays an important role in the mechanism underlying acute lung injury.

  10. Influence of heme oxygenase-1 gene transfer on the viability and function of rat islets in in vitro culture

    Institute of Scientific and Technical Information of China (English)

    Xiao-Bo Chen; Yong-Xiang Li; Yang Jiao; Wei-Ping Dong; Ge Li; Jing Chen; Jian-Ming Tan

    2007-01-01

    AIM:To ifivestigate the influence of heme oxygenase-1(HO-1)gene transfer on the viability and function of cultured rat islets in vitro.METHODS:Islets were isolated from the pancreata of Sprague-Dawley rats by intraductal collagenase digestion,and purified by discontinuous Ficoll density gradient centrifugation.Purified rat islets were transfected with adenoviral vectors containing human HO-1 gene(Ad-HO-1)or enhanced green fluorescent protein gene(Ad-EGFP),and then cultured for seven days.Transfection was confirmed by fluorescence microscopy and Western blot.Islet viability was evaluated by acridine orange/propidium iodide fluorescent staining.Glucose-stimulated insulin release was detected using insulin radioimmunoassay kits and was used to assess the function of islets.Stimulation index (SI)was calculated by dividing the insulin release upon high glucose stimulation by the insulin release upon low glucose stimulation.RESULTS:After seven days culture,the viability of cultured rat islets decreased significantly(92% ± 6% vs 52% ± 13%,P < 0.05),and glucose-stimulated insulin release also decreased significantly(6.47 ± 0.55 mIU/L/30IEQ vs 4.57 ± 0.40 mIU/L/30IEQ,14.93 ± 1.17mIU/L/30IEQ vs 9.63 ± 0.71 mIU/L/30IEQ,P < 0.05).Transfection of rat Islets with adenoviral vectors at an MOI of 20 was efficient,and did not impair islet function.At 7 d post-transfection,the viability of Ad-HO-1 transfected islets was higher than that of control islets (71% ± 15% vs 52% ± 13%,P < 0.05).There was no significant difference in insulin release upon low glucose stimulation(2.8 mmol/L)among Ad-HO-1 transfected group,Ad-EGFP transfected group,and control group(P > 0.05),while when stimulated by high glucose(16.7 mmol/L)solution,insulin release in Ad-HO-1 transfected group was significantly higher than that in Ad-EGFP transfected group and control group,respectively(12.50 ± 2.17 mIU/L/30IEQ vs 8.87 ± 0.65 mIU/L/30IEQ;12.50 ± 2.17 mIU/L/30IEQ vs 9.63 ± 0.71 mIU/L/30IEQ

  11. Photoacoustic imaging of angiogenesis in subdermal islet transplant sites

    Science.gov (United States)

    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.

  12. Hexose metabolism in pancreatic islets: the Pasteur effect.

    Science.gov (United States)

    Malaisse, W J; Rasschaert, J; Zähner, D; Sener, A

    1988-02-01

    In rat pancreatic islets, hypoxia severely decreased both the oxidation of D-[U-14C]glucose and the release of insulin evoked by D-glucose. The production of [14C]lactate was increased in the hypoxic islets, the relative magnitude of such an increment being greater at low (2.8 mM) than high (8.3 and 16.7 mM) D-glucose concentrations. Hypoxia increased the detritiation of D-[5-3H]glucose at low glucose concentration (2.8 mM), failed to affect 3H2O production at an intermediate glucose level (8.3 mM), and inhibited the utilization of D-[5-3H]glucose at a higher hexose concentration (16.7 mM). In tumoral islet cells (RINm5F line) exposed to 16.7 mM D-glucose, hypoxia decreased D-[U-14C]glucose oxidation to the same extent as in normal islet cells, but increased the production of [14C]lactate and 3H2O to a greater extent than in normal islets. These findings indicate that the Pasteur effect is operative in islet cells. The experimental data also suggest that, under normal conditions of oxygenation, high concentrations of D-glucose lead to both activation of phosphofructokinase and stimulation of mitochondrial oxidative events in normal, but not tumoral, islet cells.

  13. Comparison between cryopreserved and glycerol-preserved allografts in a partial-thickness porcine wound model.

    Science.gov (United States)

    Yoon, Cheonjae; Lim, Kihwan; Lee, Sungjun; Choi, Yanghwan; Choi, Youngwhan; Lee, Jungsuk

    2016-03-01

    Human skin allografts are one of the best temporary biological coverings for severely burned patients. Cryopreserved (CPA) and glycerol-preserved (GPA) allografts are the most widely used types. This study compared the allograft efficiency of both preservation methods under the same conditions. To simulate actual clinical conditions, we used a porcine wound model. In addition, we evaluated the macroscopic and microscopic scoring of graft performance for each method. Porcine cadaver skin 1 mm thick was obtained from one pig. Cryopreserved skin cell viability was 20.8 %, glycerol-preserved skin was 9.08 %, and fresh skin was 58.6 %. We made ten partial-thickness wounds each in two pigs. The take rates on day 2 were 96.23 and 82.65 % in the GPA and CPA group (both n = 9), respectively. After 1 week, the take rates of both groups were nearly equal. The removal rate at week 5 was 98.87 and 94.41 % in the GPA and CPA group, respectively. On microscopic findings at week 2, inflammation was greater in the CPA group. Other findings such as fibroblast hyperplasia and neovascularization were not significantly different between both groups. At week 5, the score of collagen fiber synthesis was 2.67 ± 0.47 and 2.33 ± 0.47 in the GPA and CPA group, respectively. The epidermal-dermal junction was 2.22 ± 0.79 and 2.00 ± 0.47 in the GPA and CPA group, respectively. These findings suggest that wound healing takes longer in the CPA group. The preservation method of allografts is not a absolute factor in the wound healing process in this wound model.

  14. Renal allograft rejection: examination of delayed differentiation of Treg and Th17 effector T cells.

    Science.gov (United States)

    Pekalski, Marcin; Jenkinson, Sarah E; Willet, Joseph D P; Poyner, Elizabeth F M; Alhamidi, Abdulaziz H; Robertson, Helen; Ali, Simi; Kirby, John A

    2013-03-01

    Antigen presentation after kidney transplantation occurs in lymphoid tissues remote from the allograft, with activated T cells then migrating towards the graft. This study examined the possibility that these activated T cells can differentiate to acquire Th17 or Treg phenotypes after a time consistent with their arrival within renal allograft tissues. An immunocytochemical study was performed to demonstrate the response to intragraft TGF-β and the phenotype of lymphoid cells within rejecting human renal allograft tissue. A series of in vitro experiments was then performed to determine the potential to induce these phenotypes by addition of appropriate cytokines 3days after initial T cell activation. During renal allograft rejection there was a strong response to TGF-β, and both FOXP3 and IL-17A were expressed by separate lymphoid cells in the graft infiltrate. FOXP3 could be induced to high levels by the addition of TGF-β1 3days after the initiation of allogeneic mixed leukocyte culture. This Treg marker was enriched in the sub-population of T cells expressing the cell-surface αE(CD103)β7 integrin. The RORγt transcription factor and IL-17A were induced 3days after T cell activ