Islet Transplantation in the UK.

This commentary outlines the development and current status of the UK Islet Transplant Programme in the UK. The author makes the case that it is now time for similar fully funded beta-cell programmes to be made available in many other countries as well.

Effect of Etanercept Concentration on Human Islet Integrity.

BACKGROUND: Etanercept is widely used as an antiinflammatory drug to improve engraftment after intraportal islet transplantation. In contrast to other immunosuppressive agents, very little is known about detrimental effects of etanercept on islets. The aim of this pilot study was to define the toxic range of etanercept. METHODS: Human islets isolated from 8 donors were cultured for 4-5 days at 37°C in culture medium supplemented with etanercept at concentrations from 2.5 to 40 µg/mL, corresponding to potential in vivo levels within the portal vein. After culture, islet equivalent (IEQ) yield, fragmentation index (islet number/IEQ), purity, viability, and stimulated insulin release (2 vs 20 mmol/L) were assessed and normalized to islets before culture. RESULTS: Yield (73 ± 8%) and viability (91 ± 4%) were highest with 5 µg/mL etanercept. Islet loss was evident when etanercept was ≥10 µg/mL (55 ± 7%; P < .05 vs control). Fragmentation (154 ± 34%; P < .05) was markedly increased and viability (81 ± 4%, P < .05) markedly decreased with etanercept >10 µg/mL. The accumulation of cell debris at concentrations ≥20 µg/mL resulted in a significant reduction of islet purity (84 ± 3%; P < .05). Etanercept did not interfere with stimulated insulin secretion at concentrations ≤10 µg/mL. The maximum stimulation index was noted at 2.5 µg/mL (1.8 ± 0.1). CONCLUSIONS: Etanercept is tolerated by isolated human islets at concentrations <10 µg/mL. Our data suggest that the tight range between benefit and toxicity should be considered for dosage and administration of etanercept.

Hypoxia-Induced Damage in Human Islets Is Reduced With the Use of Mesenchymal Stem Cell-Preconditioned Medium.

BACKGROUND: Mesenchymal stem cells (MSCs) are protective for islets when cotransplanted in a hypoxic environment. However, the risk of neoplasia is increased when MSCs are transplanted into immunosuppressed patients. This initial study aimed to investigate whether the production of protective factors from MSC can be stimulated by different culture conditions to benefit human islets cultured in hypoxia. METHODS: MSC were isolated from human adipose tissue and cultured for 2 days in supplemented Minimum Essential Media α (MEMα) and 21% (21%-MEMα) or 1% oxygen (1%-MEMα). Native MEMα served as control. After MSC harvesting, cell-depleted media were frozen at -20°C until use for human islet culture in 2% oxygen for 72-96 hours before islet characterization. Data were normalized to control islets cultured in native MEMα and 2% oxygen (mean ± SEM). RESULTS: After culture in 21%- or 1%-MEMα, islet recovery increased to 117 ± 12% (NS) and 138 ± 12% (P < .05), respectively. Viability did not change after culture in native MEMα (59 ± 2%), 21%-MEMα (59 ± 3%), or 1%-MEMα (61 ± 3%). Compared with control samples, the glucose stimulation index was increased after culture in 21%-MEMα (P < .05) or 1%-MEMα (P < .05). Overall survival was higher in 1%-MEMα (143 ± 14%) than in 21%-MEMα (119 ± 14%; NS) or native MEMα (P < .05). CONCLUSIONS: This study demonstrates that MSC-preconditioned MEMα increases survival and in vitro function of hypoxic human islets. These findings indicate that hypoxic MSCs seem to produce factors that improve survival of islets suffering from hypoxia.

Long-term Stability of a Compounded Suspension of Torsemide (5 mg/mL) for Oral Administration.

BACKGROUND: Torsemide use for congestive heart failure (CHF) has been reported, but prescription frequency is unknown. Commercially available tablet sizes in North America limit dosing precision, indicating a need to evaluate its strength and stability in suspension. OBJECTIVES: To determine the frequency of torsemide prescriptions and to determine a beyond use date (BUD) of a compounded suspension of torsemide for oral administration stored under 2 temperature conditions for 90 days. ANIMALS: No animals used. METHODS: Pharmacy records were retrospectively reviewed for torsemide and furosemide prescriptions from 2008 to 2015 at 2 veterinary referral centers. After preliminary strength testing, compounded torsemide suspension (5 mg/mL) for oral administration was prepared using torsemide tablets suspended in OraPlus:OraSweet 1:1, buffered to a pH of 8.3 and stored at refrigeration (2-8°C) and room temperature (20-25°C) in 2 oz amber plastic bottles. Samples were analyzed by reverse phase high-performance liquid chromatography (RP-HPLC) on days 0, 14, 30, 60, and 90. RESULTS: Prescriptions for torsemide increased from 2008 to 2015. Analysis of the torsemide 5 mg/mL suspension for oral administration at each time point met United States Pharmacopeia (USP) requirements for torsemide content of 90-110% of label claim. The average strength at 90 days decreased to 92 ± 3% at 2-8°C and 95 ± 2% at 20-25°C. Stability testing did not detect unknown impurities. CONCLUSIONS: Increasing torsemide use warrants availability of a validated and stable compounded formulation. Our results support the assignment of a 90-day BUD for torsemide 5 mg/mL suspension for oral administration compounded in OraPlus:Sweet 1:1 buffered to a pH of 8.3.

Key Matrix Proteins Within the Pancreatic Islet Basement Membrane Are Differentially Digested During Human Islet Isolation.

Clinical islet transplantation achieves insulin independence in selected patients, yet current methods for extracting islets from their surrounding pancreatic matrix are suboptimal. The islet basement membrane (BM) influences islet function and survival and is a critical marker of islet integrity following rodent islet isolation. No studies have investigated the impact of islet isolation on BM integrity in human islets, which have a unique duplex structure. To address this, samples were taken from 27 clinical human islet isolations (donor age 41-59, BMI 26-38, cold ischemic time < 10 h). Collagen IV, pan-laminin, perlecan and laminin-α5 in the islet BM were significantly digested by enzyme treatment. In isolated islets, laminin-α5 (found in both layers of the duplex BM) and perlecan were lost entirely, with no restoration evident during culture. Collagen IV and pan-laminin were present in the disorganized BM of isolated islets, yet a significant reduction in pan-laminin was seen during the initial 24 h culture period. Islet cytotoxicity increased during culture. Therefore, the human islet BM is substantially disrupted during the islet isolation procedure. Islet function and survival may be compromised as a consequence of an incomplete islet BM, which has implications for islet survival and transplanted graft longevity.

Microbial community structure across a wastewater-impacted riparian buffer zone in the southeastern coastal plain.

Riparian buffer zones are important for both natural and developed ecosystems throughout the world because of their ability to retain nutrients, prevent soil erosion, protect aquatic environments from excessive sedimentation, and filter pollutants. Despite their importance, the microbial community structures of riparian buffer zones remains poorly defined. Our objectives for this study were twofold: first, to characterize the microbial populations found in riparian buffer zone soils; and second, to determine if microbial community structure could be linked to denitrification enzyme activity (DEA). To achieve these objectives, we investigated the microbial populations of a riparian buffer zone located downslope of a pasture irrigated with swine lagoon effluent, utilizing DNA sequencing of the 16S rDNA, DEA, and quantitative PCR (qPCR) of the denitrification genes nirK, nirS, and nosZ. Clone libraries of the 16S rDNA gene were generated from each of twelve sites across the riparian buffer with a total of 986 partial sequences grouped into 654 operational taxonomic units (OTUs). The Proteobacteria were the dominant group (49.8% of all OTUs), with the Acidobacteria also well represented (19.57% of all OTUs). Analysis of qPCR results identified spatial relationships between soil series, site location, and gene abundance, which could be used to infer both incomplete and total DEA rates.

Contemporary management and outcomes for infants born with oesophageal atresia.

BACKGROUND: Reports on the management and outcome of rare conditions, such as oesophageal atresia, are frequently limited to case series reporting single-centre experience over many years. The aim of this study was to identify all infants born with oesophageal atresia in the UK and Ireland to describe current clinical practice and outcomes. METHODS: This was a prospective multicentre cohort study of all infants born with oesophageal atresia and/or tracheo-oesophageal fistula in 2008-2009 in the UK and Ireland to record current clinical management and early outcomes. RESULTS: A total of 151 infants admitted to 28 paediatric surgical units were identified. Some aspects of perioperative management were universal, including oesophageal decompression, operative technique and the use of transanastomotic tubes. However, there were a number of areas where clinical practice varied considerably, including the routine use of perioperative chest drains, postoperative contrast studies and antireflux medication, with each of these being employed in 30-50 per cent of patients. There was a trend towards routine postoperative ventilation. CONCLUSION: The prospective methodology used in this study can help identify practices that all surgeons employ and also those that few surgeons use. Areas of clinical equipoise can be recognized and avenues for further research identified.

Quality of isolated pig islets is improved using perfluorohexyloctane for pancreas storage in a split lobe model.

Pancreas transportation between donor center and islet production facility is frequently associated with prolonged ischemia impairing islet isolation and transplantation outcomes. It is foreseeable that shipment of pig pancreases from distant centralized biosecure breeding facilities to institutes that have a long-term experience in porcine islet isolation is essentially required in future clinical islet xenotransplantation. Previously, we demonstrated that perfluorohexyloctan (F6H8) is significantly more efficient to protect rat and human pancreata from ischemically induced damage compared to perfluorodecalin (PFD). To evaluate the effect of F6H8 on long-term stored pig pancreases in a prospective study, we utilized the split lobe model to minimize donor variability. Retrieved pancreases were dissected into the connecting and splenic lobe, intraductally flushed with UW solution and immersed alternately in either preoxygenated F6H8 or PFD for 8-10 h. Prior to pancreas digestion, the intrapancreatic pO2 and the ratio of ATP-to-inorganic phosphate was compared utilizing 31P-NMR spectroscopy. Isolated islets were cultured for 2-3 days at 37°C and subjected to quality assessment. Pancreatic lobes stored in preoxygenated F6H8 had a significantly higher intrapancreatic pO2 compared to pancreata in oxygen-precharged PFD (10.11 ± 3.87 vs. 1.64 ± 1.13 mmHg, p < 0.05). This correlated with a higher ATP-to-inorganic phosphate ratio (0.30 ± 0.04 vs. 0.14 ± 0.01). No effect was observed concerning yield and purity of freshly isolated islets. Nevertheless, a significantly improved glucose-stimulated insulin response, increased viability and postculture survival (57.2 ± 5.7 vs. 39.3 ± 6.4%, p < 0.01) was measured in islets isolated from F6H8-preserved pancreata. The present data suggest that F6H8 does not increase islet yield but improves quality of pig islets isolated after prolonged cold ischemia.

Outcomes of obese versus non-obese subjects undergoing robotic-assisted hysterectomy: a multi-institutional study.

The goal of our study was to determine whether there was a difference in operative outcomes in obese versus non-obese subjects undergoing robotic-assisted hysterectomies of varying levels of difficulty. Secondarily, we sought to analyze the published outcomes between robotic-assisted hysterectomy and total laparoscopic hysterectomy in obese women at each of these levels of difficulty. This was a multi-institutional retrospective cohort study of all patients undergoing robotic-assisted hysterectomy by five gynecologic oncologists at four geographically separate locations from April 2003 to March 2008. The cohort was stratified into obese vs. non-obese groups, and defined surgical outcomes compared between groups, then further divided into three subgroups based on case difficulty level. Univariate analysis and regression analysis using SAS 9.1 was performed. We then conducted a literature search of total laparoscopic hysterectomy outcomes in obese women, dividing the resulting studies into three comparative subgroups based on surgical difficulty levels for comparison with our robotic-assisted hysterectomy results. Our cohort had 228 obese and 323 non-obese subjects. Overall, the obese group had higher blood loss and longer operative time. When further stratified by level of difficulty, obese subjects also had a higher average blood loss and longer operative time in the hysterectomy-alone subgroup. No clinically significant differences in operative outcomes exist between obese and non-obese women when utilizing the da Vinci robotic system to perform a hysterectomy, independent of case difficulty level. More prospective, controlled studies which compare the two surgical approaches of robotic-assisted and laparoscopic hysterectomy approaches are needed.

Orogen styles in the East African Orogen: A review of the Neoproterozoic to Cambrian tectonic evolution.

The East African Orogen, extending from southern Israel, Sinai and Jordan in the north to Mozambique and Madagascar in the south, is the world́s largest Neoproterozoic to Cambrian orogenic complex. It comprises a collage of individual oceanic domains and continental fragments between the Archean Sahara-Congo-Kalahari Cratons in the west and Neoproterozoic India in the east. Orogen consolidation was achieved during distinct phases of orogeny between ∼850 and 550 Ma. The northern part of the orogen, the Arabian-Nubian Shield, is predominantly juvenile Neoproterozoic crust that formed in and adjacent to the Mozambique Ocean. The ocean closed during a protracted period of island-arc and microcontinent accretion between ∼850 and 620 Ma. To the south of the Arabian Nubian Shield, the Eastern Granulite-Cabo Delgado Nappe Complex of southern Kenya, Tanzania and Mozambique was an extended crust that formed adjacent to theMozambique Ocean and experienced a ∼650-620 Ma granulite-facies metamorphism. Completion of the nappe assembly around 620 Ma is defined as the East African Orogeny and was related to closure of the Mozambique Ocean. Oceans persisted after 620 Ma between East Antarctica, India, southern parts of the Congo-Tanzania-Bangweulu Cratons and the Zimbabwe-Kalahari Craton. They closed during the ∼600-500 Ma Kuungan or Malagasy Orogeny, a tectonothermal event that affected large portions of southern Tanzania, Zambia, Malawi, Mozambique, Madagascar and Antarctica. The East African and Kuungan Orogenies were followed by phases of post-orogenic extension. Early ∼600-550 Ma extension is recorded in the Arabian-Nubian Shield and the Eastern Granulite-Cabo Delgado Nappe Complex. Later ∼550-480 Ma extension affected Mozambique and southern Madagascar. Both extension phases, although diachronous,are interpreted as the result of lithospheric delamination. Along the strike of the East African Orogen, different geodynamic settings resulted in the evolution of distinctly different orogen styles. The Arabian-Nubian Shield is an accretion-type orogen comprising a stack of thin-skinned nappes resulting from the oblique convergence of bounding plates. The Eastern Granulite-Cabo Delgado Nappe Complex is interpreted as a hot- to ultra-hot orogen that evolved from a formerly extended crust. Low viscosity lower crust resisted one-sided subduction, instead a sagduction-type orogen developed. The regions of Tanzania and Madagascar affected by the Kuungan Orogeny are considered a Himalayan-type orogen composed of partly doubly thickened crust.

Perfluorocarbon emulsions prevent hypoxia of pancreatic ß-cells.

As oxygen carriers, perfluorocarbon emulsions might be useful to decrease hypoxia of pancreatic islets before transplantation. However, their hydrophobicity prevents their homogenisation in culture medium. To increase the surface of contact between islets and Perfluorooctyl bromide (PFOB), and consequently oxygen delivery, we tested effect of a PFOB emulsion in culture medium on ß-cell lines and rat pancreatic islets. RINm5F ß-cell line or pancreatic rat islets were incubated for 3 days in the presence of PFOB emulsion in media (3.5% w/v). Preoxygenation of the medium was performed before culture. Cell viability was assessed by apoptotic markers (Bax and Bcl-2) and by staining (fluoresceine diacetate and propidium iodide). ß-Cell functionality was determined by insulin release during a glucose stimulation test and. Hypoxia markers, HIF-1α and VEGF, were studied at days 1 and 3 using RT-PCR, Western blotting, and ELISA. PFOB emulsions preserved viability and functionality of RINm5F cells with a decrease of HIF-1α and VEGF expression. Islets viability was preserved during 3 days of culture. Secretion of VEGF was higher in untreated control (0.09 ± 0.041 µg VEGF/mg total protein) than in PFOB emulsion incubated islets (0.02 ± 0.19 µg VEGF/mg total protein, n = 4, p < 0.05) at day 1. At day 3, VEGF secretion was increased as compared to day 1 in control (0.23 ± 0.04 µg VEGF/mg total protein) but it was imbalance by the presence of PFOB emulsion (0.09 ± 0.03 µg VEGF/mg total protein, n = 5, p < 0.05). While insulin secretion was maintained in response to a glucose stimulation test until day 3 when islets were incubated in the presence of PFOB emulsion preoxygenated (0.81 ± 0.16 at day 1 vs. 0.75 ± 0.24 at day 3), the ability to secrete insulin in the presence of high glucose concentration was lost in islets controls (0.51 ± 0.18 at day 1 vs. 0.21 ± 0.13 at day 3). Atmospheric oxygen delivery by PFOB emulsion might be sufficient to decrease islets hypoxia. However, to improve islets functionality, overoxygenation is needed. Finally, maintenance of islet viability and functionality for several days after isolation could improve the outcome of islets transplantation.

Regulation of glucagon secretion by glucose: paracrine, intrinsic or both?

Glucagon secretion is regulated by glucose but the mechanisms involved remain hotly debated. Both intrinsic (within the α-cell itself) and paracrine (mediated by factors released ß- and/or δ-cells) have been postulated. Glucagon secretion is maximally suppressed by glucose concentrations that do not affect insulin and somatostatin secretion, a finding that highlights the significance of intrinsic regulation of glucagon secretion. Experiments on islets from mice lacking functional ATP-sensitive potassium channels (K(ATP)-channels) indicate that these channels are critical to the α-cell's capacity to sense changes in extracellular glucose. Here, we review recent data on the intrinsic and paracrine regulation of glucagon secretion in human pancreatic islets. We propose that glucose-induced closure of the K(ATP)-channels, via membrane depolarization, culminates in reduced electrical activity and glucagon secretion by voltage-dependent inactivation of the ion channels involved in action potential firing. We further demonstrate that glucagon secretion measured in islets isolated from donors with type-2 diabetes is reduced at low glucose and that glucose stimulates rather than inhibits secretion in these islets. We finally discuss the relative significance of paracrine and intrinsic regulation in the fed and fasted states and propose a unifying model for the regulation of glucagon secretion that incorporates both modes of control.

The long lifespan and low turnover of human islet beta cells estimated by mathematical modelling of lipofuscin accumulation.

AIMS/HYPOTHESIS: Defects in pancreatic beta cell turnover are implicated in the pathogenesis of type 2 diabetes by genetic markers for diabetes. Decreased beta cell neogenesis could contribute to diabetes. The longevity and turnover of human beta cells is unknown; in rodents <1 year old, a half-life of 30 days is estimated. Intracellular lipofuscin body (LB) accumulation is a hallmark of ageing in neurons. To estimate the lifespan of human beta cells, we measured beta cell LB accumulation in individuals aged 1-81 years. METHODS: LB content was determined by electron microscopical morphometry in sections of beta cells from human (non-diabetic, n = 45; type 2 diabetic, n = 10) and non-human primates (n = 10; 5-30 years) and from 15 mice aged 10-99 weeks. Total cellular LB content was estimated by three-dimensional (3D) mathematical modelling. RESULTS: LB area proportion was significantly correlated with age in human and non-human primates. The proportion of human LB-positive beta cells was significantly related to age, with no apparent differences in type 2 diabetes or obesity. LB content was low in human insulinomas (n = 5) and alpha cells and in mouse beta cells (LB content in mouse <10% human). Using 3D electron microscopy and 3D mathematical modelling, the LB-positive human beta cells (representing aged cells) increased from >or=90% (<10 years) to >or=97% (>20 years) and remained constant thereafter. CONCLUSIONS/INTERPRETATION: Human beta cells, unlike those of young rodents, are long-lived. LB proportions in type 2 diabetes and obesity suggest that little adaptive change occurs in the adult human beta cell population, which is largely established by age 20 years.

Somatostatin release, electrical activity, membrane currents and exocytosis in human pancreatic delta cells.

AIMS/HYPOTHESIS: The aim of this study was to characterise electrical activity, ion channels, exocytosis and somatostatin release in human delta cells/pancreatic islets. METHODS: Glucose-stimulated somatostatin release was measured from intact human islets. Membrane potential, currents and changes in membrane capacitance (reflecting exocytosis) were recorded from individual human delta cells identified by immunocytochemistry. RESULTS: Somatostatin secretion from human islets was stimulated by glucose and tolbutamide and inhibited by diazoxide. Human delta cells generated bursting or sporadic electrical activity, which was enhanced by tolbutamide but unaffected by glucose. Delta cells contained a tolbutamide-insensitive, Ba(2+)-sensitive inwardly rectifying K(+) current and two types of voltage-gated K(+) currents, sensitive to tetraethylammonium/stromatoxin (delayed rectifying, Kv2.1/2.2) and 4-aminopyridine (A current). Voltage-gated tetrodotoxin (TTX)-sensitive Na(+) currents contributed to the action potential upstroke but TTX had no effect on somatostatin release. Delta cells are equipped with Ca(2+) channels blocked by isradipine (L), omega-agatoxin (P/Q) and NNC 55-0396 (T). Blockade of any of these channels interferes with delta cell electrical activity and abolishes glucose-stimulated somatostatin release. Capacitance measurements revealed a slow component of depolarisation-evoked exocytosis sensitive to omega-agatoxin. CONCLUSIONS/INTERPRETATION: Action potential firing in delta cells is modulated by ATP-sensitive K(+)-channel activity. The membrane potential is stabilised by Ba(2+)-sensitive inwardly rectifying K(+) channels. Voltage-gated L- and T-type Ca(2+) channels are required for electrical activity, whereas Na(+) currents and P/Q-type Ca(2+) channels contribute to (but are not necessary for) the upstroke of the action potential. Action potential repolarisation is mediated by A-type and Kv2.1/2.2 K(+) channels. Exocytosis is tightly linked to Ca(2+)-influx via P/Q-type Ca(2+) channels. Glucose stimulation of somatostatin secretion involves both K(ATP) channel-dependent and -independent processes.

Results of a phase I/II British Society of Bone Marrow Transplantation study on PCR-based pre-emptive therapy with valganciclovir or ganciclovir for active CMV infection following alemtuzumab-based reduced intensity allogeneic stem cell transplantation.

This multi-centre randomized study assessed the bioavailability of ganciclovir in patients undergoing alemtuzumab-based reduced intensity conditioning (RIC) haematopoietic stem cell transplantation (HSCT) after oral administration of valganciclovir. Patients were randomized to 2 groups receiving either oral valganciclovir (900 mg twice daily) or intravenous ganciclovir (5mg/kg twice daily) for 14 days. Twenty-seven patients were recruited and 18 patients (67%) completed allocated treatment resulting in clearance of cytomegolovirus (CMV) DNA load at a median of 14 days. The bioavailability of ganciclovir from valganciclovir was 73% (95% CI: 34-112%). The average exposure in the valganciclovir group (36.9+/-14.9 microg h/ml) was higher than the ganciclovir cohort (27.9+/-7.5 microg h/ml). When compared with intravenous ganciclovir, oral valganciclovir had high bioavailability in patients undergoing alemtuzumab-based RIC HSCT.

Successful use of National Cancer Registry data to monitor the effective use of imatinib for treating chronic myeloid leukaemia.

UNLABELLED: Imatinib is a tyrosine kinase inhibitor, which selectively antagonises the BCR-ABL molecular pathway which causes chronic myeloid leukaemia (CML). Imatinib was first approved by the Scottish Medicines Consortium (SMC) in January 2002 with the recommendation that its use be audited. The cost of the drug has major financial implications for health resources. METHODS: All imatinib usage since its first prescription in Scotland in September 2000 to July 2003 was audited through pharmacy records and through the Scotland Leukaemia Registry (SLR), an existing national registry of patients with CML. RESULTS: One hundred and four patients in Chronic Phase (CP), 36 in Accelerated Phase (AP) and five in Blast Phase (BP) received imatinib. The median duration of therapy was not reached for CFP 17 months for AFP and two months for BP patients. Major (complete) cytogenetic response rates were 74% (63%) and 38% (24%) respectively for CP and APR Overall survival for all CP patients from the start of imatinib therapy was 94% at one year, 91% at two years and 83% at three years. An audit of the effectiveness of the SLR as an auditing agency, showed complete registration in 95% of cases. CONCLUSIONS: We believe such data collection should be an important ongoing resource for assessing outcomes in a rare form of leukaemia but one which already has major implications for health economics and will continue to do so given the future development of dual tyrosine kinase inhibitors for imatinib resistant cases.

Preparing and probing atomic number states with an atom interferometer.

We describe the controlled loading and measurement of number-squeezed states and Poisson states of atoms in individual sites of a double well optical lattice. These states are input to an atom interferometer that is realized by symmetrically splitting individual lattice sites into double wells, allowing atoms in individual sites to evolve independently. The two paths then interfere, creating a matter-wave double-slit diffraction pattern. The time evolution of the double-slit diffraction pattern is used to measure the number statistics of the input state. The flexibility of our double well lattice provides a means to detect the presence of empty lattice sites, an important and so far unmeasured factor in determining the purity of a Mott state.

Vascular endothelial growth factor as a survival factor for human islets: effect of immunosuppressive drugs.

AIMS/HYPOTHESIS: Rapamycin, part of the immunosuppressive regimen of the Edmonton protocol, has been shown to inhibit vascular endothelial growth factor (VEGF) production and VEGF-mediated survival signalling in tumour cell lines. This study investigates the survival-promoting activities of VEGF in human islets and the effects of rapamycin on islet viability. MATERIALS AND METHODS: Levels of VEGF and its receptors in isolated human islets and whole pancreas was determined by western blotting and immunostaining. Islet viability following VEGF or immunosuppressive drug treatment was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Islet VEGF release was measured by ELISA. Mouse islets infected with an adenovirus expressing the gene for VEGF were transplanted syngeneically into streptozotocin-induced diabetic mice, with blood glucose levels measured three times per week. RESULTS: Isolated human islets produced multiple isoforms of VEGF and VEGF receptors 1, 2 and 3 and the coreceptor neuropilin 1. Exogenous VEGF (10 ng/ml) prevented human islet death induced by serum starvation, which suggests that VEGF can act as a survival factor for human islets. Transplantation of mouse islets infected with a VEGF-expressing adenovirus in a syngeneic model, improved glycaemic control at day 1 post-transplantation (p < 0.05). Rapamycin at 10 and 100 ng/ml significantly reduced islet VEGF release (by 37 +/- 4% and 43 +/- 6%, respectively; p < 0.05) and at 100 ng/ml reduced islet viability (by 36 +/- 9%) and insulin release (by 47 +/- 7%, all vs vehicle-treated controls; p < 0.05). Tacrolimus had no effect on islet VEGF release or viability. CONCLUSIONS/INTERPRETATION: Our data suggest that rapamycin may have deleterious effects on islet survival post-transplantation, both through a direct effect on islet viability and indirectly through blockade of VEGF-mediated revascularisation.

Population-based demographic study of karyotypes in 1709 patients with adult acute myeloid leukemia.

Few large demographic studies of acute myeloid leukemia (AML) are derived from population-based registries. Demographic and karyotypic data were provided for AML cases from two regional leukemia registry databases in Scotland and the Northern Region of England. A population-based dataset was compiled, comprising 1709 patients aged >16 years (1235 North England/474 Scotland patients). The most common cytogenetic abnormalities involved chromosomes 5 and/or 7 (17%). Patients with the following abnormal chromosome 5/7 combinations: -5, del(5q), -5/-7 and del(5q)/-7 represented a significantly older population (P < 0.01, ANOVA). t(8;21) was the only 'favourable' karyotype found in older age. Karyotypic complexity varied within chromosome 5/7 combination groups; those containing -5, -5/-7, -5/del(7q), del(5q)/-7 or del(5q)/del(7q) combinations were significantly more frequently complex than those containing -7 and del(7q) (P < 0.01, chi2 test). Additional recurring cytogenetic abnormalities within complex karyotypes containing chromosome 5/7 combinations included (in order of frequency), abnormalities of chromosomes 17, 12, 3 and 18. Complex karyotypes not involving chromosomes 5 or 7 represented 30% of all complex karyotypes, occurred in younger patients than those involving chromosomes 5 and 7, and frequently included additional trisomy 8 (26%). In conclusion, we describe subgroups within adverse karyotypes, with different demographics, degree of complexity and additional chromosome abnormalities.