The transplantation of encapsulated islets of Langerhans is one approach to treat type 1 diabetes without the need of lifelong immunosuppression. Capillaries have been used for macroencapsulation ...because they have a favorable surface-to-volume ratio and because they can be refilled. It is unclear at present whether the outer surface of such capillaries should be smooth to prevent, or rough to promote, cell adhesions. In this study we tested a new capillary made of modified polysulfone (MWCO: 50 kDa) with a rough, open-porous outer surface for islet transplantation. Compared with free-floating islets, encapsulation of freshly isolated rat islets affected neither the kinetics nor the efficiency of glucose-induced insulin release in perifusion experiments. Free-floating islets maintained insulin secretion during cell culture but encapsulated islets gradually lost their glucose responsiveness and released VEGF. This indicated hypoxia in the capillary lumen. Transplantation of encapsulated rat islets into diabetic rats significantly reduced blood glucose concentrations from the first week of implantation. This hypoglycaemic effect persisted until explantation 4 weeks later. Transplantation of encapsulated porcine islets into diabetic rats reduced blood glucose concentrations depending on the islet purity. With semipurified islets a transient reduction of blood glucose concentrations was observed (2, 8, 18, 18 days) whereas with highly purified islets a sustained normoglycaemia was achieved (more than 28 days). Explanted capillaries containing rat islets were covered with blood vessels. Vascularization was also observed on capillaries containing porcine islets that were explanted from normoglycaemic rats. In contrast, on capillaries containing porcine islets that were explanted from hyperglycemic rats a fibrous capsule and lymphocyte accumulations were observed. No vascularization on the surface of transplanted capillaries was observed in the absence of islets. In conclusion, encapsulated islets can release VEGF, which appears to be an important signal for the vascularization of the capillary material. The rough, open-porous outer surface of the polysulfone capillary provides a site well suited for vascular tissue formation and may allow a prolonged islet function after transplantation.
Cold antihydrogen atoms were produced by mixing cold samples of antiprotons and positrons. The temperature of the positron plasma was increased by controlled radio-frequency (RF) heating, and the ...antihydrogen production was measured. Formation is observed to decrease with increased temperature but a simple power law scaling is not observed. Significant production is still present at room temperature.
The determination of islet mass is important for the normalization of islet experiments in the laboratory and for the precise dosing of islets for transplantation. The common microscopical analysis ...is based on individual islet sizing, calculation of the frequency distribution, and conversion into islet equivalents (IEQ), which is the volume of a spherical islet with a diameter of 150 μm. However, islets are of irregular form, which makes this determination user dependent, and the analysis is irreproducible once the original sample is discarded. This routine technique of islet quantification was compared with the analysis of areal density measurements. It was assumed that the entire area occupied by islets can be expressed in IEQ without sizing and counting individual islets. Porcine islets were isolated by continuous digestion/filtration and purified by gradient centrifugation. Purified islets were stained with dithizone and were repeatedly pictured under the microscope with random area selection. A total of 51 pictures was taken from 11 different purifications and stained islets were detected by digital image analysis. The correlation coefficient (r) between both analyses was 0.977 with an underestimation of islet yield by areal density detection (slope: 0.75 ± 0.03). Areal density analysis per picture took about 1 min, which is about 10 times faster than the traditional method without increasing the method error (CV 2.1% vs. 2.7%). In summary, areal density measurements allow a rapid and reproducible estimation of IEQ without counting individual islets. It can be performed in a single step analysis without computer programming and is valuable for online determinations of islet yield preceding transplantation.
At the moment autologous nerve grafting remains the only reasonable technique for reconstruction of peripheral nerve defects. Unfortunately, this technique has a lot of complications and ...disadvantages. These problems are related to the autologous nerve that is harvested for this procedure. Donor site morbidity with loss of sensitivity, painful neuroma formation and of course the restricted availability of autologous nerves stimulates the idea for alternative techniques on that field. In this paper we describe our experience with different graft materials for reconstruction of a 2 cm nerve gap in a median nerve model in rats. After implantation of various materials (biological/synthetic) the main experiments were conducted with a synthetic, biodegradable nerve conduit seeded with autologous Schwann cells. With this material we were able to reconstruct successfully a 2 cm gap in the rat median nerve. Regeneration with this material was found to be equally to an autologous nerve graft.
Thrombogenicity of small diameter vascular prostheses might be reduced by complete coverage of the luminal surface with vascular cells. We investigated cell seeding on polyurethane vascular ...prostheses (PUVP).
45 PUVP were divided into three groups of n = 15 each: Group A (diameter 20 mm, gamma-sterilized), Group B (diameter 4 mm, gamma-sterilized), and Group C (diameter 4 mm, ethylene oxid Eto-sterilized). Human smooth muscle cells (SMC), fibroblasts (FB), and endothelial cells (EC) were isolated from saphenous vein segments and expanded in culture. PUVPs were pre-seeded with a mixed culture of FBs and SMCs (mean 7.7 +/- 2.3 x 10(6) cells) followed by EC seeding (mean 4.4 +/- 0.9 x 10(6) cells). Seven days after cell seeding, PUVPs were perfused under a pulsatile flow. Flow definitions were as follows: adaption phase: low flow, resulting pressure: 60/30 mm Hg; high flow: resulting pressure: 160/50 mm Hg, lasting for 4 hours in all groups. Three subgroups were defined out of each group, differing in the perfusion strategy: high flow immediately, adaption phase of 15 minutes followed by high flow, and adaption phase of 30 minutes followed by high flow. Specimens were taken after each seeding procedure, prior to and after perfusion, and then examined using a scanning electron microscope (SEM) and immunohistochemical staining procedures.
Pre-seeding with the mixed culture revealed a better initial adhesion in Groups A and B compared to group C (76% vs. 41%). In Groups A and B, EC seeding (adhesion 72%) resulted in a confluent EC layer. Immunohistochemical stainings were positive for collagen IV, laminin, CD31, and factor VIII, but negative for eNOS. In Group C, only isolated cells were found after each seeding procedure, which rounded up and vanished during the next days. When perfused with high-flow immediately, Group A and B prostheses revealed small defects (< 10% of the surface) of all cell layers. After perfusion with an adaption phase of 15 minutes only few defects were found within the EC layer with an intact basement membrane. An adaption phase of 30 minutes resulted in a confluent cell layer without significant cell defects. After perfusion, the endothelial cells also stained positive for eNOS.
Seeding of a mixed culture consisting of FBs and SMC resulted in an excellent EC adhesion and resistance to shear stress. Cell attachment was better on gamma-sterilized PUVPs compared to Eto-sterilization. The cells obviously maintained their ability to adapt to shear stress.
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