Pancreas and islet transplantation: an unfinished journey.
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Adult↗
Biomedical subjects
R D Allen
Publications and source records attributed to R D Allen.
Recovery of diabetic neuropathy after pancreas transplantation.
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Action Potentials↗
Polymorphism of the human TNF-alpha promoter--random variation or functional diversity?
As more and more polymorphisms are discovered in the genes encoding cytokines, a crucial question is whether this polymorphism has any functional effect. One of the most widely studied cytokine genes in this respect is the gene encoding human TNF-alpha. Much of the literature investigating the issue of whether TNF-alpha promoter polymorphisms have any functional effect on TNF-alpha transcription or influence disease susceptibility appears to report negative results, giving the appearance and leading some authors to conclude that polymorphism at this locus is functionally silent and exists only because of linkage disequilibrium with selectable HLA alleles. This review presents a new analysis of the available data which suggests that polymorphism in the TNF-alpha promoter is not randomly distributed and therefore that it most likely does have some functional and selectable effect. Further, a comparison of available data suggests that there is more consensus in the literature than may at first appear to be the case.
Alleles↗
Gelsolin inhibition of fast axonal transport indicates a requirement for actin microfilaments.
The actions of actin-based microfilaments in cell motility suggest a possible role in the mechanism of fast axonal transport, but the pharmacological data evaluating their role in this process are equivocal. Moreover, microfilaments are difficult to preserve and identify in ultrastructural studies, so the organization and function of axonal actin has remained uncertain. We have now evaluated the role of actin microfilaments in intracellular transport of membranous organelles using video-enhanced contrast microscopy and gelsolin to analyse fast axonal transport directly in isolated axoplasm from the squid giant axon. With this preparation it is possible to perfuse axoplasm with large molecules that do not cross the plasmalemma, while controlling cation levels. The 90,000-molecular weight protein gelsolin depolymerizes actin microfilaments in micromolar Ca2+, but not in the absence of Ca2+. Axonal transport of membranous organelles has previously been shown to be unaffected by levels of Ca2+ up to 10 microM. In the presence of EGTA, gelsolin has no effect on the movement of membranous organelles, but in the presence of 10 microM Ca2+ it completely blocks transport of all membranous organelles. No changes in the organization of the axoplasm were detected. These results and results using other probes for actin are consistent with the hypothesis that actin-based microfilaments are involved in the movement of membranous organelles in the axon.
Actins↗
Translocation of vesicles from squid axoplasm on flagellar microtubules.
Directed intracellular particle movement is a fundamental process characteristic of all cells. During fast axonal transport, membranous organelles move at rapid rates, from 1 to 5 micron s-1, in either the orthograde or retrograde direction along the neurone and can traverse distances as long as 1 m (for reviews, see refs 1-3). Recent studies indicate that this extreme example of intracellular motility can occur along single microtubules, but the molecules generating the motile force have not been identified or localized. It is not known whether the force-transducing 'motor' is associated with the moving particle or with the microtubule lattice. To distinguish between these hypotheses and to characterize the membrane-cytoskeletal interactions that occur during vesicle translocations, we have developed a reconstituted model for microtubule-based motility. We isolated axoplasmic vesicles from the giant axon of the squid Loligo pealei as described previously. The vesicles (35-475 nm in diameter) were then added to axonemes of Arbacia punctulata spermatozoa that served as a source of microtubules. Axonemes were used because the tubulin subunit lattice of the A-subfibre of a given outer doublet is the same as the subunit lattice of neuronal microtubules along which motility occurs. Moreover, all the microtubules of a single axoneme show the same structural polarity, indicating that the axoneme represents an oriented microtubule substrate. Here we demonstrate that vesicle motility is ATP-dependent, that it is not mediated by the flagellar force-transducing molecule dynein and that the direction of movement is not specified by microtubule polarity.
Adenosine Triphosphate↗
Monoclonal antibody to a bacterial endonuclear symbiont Holospora cross reacts with proteins of contractile vacuole radial canals of Paramecium species.
A monoclonal antibody (mAb) IR-2-1 was raised against a 67-kDa protein purified from the macronucleus-specific bacterial symbiont Holospora obtusa of Paramecium caudatum. The mAb was found to react with two bands (31 and 67-kDa) on gels of H. obtusa. Indirect immunofluorescence microscopy showed that these antigens were distributed inside the cells. However, unexpectedly, this mAb also cross reacted with the radial arms of the contractile vacuole in P. caudatum, P. tetraurelia, P. multimicronucleatum, P. jenningsi and P. bursaria as well as with their cytoplasm. Immunoelectron microscopy showed that the antigens were located on the decorated spongiome of the radial arms. In immunoblots, mAb IR-2-1 reacted with a band of 67 kDa in all Paramecium species examined. However, no band appeared in the immunoblot of isolated macronuclei of H. obtusa-free P. caudatum and no label was seen in the nuclear matrix of the macronucleus of air-dried P. caudatum. These results suggest that the 67-kDa antigen found in H. obtusa was not imported from the host macronucleus and the same antigen in the host contractile vacuoles and cytoplasm were not derived from the symbiont. These results also showed that an epitope on the decorated spongiome of the Paramecium species is shared by its bacterial symbiont. In contrast to the decorated tubule-specific mAb, DS-1, the antigens for IR-2-1 appeared to be loosely membrane bound as they were lost in paraformaldehyde fixed and acetone permeabilized Paramecium.
Animals↗
The striated bands of Paramecium are immunologically distinct from the centrin-specific infraciliary lattice and cytostomal cord.
The pellicle of Paramecium has three two-dimensionally arrayed systems that occupy separate but closely paralleling planes. All three systems are now distinguishable by their differing immunological properties. This study focused on the two deeper systems. The infraciliary lattice lies innermost and labels with centrin-specific antibodies. The middle system, the striated bands, is specifically labeled with a monoclonal antibody that we have raised to a 110 kDa cortical antigen in P. multimicronucleatum. This antibody labels a similar geometric cortical pattern in at least two species, P. multimicronucleatum and P. tetraurelia. Centrin-specific structures appear to be net-like in the above two species but show a more interrupted pattern in P. caudatum. The cytostomal cord is an essentially unbranched extension of the net-like infraciliary lattice and, like it, is centrin-specific. The cord has a unique association with the alveolar sacs which suggest these calcium-storing compartments contribute to the calcium fluxes required for contraction of the cord. A structural rather than a contractile function is favored for the striated bands, based solely on their morphology.
Animals↗
Phagosome formation in Paramecium: roles of somatic and oral cilia and of solid particles as revealed by video microscopy.
The roles of somatic and oral cilia and solid particles during digestive vacuole (DV) formation in Paramecium multimicronucleatum were investigated using video-enhanced and immunofluorescence microscopy. Membrane incorporation into DVs was found to increase linearly with increasing particle concentration. The rate of discoidal vesicle transport to the cytopharynx was not affected by particles, showing that particles are not required for membrane trafficking to the cytopharynx. However, the presence of particles leads to an increased membrane fusion between the cytopharyngeal membrane and the discoidal vesicles. When live cells lost their somatic cilia on the left-ventral side anterior to the oral region due to deciliation, membrane incorporation into newly formed DVs was strongly inhibited. Using video-enhanced microscopy, latex beads were seen to be loaded along the quadrulus on the dorsal surface of the buccal cavity, but few beads were seen next to the dorsal and ventral peniculi. Particle sequestration into a pre-formed nascent digestive vacuole (NDV) was studied in Triton X-100-permeabilized cells whose ciliary beating was reactivated by the addition of Mg-ATP. Both beat frequency and the percentage of cells containing bead-labeled NDV were dependent on the Mg-ATP concentration: the higher the beat frequency, the higher the percentage of cells with a bead-labeled NDV. These results suggest that ciliary beating is probably the only mechanism required for particle accumulation in the NDV, while a coordinated beating of the somatic cilia on the left-ventral side anterior to the oral region as well as the quadrulus moves particles into the NDV. The beating of the peniculi may somehow prevent the backward flow of particles out of the NDV.
Animals↗
A large-animal model to evaluate the clinical potential of fetal pig pancreas fragment transplantation.
The long-term goal of this study is to assess the feasibility of using fetal pig pancreas fragment (FPPF) transplantation to treat patients with type I diabetes. Using the highly inbred Westran Pigs, our initial aim was to establish a rejection-free transplant model of FPPF grafted into sibling recipient pigs without immunosuppression. FPPFs were isolated from 80-100-day-old fetuses of either Westran Pigs or outbred pigs and transplanted into the thymus, spleen, liver, or kidney of the recipient Westran pig. Biopsies were taken from each transplant site at set time points and assessed histologically for islet viability, rejection, and endocrine function. Fifty-eight fetal donors were used to transplant 16 recipient pigs. A nonspecific inflammation was seen for both outbred and inbred FPPF donor tissue at day 3 and was considered a response to ischemic necrosis. However, all the transplanted outbred FPPF donor tissue was acutely rejected and lost by day 10-14. In contrast, inbred FPPF tissue showed little evidence of graft necrosis after 3 days, and growth and formation of epithelial islet cell nest-like structures were seen to 28 days after transplantation. With time after transplantation, increasing amounts of insulin immunoperoxidase staining was seen together with chromogranin and somatostatin staining. In summary, this study confirms the potential of the Westran pig to answer the unproven ability of fetal pancreatic tissue to reverse type I diabetes in a large animal model.
Animals↗
Induction of allogeneic islet tolerance in a large-animal model.
For islet allotransplantation to become a therapy widely applicable to patients with insulin-dependent diabetes, it will be important to avoid conventional immunosuppression and yet maintain long-term rejection-free islet survival. This possibility was tested in a large-animal model using mixed allogeneic chimeras established using total lymphoid irradiation (TLI) and donor-specific bone marrow transplantation (BMTX). Four recipient sex-mismatched and DLA class II-matched English springer spaniels became chimeric after TLI and donor-specific BMTX. Subsequent donor-specific renal allografts survived for more than a year. Acceptance of a donor-specific skin graft and rejection of a third-party graft demonstrated tolerance with maintenance of immunocompetence. Pancreatic microfragments containing islets were refluxed into the splenic vein of the recipient. Purified islets were placed under the capsule of spleen and liver. After 75 days, recipients underwent total native pancreatectomy. All four chimeric pancreatectomized dogs had functioning islet grafts 75 days after transplantation, evidenced by a prompt rise in serum insulin levels following an IVGTT and histological demonstration of islet tissue at the site of transplantation. After removal of the transplanted islet tissue, no insulin was released after IVGTT. In summary, intrasplenic allogeneic canine islets transplanted into chimeric dogs rendered tolerant to donor MHC survive and function for greater than 75 days in the absence of immunosuppression. This study represents proof of the concept that allogeneic islet transplants have the potential to reverse diabetes without the use of conventional immunosuppression.
Amylases↗
National health insurance and the single specialist.
The factors affecting medical economics in the future will necessitate not only changes in existing systems of medical care, but even the implementation of new systems.
Delivery of Health Care↗