bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular dynamics simulations of steady-state crystal growth and homogeneous nucleation in polyethylene-like polymer
Authors: Yamamoto Takashi
Published: November 2008

Molecular mechanisms of crystal growth and homogeneous nucleation from the melt of polyethylene-like linear polymer are investigated by molecular dynamics simulations. The present paper is aimed at extending our previous work with respect to the system size and the boundary condition, thereby enabling detailed studies on the structures of sufficiently large lamellae and fully equilibrated melt. Lamellae of uniform thickness but with marked tapered edges are found to grow at constant velocity from the substrate. Three-dimensional shape of the growing lamellae exhibits peculiar undulation at the growth front, the origin of which is suggested to be the inhomogeneous thickness distribution within the lamellae. Trajectories of chains crystallizing onto the growth front reveal an unexpected pathway for chain folding, where a partially attached chain stem forms a new fold by plunging its head back into a neighboring stem position through slithering snake motions of the chain. Detailed statistics of folds and cilia show that the folds are rather neat and mostly make re-entries into the nearest or the second or third nearest neighboring stem positions, whereas the cilia are generally short but with a small number of longer cilia forming thick amorphous layers. Structure of supercooled melt investigated versus temperature reveals that, at moderate degree of supercooling, the overall chain conformation remains Gaussian random coil but the persistent length of chains increases monotonically with increasing supercooling. Exceptions are at the largest supercooling where homogeneous nucleation takes place; usual melt structure becomes rapidly unstable and emerges many crystallites of random orientations. During early 10–20ns after the quench, density of melt, radius of gyration of chains, and fraction of kinked bonds show marked alterations. These structural changes are highly cooperative and are considered simply due to the emergence of many embryonic crystals in the melt. Conformations of the chains forming nuclei are also traced to reveal that the homogeneous nuclei are fringed micelle like aggregates of chains, but the chains as a whole have folded conformations, which are similar to those reported in previous simulations on a single polyethylene in a vacuum.
Asymmetric oscillations during phase separation under continuous cooling: A simple model
Authors: Hayase Yumino , Kobayashi Mika , Vollmer Doris , Pleiner Harald , Auernhammer Günter K.
Published: November 2008

We investigate the phase separation of binary mixtures under continuous cooling using the Cahn–Hilliard equation including the effect of gravity. In our simple model, sedimentation is accounted for by instantaneously “removing” droplets from the supersaturated mixture into the coexisting phase once the droplets have reached a defined maximum size. Our model predicts an oscillatory variation of turbidity. Depending on the composition, either both phases oscillate (symmetric oscillations) or only one of the phases oscillates (asymmetric oscillations). In the asymmetric case, droplet sedimentation from the majority phase into the minority phase reduces supersaturation in the minority phase. This inhibits droplet formation in the minority phase. The cooling rate dependence of the period agrees with experimental results.
Luminescence of Tb3+ doped TeO2–ZnO–Na2O–PbO glasses containing silver nanoparticles
Authors: Kassab Luciana R. P. , de Almeida Ricardo , da Silva Davinson M. , de Araújo Cid B.
Published: November 2008

Luminescence properties of Tb3+ doped TeO2–ZnO–Na2O–PbO glasses containing silver nanoparticles (NPs) were investigated. The absorption band due to the surface plasmon resonance in the NPs was observed. Its amplitude increases with the heat treatment of the samples that controls the nucleation of the NPs. Tb3+ emission bands centered at ≈485, ≈550, ≈585, and ≈623 nm were detected for excitation at 377 nm. The whole spectrum is intensified by the appropriate annealing time of the samples. Enhancement by ≈200% of the Tb3+ luminescence at 550 nm was observed for samples annealed at 270 °C during 62 h. This enhancement effect is due to the local field amplitude that increases with the amount of silver NPs and their aggregates.
Role of oxygen partial pressure and seed layer chemistry in flux mediated epitaxy of single phase multiferroic BiFeO3 thin films
Authors: Anbusathaiah Varatharajan , Cheng Ching Jung , Lim Sung Hwan , Murakami Makoto , Salamanca-Riba Lourdes G. , Takeuchi Ichiro , Nagarajan Valanoor
Published: November 2008

Multiferroic BiFeO3 (BFO) thin films have been fabricated via flux mediated epitaxy with varying oxygen partial pressure and flux composition (Bi2O3:CuO) conditions. Transmission electron microscopy coupled with energy dispersive x-ray spectroscopy as well as piezoresponse force microscopy confirm, that with the correct flux and seed layer conditions, even at very low partial pressures (3mTorr) no secondary phases are formed. The study reveals the crucial role of the bottom seed layer and flux chemistry in epitaxy of BFO thin films and provides alternate routes to BFO epitaxy in oxygen-deficient environments.
Effects of 2 mass % Si admixture in a laser-produced Fe plasma
Authors: Krása Josef , Láska Leoš , Rohlena Karel , Velyhan Andriy , Lorusso Antonella , Nassisi Vincenzo , Czarnecka Agata , Parys Piotr , Ryć Leszek , Wolowski Jerzy
Published: November 2008

Emission of multiply charged ions and soft x-rays from the plasmas produced by laser pulses focused on (111) surface of Fe and Fe−2 mass % Si single crystals is investigated for wavelengths of 1064 and 248 nm and intensities up to ≈1×1010 W/cm2. It is demonstrated that the Si admixture in the Fe plasma results in a higher emission of Feq+ ions (1≤q≤4) but in a markedly lower x-ray emission. The relation of wavelengths and pulse durations of laser beams used is figured in the fluence dependence of the ion emission.
Observation of magnetoelectric coupling in glassy epitaxial PbFe0.5Nb0.5O3 thin films
Authors: Correa Margarita , Kumar Ashok , Katiyar R. S. , Rinaldi Carlos
Published: November 2008

The temperature evolution of the dielectric spectra in PbFe0.5Nb0.5O3 (PFN) thin films showed a kink near the Néel temperature (∼170 K), diffuse phase transition (327–380 K) with frequency dispersion at dielectric maxima temperature, and almost frequency independent maxima at 600 K, suggesting weak magnetoelectric coupling, relaxor behavior, and structural phase transition, respectively. Dielectric properties and magnetic hysteresis revealed the coexistence of relaxor ferroelectricity and weak ferromagnetism at room temperature in PFN films. The magnetization as a function of temperature revealed a cusp in the zero field cooled curve for low magnetic field (∼100 Oe) near 170 K, suggesting spin glasslike behavior.
Magneto-optical and magnetotransport properties of amorphous ferromagnetic semiconductor Ge1−xMnx thin films
Authors: Yada Shinsuke , Sugahara Satoshi , Tanaka Masaaki
Published: November 2008

The authors investigate the magnetooptical and magnetotransport properties of Mn-doped amorphous Ge (a-Ge1−xMnx) thin films deposited on thermally oxidized Si substrates. Magnetic circular dichroism measurements reveal that their ferromagnetic ordering appears for x>0.02 and the films show magnetically homogeneous behavior. The resistivity of the films decreases with increasing x, and the temperature dependence of the resistivity shows semiconductor-like behavior. The anomalous Hall effect of the a-Ge1−xMnx films also exhibits ferromagnetic behavior. The temperature dependence of magnetization is consistent with that of the magnetooptical and magnetotransport results. These results indicate that the a-Ge1−xMnx films behave as a ferromagnetic semiconductor.
Fiber lasers generating radially and azimuthally polarized light
Authors: Fridman Moti , Machavariani Galina , Davidson Nir , Friesem Asher A.
Published: November 2008

A simple, robust, and efficient method to produce either radially or azimuthally polarized output beam from a fiber laser is presented. Experimental results reveal that polarization purity of 90% or better can be obtained.
A microstrip tunable negative refractive index metamaterial and phase shifter
Authors: He P. , Gao J. , Marinis C. T. , Parimi P. V. , Vittoria C. , Harris V. G.
Published: November 2008

A tunable negative refractive index metamaterial and miniature phase shifter have been designed and fabricated in a microstrip configuration for applications in radio frequency integrated circuits. The metamaterial consists of plasmonic copper wires and yttrium iron garnet slabs having a low insertion loss of 5dB at the center of the transmission band. The yttrium iron garnet material enables the magnetic field tuning of the negative refractive index in a dynamic frequency band from 7.0to11.0GHz. The insertion phase can be tuned by 45° continuously by varying the bias field from 3.8to4.6kOe at 9.0GHz.
Silicon out-diffusion and aluminum in-diffusion in devices with atomic-layer deposited La2O3 thin films
Authors: Jinesh K. B. , Lamy Y. , Wolters R. A. M. , Klootwijk J. H. , Tois E. , Roozeboom F. , Besling W. F. A.
Published: November 2008

The use of aluminum as an electrode in metal-insulator-semiconductor devices containing lanthanum oxide is impaired by unacceptable leakage current levels. Time of flight secondary ion mass spectroscopy depth profiling shows a significant amount of silicon out-diffusion from the substrate and aluminum in-diffusion towards the oxide. By using titanium nitride as the electrode, the silicon out-diffusion is suppressed, which improves the device performance. This indicates that, despite the larger coordination number of the lanthanum ions in the oxide, aluminum acts as a sink for silicon, thus driving the out-diffusion of silicon.
Translator awareness of semantic prosodies
Authors: Dam-Jensen Helle , Zethsen Karen Korning
Published: November 2008

An increasing amount of linguistic corpus-based research is being carried out and especially within semantics, some groundbreaking results are highly relevant to Translation Studies. Research has shown that evaluation in language is very much anchored in extended lexical units and not merely in the lexeme. Corpusbased semantic profiles of words reveal evaluative features which may not be accessible through introspection and which have therefore not been part of dictionary information. Previous empirical research has almost exclusively been concerned with native speakers of English. The present study, however, tests and compares the awareness of these evaluative features in the behaviour of soonto-be translators and first year LSP English students when using their foreign language.
Translations of ‘-ly’ adverbs of degree in an English-Spanish Parallel Corpus
Authors: Ramón Noelia , Labrador Belén
Published: November 2008

This paper focuses on the translations of English -ly adverbs of degree into Spanish. The English suffix -ly has been traditionally associated with the expression of manner. However, it also actualises other meanings, in particular degree. In Spanish, the formal equivalents of -ly adverbs are adverbs ending in -mente, but the latter occur less frequently and with different pragmatic nuances. Adverbs of degree can be translated into -menteadverbs but also into other resources such as non -menteadverbs, prepositional phrases, adjectives, etc. The aim is to establish a taxonomy of translation solutions extracted from a parallel corpus in order to reveal cross-linguistic correspondences useful in translator training and translation quality assessment.
Oxygen uptake of the human fetus at term
Authors: ACHARYA GANESH , SITRAS VASILIS
Published: November 2008

Objective. To measure oxygen uptake of term human fetuses and compare the values between those delivered vaginally following normal labor and those delivered by cesarean section before the onset of labor. Design. This was a prospective cross‐sectional study. Setting. University teaching hospital. Sample. Twenty healthy pregnant women at term (38–42 weeks) were included in this study. Among them, 10 were delivered by elective cesarean section and 10 had normal vaginal delivery. Methods. Umbilical vein volume blood flow was measured <24 hours before delivery. Immediately after delivery, the fetal weight was determined, and the umbilical venous and arterial blood samples were obtained. Blood gas analysis was performed and hemoglobin content was measured. Fetal oxygen uptake was calculated as a product of umbilical venous blood flow and the difference in the umbilical arterial and venous oxygen content. Results. We found that the mean oxygen uptake in human fetuses at term (median gestational age 39 weeks) to be 6.58 ml/min/kg (i.e. 0.29 mmol/min/kg). There was no significant difference in oxygen uptake between fetuses delivered following uncomplicated normal labor and those delivered by elective cesarean section before the onset of labor. Conclusion. Oxygen uptake of the appropriately grown normal human fetus at term is approximately 6.6 ml/kg/min and is not significantly affected by normal labor and delivery. Human fetuses tolerate intermittent reductions in uterine blood flow and oxygen supply associated with myometrial contractions during normal labor quite well.
Beyond the Tarmac Road: Local Forms of Policing in Sierra Leone & Rwanda
Authors: Baker Bruce
Published: November 2008

Civil war deeply disrupted policing in Sierra Leone and Rwanda, leaving their state police forces inadequate in numbers, skills and resources to serve all citizens. In this security vacuum local forms of policing play an important role. The article argues that the country-specific pattern of local forms of policing depends on three factors: the nature of the conflict and peace settlement; the regime ideology; and the level of regime insecurity and fear of conflict recurring. The empirical data concerning the local policing groups is presented under three headings: crime prevention and intervention; investigation and resolution; and punishment. The article concludes with an assessment of the hazards and potential for states and donors supporting such groups. They are certainly flawed agencies in the eyes of both users and government, but in a context of less than fair and accountable state policing, their widespread provision and support is not to be dismissed lightly.
Private Security & Local Politics in Somalia
Authors: Hansen Stig
                      Jarle
Published: November 2008

The use of private security by weak states is often seen to erode state power and prevent national institution building. This article investigates the use of private military force in Somalia and the three different entities that exercise political authority within this geographically defined territory, namely the Transitional Federal Government, Puntland, and Somaliland. All three have contracted private security companies, primarily to prevent piracy and illegal fishing in their costal waters. The article shows that while the turmoil in Somalia continues to offer lucrative investment opportunities for private security and military companies of various sorts, it cannot be uniformly concluded that private security always serves to weaken already fragile public authorities. On the contrary, in some cases the activities of private military companies have served to strengthen the power of local authorities.
Casein Kinase Iγ2 Down-Regulates Trafficking of Ceramide in the Synthesis of Sphingomyelin
Authors: Tomishige Nario , Kumagai Keigo , Kusuda Jun , Nishijima Masahiro , Hanada Kentaro
Published: November 2008

Intracellullar trafficking of lipids is fundamental to membrane biogenesis. For the synthesis of sphingomyelin, ceramide is transported from the endoplasmic reticulum to the Golgi apparatus by the ceramide transfer protein CERT. CERT is phosphorylated by protein kinase D at S132 and subsequently multiple times in a serine-repeat motif, resulting in its inactivation. However, the kinase involved in the multiple phosphorylation remains unclear. Here, we identify the γ2 isoform of casein kinase I (CKIγ2) as a kinase whose overexpression confers sphingomyelin-directed toxin-resistance to Chinese hamster ovary cells. In a transformant stably expressing CKIγ2, CERT was hyperphosphorylated, and the intracellular trafficking of ceramide was retarded, thereby reducing de novo sphingomyelin synthesis. The reduction in the synthesis of sphingomyelin caused by CKIγ2 was reversed by the expression of CERT mutants that are not hyperphosphorylated. Furthermore, CKIγ2 directly phosphorylated CERT in vitro. Among three γ isoforms, only knockdown of γ2 isoform caused drastic changes in the ratio of hypo- to hyperphosphorylated form of CERT in HeLa cells. These results indicate that CKIγ2 hyperphosphorylates the serine-repeat motif of CERT, thereby inactivating CERT and down-regulating the synthesis of sphingomyelin.
Thioredoxin h System and Wheat Seed Quality
Authors: Zahid Abderrakib , Afoulous Samia , Cazalis Roland
Published: November 2008

Thioredoxin is one of the key systems controlling cellular redox balance in all living organisms. Plant thioredoxins are a diverse multigene family divided into two systems, the chloroplastic and the cytoplasmic systems, which are distinguishable by the electron donor and by the enzyme that catalyses thioredoxin reduction. In cereal seed, the thioredoxin (Trx) h system acts in the developing phase, controlling the delivery of compounds during seed filling. Early in the development of the imbibed seed, it promotes the mobilization of storage nitrogen and carbon in the endosperm by inhibiting the inactivators of amylolytic enzyme and by activating a specific serine protease, thiocalsin. During seed maturation and germination, the Trx h system controls oxidative stress in the living tissues, specifically in the scutellum and the aleurone layer, where it accumulates in the nucleus. The overexpression and the suppression of Trx confirm these features and constitute a powerful tool to manage seed quality, due to the large number of Trx h isoforms and to the specific nature of some of them.
Sorghum Protein Extraction by Sonication and Its Relationship to Ethanol Fermentation
Authors: Zhao Renyong , Bean Scott R. , Wang Donghai
Published: November 2008

The objectives of this research were to develop a rapid method for extracting proteins from mashed and nonmashed sorghum meal using sonication (ultrasound), and to determine the relationships between the levels of extractable proteins and ethanol fermentation properties. Nine grain sorghum hybrids with a broad range of ethanol fermentation efficiencies were used. Proteins were extracted in an alkaline borate buffer using sonication and characterized and quantified by size‐exclusion HPLC. A 30‐sec sonication treatment extracted a lower level of proteins from nonmashed sorghum meal than extracting the proteins for 24 hr with buffer only (no sonication). However, more protein was extracted by sonication from the mashed samples than from the buffer‐only 24‐hr extraction. In addition, sonication extracted more polymeric proteins from both the mashed and nonmashed samples compared with the buffer‐only extraction method. Confocal laser‐scanning microscopy images showed that the web‐like protein microstructures were disrupted during sonication. The results showed that there were strong relationships between extractable proteins and fermentation parameters. Ethanol yield increased and conversion efficiency improved significantly as the amount of extractable proteins from sonication of mashed samples increased. The absolute amount of polymeric proteins extracted through sonication were also highly related to ethanol fermentation. Thus, the SE‐HPLC area of proteins extracted from mashed sorghum using sonication could be used as an indicator for predicting fermentation quality of sorghum.
Changes in aquatic insect emergence in response to whole‐lake experimental manipulations of introduced trout
Authors: POPE KAREN L. , PIOVIA‐SCOTT JONAH , LAWLER SHARON P.
Published: November 2008

1. Insects emerging from mountain lakes provide an important food source for many terrestrial predators. The amount of insect subsidy that emerges from lakes is influenced by predator composition, but predator effects could be ameliorated by greater habitat complexity. We conducted a replicated whole‐lake experiment to test the effects of introduced fish predators on the abundance and composition of aquatic insects within and emerging from the littoral zone of 16 mountain lakes in the Trinity Alps Wilderness in northwestern California.

2. Study treatments matched the fisheries management options being implemented in California’s wilderness areas: (i) continued stocking with non‐native trout, (ii) suspension of stocking, and (iii) suspension of stocking and removal of fish. We also included four naturally fishless ‘reference’ lakes. We compared abundances and biomass of emerging aquatic insects before treatments were initiated and for 3 years following their establishment. Abundances of benthic insects were also compared in the third year post‐treatment.

3. Trout removal rapidly increased abundances of mayflies, caddisflies, and insect predators, and overall insect biomass emerging from lakes. No significant differences were found between the suspension of stocking lakes and continued stocking lakes. Fish density was a more important predictor of aquatic insect emergence than habitat complexity.

4. Mayfly larvae responded positively to fish removal and caddisfly larvae tended to be more abundant in lakes without fish, but we did not detect effects on abundance of predatory insects. However, we found large insect predators in shallower water in lakes with fish compared to fish removal or fish‐free reference lakes.

5. These results provide insights into the continuing effects of past and current fish stocking practices on the flow of insect prey from mountain lakes into the neighbouring terrestrial environment. We also show that these consequences can rapidly be reversed by removing non‐native fishes.
Auto‐ and cross‐reactivity to thioredoxin allergens in allergic bronchopulmonary aspergillosis
Authors: Glaser A. G. , Menz G. , Kirsch A. I. , Zeller S. , Crameri R. , Rhyner C.
Published: November 2008

Background:  Thioredoxins are cross‐reactive allergens involved in the pathogenesis of atopic eczema and asthma. Cross‐reactivity to human thioredoxin can contribute to the exacerbation of severe atopic diseases.

Methods:  Human thioredoxin, Asp f28 and Asp f29, two thioredoxins of Aspergillus fumigatus, and thioredoxin of Malassezia sympodialis were cloned and produced as recombinant proteins. Allergenicity and cross‐reactivity to thioredoxins in allergic bronchopulmonary aspergillosis patients were assessed by enzyme‐linked immunosorbent assay (ELISA), inhibition ELISA, immunoblot analysis, proliferation assays and skin tests. Molecular homology modelling was used to identify conserved, surface‐exposed amino acids potentially involved in immunoglobulin E (IgE)‐binding.

Results:  All thioredoxins, including the human enzyme, bind IgE from patients with allergic bronchopulmonary aspergillosis and induce allergen‐specific proliferation in peripheral blood mononuclear cells and positive skin reactions in thioredoxin‐sensitized patients. Inhibition experiments showed that the thioredoxins are cross‐reactive indicating humoral immune responses based on molecular mimicry. To identify structural surface elements involved in cross‐reactivity, the three‐dimensional structures were modelled based on solved thioredoxin structures. Analysis of the molecular surfaces combined with sequence alignments allowed identification of conserved solvent exposed amino acids distantly located in the linear sequences which cluster to patches of continuous surface areas. The size of the surface areas conserved between human and fungal thioredoxins correlates well with the inhibitory potential of the molecules in inhibition ELISA indicating that the shared amino acids are involved in IgE‐binding.

Conclusions:  Conserved, solvent exposed residues shared between different thioredoxins cluster to continuous surface regions potentially forming cross‐reactive conformational B‐cell epitopes responsible for IgE‐mediated cross‐reactivity and autoreactivity.
CD4+ T cells, without CD8+ or B lymphocytes, can reject xenogeneic skin grafts
Authors: Wecker Hermann , Winn Henry J. , Auchincloss Hugh
Published: November 2008

Abstract: Previous experiments have shown that rejection of xenogeneic skin grafts by mice is particularly dependent on CD4+ T cells. There are two possible explantations for this finding: either 1) “help” provided by CD4+ T cells is essential for CD8+ T cell‐, B cell‐, or NK cell‐mediated effector mechanisms of rejection, or 2) CD4+ cells are themselves responsible for rejection, perhaps by some nonspecific effector mechanism. To examine these two hypotheses, we transplanted pig skin onto SCID mice and then reconstituted the mice with selected subpopulations of lymphocytes. Mice that did not received CD4+ T cells were unable to reject their xenografts, whereas those receiving CD4+ cells could do so in the absence of CD8+ cells or B cells and even when additionally depleted of NK cells by treatment with anti‐Asialo GM1 antibody. Additional experiments were performed both in vivo and vitro to confirm the absence in test mice of CD4+ or CD8+ and B lymphocytes, respectively. These results suggest that CD4+ T cells are not only necessary for rejection of xenogeneic skin grafts by mice, but that they can do so without CD8+ cells or B cells, and probably without NK cells. Since CD4+ cells in mice have been shown to recognize xenogeneic antigens indirectly, this suggests that a nonspecific effector mechanism may be involved in the rejection of xenografts. In these experiments allogeneic skin grafts behave quite differently as they could not be rejected by this mechanism.
Characterization of porcine endothelial cell determinants recognized by human natural antibodies
Authors: Collins Bradley H. , Parker William , Piatt Jeffrey L.
Published: November 2008

Abstract: Organs transplanted from pigs to primates are subject to hyperacute rejection. This immunologic reaction is initiated by the recipient's natural antibodies that bind to endothelial cell antigens of the organ, resulting in the activation of the complement system and rapid destruction of the graft. Various lines of evidence, particularly blocking studies, using purified carbohydrates have suggested that the endothelial cell determinant recognized by human natural antibodies is a terminal galactose in an α configuration (α‐Gal). Although these studies are compelling, they fall short of proof because xenoreactive natural antibodies, being polyreactive, might bind to structures other than those used for blocking. Moreover, in vivo evidence that anti‐α‐Gal antibodies participate in hyperacute rejection has not been reported. Here we report that an enzyme specific for α‐Gal, α‐galactosidase, removes α‐galactose residues from both intact porcine aortic endothelial cells and immobilized porcine aortic endothelial cell membrane extracts and that as a result of this, xenoreactive natural antibody binding is lowered by 70 to 80%. This decrease in binding of human IgM causes a corresponding decrease in complement activation. Similar results were obtained using the sera of other Old World primates. Digestion of immobilized porcine aortic endothelial cell membrane extracts with α‐galactosidase produced a similar reduction in human IgM binding to gp 115/135. These results indicate that a terminal α‐galactose is an important component of the gp 115/135 porcine endothelial cell antigen. We also report that perfusion of porcine organs I with primate serum removes anti‐α‐Gal IgM from the serum.
Specificities of human IgM and IgG anticarbohydrate xenoantibodies found in the sera of diabetic patients grafted with fetal pig islets
Authors: Rydberg Lennart , Cairns Thomas D.H. , Groth Carl‐Gustav , Gustavsson Mikael L. , Karlsson Eva C. , Mölier Erna , Satake Masahiro , Tibell Annika , Samuelsson B.E.
Published: November 2008

Abstract: Serum samples were collected from four diabetic patients who had received intraportal injections of pig fetal islet‐like cell clusters (ICC). The binding of IgM and IgG antibodies to glycosphingolipid antigens prepared from different pig organs and separated on thin layer plates was investigated in pre‐ and posttransplant serum samples. Both IgM and IgG antibodies in the pretransplant serum samples bind to several glycolipid fractions from the tri‐, tetra‐ and pentasaccharide regions but also to compounds with longer carbohydrate chains. In the posttransplant serum samples stronger binding, compared to the pretransplant samples, was noted for both Ig‐classes. Strong binding was seen in the pentasaccharide region known to contain the “linear blood group B” structure Galαl‐3Galβl‐4GlcNAcβl‐3Galβl‐4Glcβl‐lCer. There was no convincing evidence of the I recognition of new specificities. Adsorption of a patient serum to a Synsorbcolumn with Galαl‐3Gal specificity did not grossly change the binding pattern of the IgG antibodies in the effluate. However, the eluate from the column showed strong binding to the “linear B” compound but also to glycolipids with longer carbohydrate chains presumably with the same terminal epitope. Identification of these are in progress.

The human natural anti‐pig antibodies against carbohydrate antigens expressed on pig cells seem to detect a rather limitted number of epitopes. The “linear B” structure appears to be one major target. However, other structures may also be targets for xenoreactive antibodies. Immunisation with pig cells does not seem to initiate antibody production against new carbohydrate epitopes but leads to an increased production against the existing ones.
PEC‐A: An immortalized porcine aortic endothelial cell
Authors: Khodadoust Mehran M. , Candal Francisco J. , Maher Stephen E. , Murray Allan G. , Pober Jordan S. , Davis William C. , Ades Edwin W. , Bothwell Alfred L.M.
Published: November 2008

Abstract: Cultured porcine aortic endothelium was transfected with sequences that encode the SV40 T antigen, resulting in an immortalized cell line that retains the differentiated properties of normal aortic endothelial cells. Specifically, these cells form cobblestone monolayers, synthesize von Willebrand factor, and endocytose acetylated LDL. These cells can be readily propagated in culture and have been passaged over a year in culture. The cells form tubular structures when grown on Matrigel. The cells in culture express class I but do not express MHC class II antigens. Both class I and class II expression can be induced by treatment with recombinant swine interferon‐γ. Expression of CD44 and several other cell surface antigens have been observed. Co‐culture of these cells with purified human CD4+ T cells resulted in a significant T‐cell proliferative response similar to that observed for primary porcine EC. Finally, the cells are readily susceptible to transfection and express exogenous genes. These cells should be valuable for study of human anti‐porcine endothelial responses.
Expression of Galα(1,3)Gal by porcine islet cells and its relevance to xenotransplantation
Authors: McKenzie Ian F.C. , Koulmanda Maria , Sandrin Mauro S. , Mandel Tom E.
Published: November 2008

Abstract: In pig‐to‐human transplantation, one of the major obstacles is the expression of Galα(1,3)Galactose by the endothelium of vascularized human tissues and the presence in all humans of IgG and IgM antibodies to this epitope: xenotransplantation would be followed by hyperacute rejection (HAR). However, the findings in endothelial cells of all vessels and the parenchyma of tissues such as kidney and liver do not extend to islet cells. Histological studies demonstrate that the adult pig islet (apart from endothelial cells) does not express Galα(1,3)Gal, nor do fresh neonatal or fetal pig islet cells; after tissue culture (under a variety of conditions) large amounts of Galα(1,3)Gal are expressed by the pig islet cells. However, double staining studies show that Gal+ cells do not secrete insulin, glucagon, or somatostatin and these cells may be spared from potential antigen antibody‐mediated rejection after transplantation. The relevance of Gal expression in islet cells is discussed–short term studies–preferably in pig to Old World Monkey transplants could provide the answer to the relevance of Galα(1,3)Gal expression in islet cells. Thus far it appears that some islets can be destroyed by antibody; others are spared and it is important to determine the nature of the sparing mechanism.
Long‐term survival of pig‐to‐mouse Islet xenografts
Authors: Marchetti Piero , Scharp David W. , Lacy Paul E. , Navalesi Renzo
Published: November 2008

Abstract: Several laboratories are currently able to prepare large amounts of purified porcine islets of proven in vitro viability. The long‐term in vivo function of pig islet xenografts has been evaluated in both “nonimmunocompetent” animals (i.e., the nude mouse) and “immunocompetent” animals. In the nude mouse, documentation has been provided for pig islet function for up to 4 months, even though the issue of how quickly porcine islet xenografts restore normal blood glucose in this animal model is still controversial. Interestingly, pig islet xenografts drive glucose metabolism to maintain plasma glucose concentrations at the donor species levels. Porcine islets have been also transplanted into varying “immunocompetent” animals species. Long‐term pig islet xenograft survival in rats and larger animals has been achieved by transplanting islets immunoisolated by either macro‐ or microencapsulation techniques. In the pig‐to‐mouse experimental model, freshly prepared, nonimmunoisolated islets survived long‐term (for up to 50–60 days) when anti‐CD4 antibody treatment was given temporarily posttransplant. Neither the addition of either mouse and/or pig anti‐lymphocyte serum, nor the use of 1 week, low‐temperature cultured, or cryopreserved islets did further prolong the survival. When 2 to 3 week cultured islets were transplanted into anti‐CD4 antibody treated mice, function of the xenografts was observed at 100 days posttransplant in 75% of the animals. Thus, long‐term survival of pig‐to‐mouse islet xenografts in both nonimmunocompetent and immunocompetent animals is achievable. Although further studies are needed to fully understand the hormonal and metabolic effects of the islet xenografts, as well as to extend some of the results obtained in mice to larger animal models, the in vivo data available so far support the use of pig islets for potential use in human xenotransplantation studies.
Effects of collagenase concentration on the purity and viability of isolated porcine pancreatic islets for use in xenotransplantation studies
Authors: Zhou Daobiao , Yang Bing , Sun Yilu , Vacek Ivan , Sun Anthony M.
Published: November 2008

Abstract: The use of lower concentrations of collagenase (0.8 mg/ml as compared to 1.25 and 1.5 mg/ml) resulted in isolation of porcine pancreatic islets retaining their original compact, wholesome appearance. In in vitro experiments, 100% of islet batches isolated with the low collagenase concentration responded to glucose stimulation, while only 63.1% of islet batches isolated with the high concentration of the enzyme did. The response to high glucose challenge of islets isolated with the low enzyme concentration was considerably higher compared to high collagenase concentration islets.

In in vivo experiments, intraperitoneal transplants of microencapsulated islets isolated using 0.8 mg/ml of collagenase reversed diabetes in diabetic BALB‐c mice in a significantly larger proportion of recipients (87.5%) as compared to 33.3% of recipients grafted with 1.5 mg/ml collagenase islets.

The results of this study demonstrate that the use of a low collagenase concentration is conducive to isolation of islets of a superior morphology, purity, viability, and functioning ability.
Human IgM xenoreactive natural antibodies can induce resistance of porcine endothelial cells to complement‐mediated injury
Authors: Dalmasso Agustin R , He Tao , Benson Barbara A.
Published: November 2008

Abstract: It is thought that human IgM xenoreactive natural antibodies (nAbs) can induce activation of porcine endothelial cells independent of complement. Therefore we hypothesized that pretreatment of porcine endothelial cells with anti‐pig nAbs may affect the ability of the endothelial cells, when subsequently incubated with a source of nAbs and complement, to bind antibodies and complement components and to undergo complement‐mediated cytotoxicity. We preincubated porcine endothelial cells at 37°C for 1 hr or 40 hr with a source of nAbs. We then incubated these pretreated endothelial cells with a complement source that contained a normal complement level and a low level of IgM nAbs for 1 hr to measure bound IgM and IgG and complement components, and for 4 hr to measure cytotoxicity. We found that preincubation for as long as 40 hr did not impair the binding of IgM and IgG, implying no antibody‐induced loss of membrane antigens from the endothelial cells, or the binding of C3bi, C4d, C6, and C9 upon complement activation. In contrast, preincubation for 40 hr with a nAb source induced in the endothelial cells marked resistance to complement‐mediated killing. Resistance could be induced with purified human IgM but not with purified IgG or IgM‐depleted human serum. The ability of purified IgM to induce resistance was abrogated by removal of anti‐pig xenoreactive nAbs by absorption with pig endothelial cells, and induction of resistance required protein synthesis. We conclude that prolonged incubation of human anti‐pig nAbs with pig endothelial cells does not cause loss of endothelial cell membrane antigens or impairment in binding of nAbs or complement components; instead, it induces marked resistance to complement‐mediated cytotoxicity. These observations may be of value to develop strategies that enhance survival of a xenograft.
Reduction in Gal‐α1,3‐Gal epitope expression in transgenic mice expressing human H‐transferase
Authors: Chen Chao‐Guang , Fisicaro Nella , Shinkel Trixie A. , Aitken Victoria , Katerelos Marina , van Denderen Bryce J.W. , Tange Margaret J. , Crawford Robert J. , Robins Allan J. , Pearse Martin J. , D'Apice Anthony J.F.
Published: November 2008

Abstract: There is now considerable evidence implicating anti‐Gal xenoantibodies as a key instigator in the hyperacute rejection of discordant xenografts. As a consequence it is generally held that elimination or reduction of the Gal/anti‐Gal component is critical to overcoming hyperacute rejection. We have recently shown that in mice inactivation of the GalT gene by homologous recombination completely eliminates the expression of the Gal‐epitope and that hearts from these mice demonstrate prolonged survival when perfused ex vivo with human plasma. Unfortunately this strategy is currently not feasible in pigs because the technology to isolate porcine embryonic stem cells, which are critical for homologous recombination, is not yet available. This study investigates an alternative competition‐based transgenic strategy to suppress the level of the Gal epitope by expression of H‐transferase (α1,2‐fucosyltransferase) an enzyme which has the same substrate specificity (lactobiose) as α1,3‐galactosyltransferase. In vitro transfection of murine cells with H‐transferase reduced Gal‐epitope expression by 80–90%. A similar reduction in Gal expression was observed on PBL and thymocytes from H‐transferase transgenic mice. This reduction in Gal epitope expression resulted in a marked reduction in the reactivity of these cells with human serum. In tissues from these mice the reduction in Gal expression was inversely proportional to the endogenous level of Gal. The results of this study support pursuing this strategy as a means to reduce the xenoantigenicity of porcine tissues.
Longitudinal analysis of the expression of human decay accelerating factor (HDAF) on lymphocytes, in the plasma, and in the skin biopsies of transgenic pigs
Authors: Cozzi E. , Langford G.A. , Pino‐Chávez G. , Wright L. , Levy A. , Miller N , Davies H.f.f.S. , Chatterjee M. , Lancaster R. , Tolan M.J. , White D.J.G.
Published: November 2008

Abstract: Pigs transgenic for the regulators of human complement activation, such as decay accelerating factor (DAF), membrane cofactor protein (MCP), and CD59, have recently been produced as a means to overcome the organ shortage for clinical transplantation. Histological investigations in DAF transgenic pigs have demonstrated that large amounts of the transgenic protein are expressed on the endothelium of the organs, where hyperacute rejection is known to initiate. Several papers have recently appeared in the literature that show considerable variability in the expression of a transgene over the lifespan of a transgenic animal. In order to evaluate the consistency in the expression of the transgene in our human DAF (HDAF) transgenic pigs, we have analyzed 22 newborn transgenic pigs and 5 control littermates (group A) over a 6‐month period and 11 long‐term surviving transgenic animals (group B). In all the animals from group A, HDAF expression was investigated on peripheral blood mononuclear cells (PBMC) and in the plasma as a free circulating molecule at weekly intervals in the first month of life and monthly thereafter. In the same animals, HDAF expression was also analyzed on endothelial cells using sequential ear biopsies. Moreover, HDAF expression was evaluated on the endothelial cells of our longest surviving transgenic animals (group B) and compared with that observed in the biopsy taken at birth. Over the observation period, considerable variability was found in the blood of most animals from group A, both in the percentage of cells expressing the transgene and the mean number of HDAF molecules per cell. In the same animals, free circulating HDAF levels were shown to be very high at birth, progressively declining to a nadir reached between the second and the fifth week of life. No statistically significant correlation could be found between these three hematological parameters. Over the duration of the study, consistent expression of HDAF in the tissues was, however, found at the different time points in all the newborn animals from group A. Moreover, very high levels of HDAF, at least comparable to those observed at birth, were still detected in the skin biopsies of all the animals from group B after more than 18 months of life (mean: 25.18 ± 3.46; range: 21–31 months). Thus, our data show that over an observation period of up to 6 months, there is consistency in the expression of HDAF on the endothelium of serially collected skin biopsies from HDAF transgenic pigs and that levels of HDAF similar to those observed at birth are expressed by these transgenic animals after more than 2 years of life. Hematological parameters such as expression of HDAF on PBMC or measurements of free circulating HDAF in the plasma do not seem to correlate with the expression of HDAF on the endothelial cells.
Cellular xenoresponses: Although vigorous, direct human T cell anti‐pig primary xenoresponses are significantly weaker than equivalent alloresponses
Authors: Dorling Anthony , Binns Richard , Lechler Robert I.
Published: November 2008

Abstract: The in vitro human T cell xenoresponse to porcine stimulators has been examined in detail. Alveolar lavage cells (AL cells), derived from washouts of porcine lungs were found to possess phenotypic and functional properties of porcine dendritic cells. Using AL cells that had been allowed to mature during overnight incubation, we have shown that the “direct” T cell xenoresponse is mediated predominantly by CD4+ T cells that recognize SLA‐DR molecules. Limiting dilution analysis assays indicate a high precursor frequency of such cells in the peripheral blood of human responders. However, in a formal comparison with direct alloresponses, the proliferative response of human T cells to porcine dendritic cells was found to be significantly weaker. Using HLA‐DR and SLA‐DR‐transfected murine and porcine cell lines, we have shown that this relative weakness is due in part to an inefficiency of cognate interactions. In contrast, the non‐cognate interactions between human T cells and porcine stimulators are as efficient as those with human APC.
Aorta transplantation as a model to study hyperacute, acute, and chronic rejection of xenografts
Authors: Scheringa M. , Alwayn I.P.J. , Bouwman E. , Ijzermans J.N.M. , van Bockel J.H. , Marquet R.L.
Published: November 2008

Abstract: To explore the mechanism of rejection of vascularized xenografts, we performed discordant and concordant transplantation of aortic grafts. Forty‐one guinea pig‐to‐rat and 50 hamster‐to‐rat aortic transplantations were performed successfully. Recipient rats were either untreated or treated three times per week with 10 mg/kg of cyclosporine (CsA) starting the day before transplantation. Recipients were sacrificed at various timepoints after transplantation and the grafts were processed for histology and immunohistochemistry. Guinea pig aortic grafts were rejected hyperacutely in both the control and CsA treated group. Four hours after transplantation all endothelial cells and some of the medial smooth muscle cells had disappeared. Seven days after transplantation the media were totally acellular, whereas the adventitia was enlarged due to infiltrating cells, consisting mainly of macrophages. At day 56, only traces of the original graft could be identified.

Hamster aortic grafts developed signs of chronic rejection. In control rats, the intimal lesions could already be demonstrated in 17% of the rats at day 14, and in all rats from day 28. CsA treatment inhibited the development of the intimal lesions. At days 56 and 84 the difference in intimal thickness between the control group and the CsA treated group was statistically significant. The cellularity of the media of the hamster grafts remained constant during the first 21 days in both groups but declined sharply thereafter as a consequence of migration or necrosis of myocytes. The thickness of the adventitia in both groups increased after transplantation, due to infiltrating macrophages and T cells, reaching a peak at day 14. In conclusion, the aorta transplantation model provides useful information with regard to the development of chronic rejection in concordant grafts. Discordant aortic grafts are rejected hyperacutely just like discordant heart grafts.