Delayed allograft rejection in mice transgenic for a soluble form of the IL-4 receptor.

Accumulating evidence suggests that in serum and other biological fluids, cytokine binding is a property associated with soluble proteins, including a high-affinity soluble version of the IL-4 receptor (sIL-4R). While it is tempting to speculate that sIL-4R might act as a serum carrier protein or serve to inhibit or modulate IL-4 action, specific biological roles for sIL-4R remain to be established. To further assess the immunoregulatory and therapeutic potential of sIL-4R and other soluble receptors, we have created transgenic mice which constitutively express elevated levels of biologically active sIL-4R. Phenotypic characterization of lymphoid organs in sIL-4R transgenic mice revealed normal numbers of B and T cells and normal surface marker expression. Splenic lymphocytes displayed normal in vitro activities as measured by the PFC response and generation of cytotoxic T cells. In addition, antigen-specific IgE and IgG1 in vivo responses were similar in control and transgenic mice. Despite the apparent developmental normality of the sIL-4R transgenic mice, these animals were markedly deficient in the ability to reject cardiac allografts, suggesting that IL-4 is critical for the generation of alloreactivity. The results further suggest that the ability of sIL-4R to regulate IL-4 activities may be under the control of complex interactions that remain to be elucidated.     

Biodiversity of 52 chicken populations assessed by microsatellite typing of DNA pools

In a project on the biodiversity of chickens funded by the European Commission (EC), eight laboratories collaborated to assess the genetic variation within and between 52 populations from a wide range of chicken types. Twenty-two di-nucleotide microsatellite markers were used to genotype DNA pools of 50 birds from each population. The polymorphism measures for the average, the least polymorphic population (inbred C line) and the most polymorphic population (Gallus gallus spadiceus) were, respectively, as follows: number of alleles per locus, per population: 3.5, 1.3 and 5.2; average gene diversity across markers: 0.47, 0.05 and 0.64; and proportion of polymorphic markers: 0.91, 0.25 and 1.0. These were in good agreement with the breeding history of the populations. For instance, unselected populations were found to be more polymorphic than selected breeds such as layers. Thus DNA pools are effective in the preliminary assessment of genetic variation of populations and markers. Mean genetic distance indicates the extent to which a given population shares its genetic diversity with that of the whole tested gene pool and is a useful criterion for conservation of diversity. The distribution of population-specific (private) alleles and the amount of genetic variation shared among populations supports the hypothesis that the red jungle fowl is the main progenitor of the domesticated chicken.     

Effect of temperature on the saccharide composition obtained after alpha-amylolysis of starch

The hydrolysis of starch to low-molecular-weight products (normally characterised by their dextrose equivalent (DE), which is directly related to the number-average molecular mass) was studied at different temperatures. Amylopectin potato starch, lacking amylose, was selected because of its low tendency towards retrogradation at lower temperatures. Bacillus licheniformis alpha-amylase was added to 10% [w/w] gelatinised starch solutions. The hydrolysis experiments were done at 50, 70, and 90 degrees C. Samples were taken at defined DE values and these were analysed with respect to their saccharide composition. At the same DE the oligosaccharide composition depended on the hydrolysis temperature. This implies that at the same net number of bonds hydrolysed by the enzyme, the saccharide composition was different. The hydrolysis temperature also influenced the initial overall molecular-weight distribution. Higher temperatures led to a more homogenous molecular weight distribution. Similar effects were observed for alpha-amylases from other microbial sources such as Bacillus amyloliquefaciens and Bacillus stearothermophilus. Varying the pH (5.1, 6.2, and 7.6) at 70 degrees C did not significantly influence the saccharide composition obtained during B. licheniformis alpha-amylase hydrolysis. The underlying mechanisms for B. licheniformis alpha-amylase were studied using pure linear oligosaccharides, ranging from maltotriose to maltoheptaose as substrates. Activation energies for the hydrolysis of individual oligosaccharides were calculated from Arrhenius plots at 60, 70, 80, and 90 degrees C. Oligosaccharides with a degree of polymerisation exceeding that of the substrate could be detected. The contribution of these oligosaccharides increased as the degree of polymerisation of the substrate decreased and the temperature of hydrolysis increased. The product specificity decreased with increasing temperature of hydrolysis, which led to a more equal distribution between the possible products formed. Calculations with the subsite map as determined for the closely related alpha-amylase from B. amyloliquefaciens reconfirmed this finding of a decreased substrate specificity with increased temperature of hydrolysis. Copyright 1999 John Wiley & Sons, Inc.     

Antigen presentation by CD1d contributes to the amplification of Th2 responses to Schistosoma mansoni glycoconjugates in mice

During murine schistosomiasis, there is a gradual switch from a predominant Th1 cytokine response to a Th2-dominated response after egg laying, an event that favors the formation of granuloma around viable eggs. Egg-derived glycoconjugates, including glycolipids, may play a crucial role in this phenomenon. In this study, we used a model of dendritic cell sensitization to study the role of egg glycoconjugates in the induction of specific immune response to soluble egg Ag (SEA) and to investigate the possibility that CD1d, a molecule implicated in glycolipid presentation, may be involved in such a phenomenon. We show that, when captured, processed, and presented to naive T lymphocytes by dendritic cells, egg, but not larval, Ag skew the immune response toward a Th2 response. Periodate treatment reversed this effect, indicating that the sugar moiety of SEA is important in this phenomenon. Using DC treated ex vivo with a neutralizing anti-CD1d Ab or isolated from CD1d knockout mice, we show that CD1d is crucial in the priming of SEA-specific Th2 lymphocytes. We then evaluated the contribution of CD1d on the development of the SEA-specific immune response and on the formation of the egg-induced liver granuloma during murine schistosomiasis. We find that CD1d knockout mice have a reduced Th2 response after egg laying and develop a less marked fibrotic pathology compared with wild-type mice. Altogether, our results suggest that Ag presentation of parasite glycoconjugates to CD1d-restricted T cells may be important in the early events leading to the induction of Th2 responses and to egg-induced pathology during murine schistosomiasis.     

Comparison of the functional properties of murine dendritic cells generated in vivo with Flt3 ligand, GM-CSF and Flt3 ligand plus GM-SCF

Flt3 ligand (FL) and granulocyte-macrophage colony-stimulating factor (GM-CSF) are important growth factors for dendritic cells (DC). Substantial numbers of DC can be generated in vivo following the administration of either factor. We sought to extend our knowledge of the functional properties of these cells including their ability to prime na?ve CD8(+) T cells. In addition, we compared the nature of the DC generated in vivo with the single cytokines to those generated with the combination of FL+polyethylene glycol-modified GM-CSF (pGM-CSF). Treatment with FL+pGM-CSF yielded greater numbers of both CD11b(low) and CD11b(high) DC than with either cytokine alone, and these DC were more efficient at antigen (Ag) capture. The FL+pGM-CSF-generated CD11b(low) DC lacked expression of CD8alpha. Following treatment with LPS in vivo, all DC subsets upregulated CD40, CD80, CD86, and MHC class II expression, but surprisingly Ag capture was not downregulated and some DC subsets retained expression of intracellular MHC class II vesicles. Thus, even after activation in vivo with LPS, DC retained Ag capture properties of immature DC, and Ag presentation/costimulation properties of mature DC. Though all DC subsets stimulated CD4(+) T cell proliferation equivalently, FL-generated DC were more efficient at priming Ag-specific CD8(+) cytolytic T cells than DC generated with either pGM-CSF alone or FL+pGM-CSF, and CD11b(high) DC were more efficient at priming CD8(+) T cells than CD11b(low) DC.