Selective suppression of leukocyte recruitment in allergic inflammation

Allergic diseases result in a considerable socioeconomic burden. The incidence of allergic diseases, notably allergic asthma, has risen to high levels for reasons that are not entirely understood. With an increasing knowledge of underlying mechanisms, there is now more potential to target the inflammatory process rather than the overt symptoms. This focuses attention on the role of leukocytes especially Th2 lymphocytes that regulate allergic inflammation and effector cells where eosinophils have received much attention. Eosinophils are thought to be important based on the high numbers that are recruited to sites of allergic inflammation and the potential of these cells to effect both tissue injury and remodelling. It is hoped that future therapy will be directed towards specific leukocyte types, without overtly compromising essential host defence responses. One obvious target is leukocyte recruitment. This necessitates a detailed understanding of underlying mechanisms, particularly those involving soluble che-moattractants signals and cell-cell adhesion molecules.

Absence of VLDL secretion does not affect alpha-tocopherol content in peripheral tissues

alpha-Tocopherol is a lipid-soluble antioxidant that helps to prevent oxidative damage to cellular lipids. alpha-Tocopherol is absorbed by the intestine and is taken up and retained by the liver; it is widely presumed that alpha-tocopherol is then delivered to peripheral tissues by the secretion of VLDL. To determine whether VLDL secretion is truly important for the delivery of alpha-tocopherol to peripheral tissues, we examined alpha-tocopherol metabolism in mice that lack microsomal triglyceride transfer protein (Mttp) expression in the liver and therefore cannot secrete VLDL (Mttp(Delta/Delta) mice). Mttp(Delta/Delta) mice have low plasma lipid levels and increased stores of lipids in the liver. Similarly, alpha-tocopherol levels in the plasma were lower in Mttp(Delta/Delta) mice than in controls, whereas hepatic alpha-tocopherol stores were higher. However, alpha-tocopherol levels in the peripheral tissues of Mttp(Delta/Delta) mice were nearly identical to those of control mice, suggesting that VLDL secretion is not critical for the delivery of alpha-tocopherol to peripheral tissues. When fed a diet containing deuterated alpha-tocopherol, Mttp(Delta/Delta) and control mice had similar incorporation of deuterated alpha-tocopherol into plasma and various peripheral tissues. We conclude that the absence of VLDL secretion has little effect on the stores of alpha-tocopherol in peripheral tissues, at least in the mouse.

Schistosoma mansoni major egg antigen Smp40: molecular modeling and potential immunoreactivity for anti-pathology vaccine development

The pathogenesis of Schistosoma mansoni infection is largely determined by host T-cell mediated immune responses such as the granulomatous response to tissue deposited eggs and subsequent fibrosis. The major egg antigens have a valuable role in desensitizing the CD4+ Th cells that mediate granuloma formation, which may prevent or ameliorate clinical signs of schistosomiasis.S. mansoni major egg antigen Smp40 was expressed and completely purified. It was found that the expressed Smp40 reacts specifically with anti-Smp40 monoclonal antibody in Western blotting. Three-dimensional structure was elucidated based on the similarity of Smp40 with the small heat shock protein coded in the protein database as 1SHS as a template in the molecular modeling. It was figured out that the C-terminal of the Smp40 protein (residues 130 onward) contains two alpha crystallin domains. The fold consists of eight beta strands sandwiched in two sheets forming Greek key. The purified Smp40 was used for in vitro stimulation of peripheral blood mononuclear cells from patients infected with S. mansoni using phytohemagglutinin mitogen as a positive control. The obtained results showed that there is no statistical difference in interferon-g, interleukin (IL)-4 and IL-13 levels obtained with Smp40 stimulation compared with the control group (P > 0.05 for each). On the other hand, there were significant differences after Smp40 stimulation in IL-5 (P = 0.006) and IL-10 levels (P < 0.001) compared with the control group. Gaining the knowledge by reviewing the literature, it was found that the overall pattern of cytokine profile obtained with Smp40 stimulation is reported to be associated with reduced collagen deposition, decreased fibrosis, and granuloma formation inhibition. This may reflect its future prospect as a leading anti-pathology schistosomal vaccine candidate.

Latex constituents from Calotropis procera (R. Br.) display toxicity upon egg hatching and larvae of Aedes aegypti (Linn.)

Calotropis procera R. Br. (Asclepiadaceae) is a well-known medicinal plant with leaves, roots, and bark being exploited by popular medicine to fight many human and animal diseases. This work deals with the fractionation of the crude latex produced by the green parts of the plant and aims to evaluate its toxic effects upon egg hatching and larval development of Aedes aegypti. The whole latex was shown to cause 100% mortality of 3rd instars within 5 min. It was fractionated into water-soluble dialyzable (DF) and non-dialyzable (NDF) rubber-free materials. Both fractions were partially effective to prevent egg hatching and most of individuals growing under experimental conditions died before reaching 2nd instars or stayed in 1st instars. Besides, the fractions were very toxic to 3rd instars causing 100% mortality within 24 h. When both fractions were submitted to heat-treatment the toxic effects were diminished considerably suggesting low thermostability of the toxic compounds. Polyacrylamide gel electrophoresis of both fractions and their newly fractionated peaks obtained through ion exchange chromatography or desalting attested the presence of proteins in both materials. When submitted to protease digestion prior to larvicidal assays NDF lost most of its toxicity but DF was still strongly active. It may be possible that the highly toxic effects of the whole latex from C. procera upon egg hatching and larvae development should be at least in part due to its protein content found in NDF. However the toxicity seems also to involve non protein molecules present in DF.

Use of an immunodominant p17 antigenic fraction of Neospora caninum in detection of antibody response in cattle

A Neospora caninum 17 kDa protein fraction (p17) has been described as an immunodominant antigen (IDA) under reducing and non-reducing conditions. The aim of the present study was to investigate the diagnostic utility of p17 in cattle. In order to achieve this, p17 was purified by electroelution from whole N. caninum tachyzoite soluble extract and a p17-based Western blot (WB-p17) was developed. The p17 recognition was measured by densitometry and expressed as OD values to check the validity of the WB-p17. A total of 131 sera including sequential samples from naturally- and experimentally-infected calves and breeding cattle were analysed by WB-p17 and compared with IFAT using whole formalin-fixed tachyzoites as a reference test. The results obtained highlight the feasibility of using the N. caninum p17 in a diagnostic test in cattle. Firstly, the assay based on the p-17 antigen discriminated between known positive and negative sera from different cattle populations, breeding cattle and calves. Secondly, the p17 antigen detected fluctuations in the antibody levels and seroconversion in naturally- and experimentally-infected cattle. Significant differences in p-17 antigen recognition were observed between naturally infected aborting and non-aborting cattle, as well as significant antibody fluctuations over time in experimentally infected cattle, which varied between groups. Furthermore, the results obtained with WB-p17 are in accordance with the results obtained with the IFAT, as high agreement values were obtained when all bovine subpopulations were included (kappa = 0.86).

Preliminary results on interleukin-4 and interleukin-10 cytokine production in malnourished, inducible nitric oxide synthase-deficient mice with schistosomiasis mansoni infection

Schistosoma mansoni infected C57Bl/6 inducible nitric oxide synthase (iNOS)-deficient and non-deficient malnourished mice, both fed a balanced controlled diet were studied. Interleukins, IL-4 and IL-10 responses to soluble egg antigens (SEA) 90 days after infection, were determined. Our results suggest that in iNOS deficient, malnourished mice, 90 days after of infection, nitric oxide has a downregulating effect on IL-4 and IL-10 production. We are currently investigating the biological significance of these findings.

Effects of exercise on the secondary blood markers commonly used to suspect erythropoietin doping.

Numerous trials have reported that some haematological and biochemical parameters could be put together and be used to detect and fight recombinant erythropoietin doping. Unfortunately, none of the studies mentioned the necessity of taking pre-analytical precautions to avoid possible suspicious results coming from major plasma volume changes caused notably by dehydration. Therefore we studied the behaviour of the most common secondary blood markers before and after a strenuous physical activity to find out how reliable these parameters were. The soluble transferrin receptor and the haemoglobin concentrations as well as the haematocrit level increased significantly after effort, whereas the plasma EPO concentration and the reticulocyte count remained constant. On the other hand, if the values were corrected for haemoconcentration, the soluble transferrin receptor concentration remained stable.

Treatment of human acute schistosomiasis with oxamniquine induces an increase in interferon-gamma response to Schistosoma mansoni antigens

Patients with acute schistosomiasis were studied before and after oxamniquine treatment. They had been exposed to cercariae 5 to 9 weeks before, and presented compatible clinical manifestations, eosinophilia, and high levels of total IgE. Interferon-gamma (IFN-gamma) and interleukin-4 were measured by ELISA in whole blood samples under soluble egg antigen or soluble adult worm preparation stimulation. After treatment, the reduction of leukocytosis and eosinophilia were not significant, but total IgE levels decreased significantly, in contrast to IFN-gamma levels that were significantly increased. The oxamniquine treatment of acute schistosomiasis patients is followed by an improvement of a Th1 response in vitro. If this response has a protective aspect is unknown, and some investigations need to be realized.

Junctional adhesion molecule-2 (JAM-2) promotes lymphocyte transendothelial migration.

The molecular mechanisms underlying lymphocyte extravasation remain poorly characterized. We have recently identified junctional adhesion molecule-2 (JAM-2), and have shown that antibodies to JAM-2 stain high endothelial venules (HEVs) within lymph nodes and Peyer patches of adult mice. Here we show that mouse lymphocytes migrate in greater numbers across monolayers of endothelioma cells transfected with JAM-2. The significance of these findings to an understanding of both normal and pathologic lymphocyte extravasation prompted us to clone the human homologue of JAM-2. We herein demonstrate that an anti-JAM-2 antibody, or a soluble JAM-2 molecule, blocks the transmigration of primary human peripheral blood leukocytes across human umbilical vein endothelial cells expressing endogenous JAM-2. Furthermore, we show that JAM-2 is expressed on HEVs in human tonsil and on a subset of human leukocytes, suggesting that JAM-2 plays a central role in the regulation of transendothelial migration.