Local immune response against larvae of Rhipicephalus (Boophilus) microplus in Bos taurus indicus and Bos taurus taurus cattle.

Bos taurus indicus cattle are less susceptible to infestation with Rhipicephalus (Boophilus) microplus than Bos taurus taurus cattle but the immunological basis of this difference is not understood. We compared the dynamics of leukocyte infiltrations (T cell subsets, B cells, major histocompatibility complex (MHC) class II-expressing cells, granulocytes) in the skin near the mouthparts of larvae of R. microplus in B. t. indicus and B. t. taurus cattle. Previously naïve cattle were infested with 50,000 larvae (B. t. indicus) or 10,000 larvae (B. t. taurus) weekly for 6 weeks. One week after the last infestation all of the animals were infested with 20,000 larvae of R. microplus. Skin punch biopsies were taken from all animals on the day before the primary infestation and from sites of larval attachment on the day after the first, second, fourth and final infestations. Infiltrations with CD3+, CD4+, CD8+ and [gamma][delta] T cells followed the same pattern in both breeds, showing relatively little change during the first four weekly infestations, followed by substantial increases at 7 weeks post-primary infestation. There was a tendency for more of all cell types except granulocytes to be observed in the skin of B. t. indicus cattle but the differences between the two breeds were consistently significant only for [gamma][delta] T cells. Granulocyte infiltrations increased more rapidly from the day after infestation and were higher in B. t. taurus cattle than in B. t. indicus. Granulocytes and MHC class II-expressing cells infiltrated the areas closest to the mouthparts of larvae. A large volume of granulocyte antigens was seen in the gut of attached, feeding larvae.

Effect of the antitumor antibiotic chromomycin A3 on the humoral immune response in rats

Chromomycin A3 (250 mug/kg) suppressed the humoral immune response in rats against sheep erythrocytes when administered 48 h or later after antigenic stimulus. The antibiotic at this dose enhanced immunity when given along with or before antigen administration. The natural heterohemagglutinin levels in rabbits and guinea pigs were not affected by the antibiotic (10 mug/kg per day x 7).

Varied Immune Response to FVIII: Presence of Proteolytic Antibodies Directed to Factor VIII in Different Human Pathologies

The versatility of antibodies is demonstrated by the various functions that they mediate such as neutralization, agglutination, fixation of the complement and its activation, and activation of effector cells. In addition to this plethora of functions, antibodies are capable of expressing enzymatic activity. Antibodies with catalytic function are a result of the productive interplay between the highly evolved machinery of the immune system and the chemical framework used to induce them (antigens). Catalytic antibodies are immunoglobulins with an ability to catalyze the reactions involving the antigen for which they are specific. Catalytic immunoglobulins of the IgM and IgG isotypes have been detected in the serum of healthy donors. In addition, catalytic immunoglobulins of the IgA isotype have been detected in the milk of healthy mothers. Conversely, antigen-specific hydrolytic antibodies have been reported in a number of inflammatory, autoimmune, and neoplastic disorders. The pathophysiological occurrence and relevance of catalytic antibodies remains a debated issue. Through the description of the hydrolysis of coagulation factor VIII as model target antigen, we propose that catalytic antibodies directed to the coagulation factor VIII may play a beneficial or a deleterious role depending on the immuno-inflammatory condition under which they occur.

Enhancement of immune response by the proteinaceous crystal of Bacillus thuringiensis Var thuringiensis

The proteinaceous crystal of Bacillus thuringiensis Var thuringiensis was found to enhance humoral immune response in rats and guinea pigs immunised with sheep red blood cells. The enhancement was due to the increased levels of both 19S and 7S antibodies in the sera of the treated animals. A novel synthesis of 7S haemolytic antibodies was observed in case of crystal treated animals.

Immune response to insulin and changes in the gut immune system in children with or at risk for type 1 diabetes

Type 1 diabetes (T1D) is considered to be an autoimmune disease. The cause of T1D is the destruction of insulin-producing β-cells in the pancreatic islets. The autoimmune nature of T1D is characterized by the presence of autoreactive T-cells and autoantibodies against β-cell molecules. Insulin is the only β-cell-specific autoantigen associated with T1D but the insulin autoantibodies (IAAs) are difficult to measure with proper sensitivity. T-cell assays for detection of autoreactive T-cells, such as insulin-specific T-cells, have also proven to be difficult to perform. The genetic risk of T1D is associated with the HLA gene region but the environmental factors also play an important role. The most studied environmental risk factors of T1D are enteroviruses and cow's milk which both affect the immune system through the gut. One hypothesis is that the insulin-specific immune response develops against bovine insulin in cow's milk during early infancy and later spreads to include human insulin. The aims of this study were to determine whether the separation of immunoglobulin (Ig)G from plasma would improve the sensitivity of the IAA assay and how insulin treatment affects the cellular immune response to insulin in newly diagnosed patients. Furthermore, the effect of insulin concentration in mother's breast milk on the development of antibodies to dietary insulin in the child was examined. Small intestinal biopsies were also obtained from children with T1D to characterize any immunological changes associated with T1D in the gut. The isolation of the IgG fraction from the plasma of T1D patients negative for plasma IAA led to detectable IAA levels that exceeded those in the control children. Thus the isolation of IgG may improve the sensitivity of the IAA assay. The effect of insulin treatment on insulin-specific T-cells was studied by culturing peripheral blood mononuclear cells with insulin. The insulin stimulation induced increased expression of regulatory T-cell markers, such as Foxp3, in those patients treated with insulin than in patients examined before initiating insulin treatment. This finding suggests that insulin treatment in patients with T1D stimulates regulatory T-cells in vivo and this may partly explain the difficulties in measuring autoantigen-specific T-cell responses in recently diagnosed patients. The stimulation of regulatory T-cells by insulin treatment may also explain the remission period often seen after initiating insulin treatment. In the third study we showed that insulin concentration in mother's breast milk correlates inversely with the levels of bovine insulin-specific antibodies in those infants who were exposed to cow's milk proteins in their diet, suggesting that human insulin in breast milk induces tolerance to dietary bovine insulin. However, in infants who later developed T1D-associated autoantibodies, the insulin concentration in their mother's breast milk was increased. This finding may indicate that in those children prone to β-cell autoimmunity, breast milk insulin does not promote tolerance to insulin. In the small intestinal biopsies the presence of several immunological markers were quantified with the RT-PCR. From these markers the expression of the interleukin (IL)-18 cytokine was significantly increased in the gut in patients with T1D compared with children with celiac disease or control children. The increased IL-18 expression lends further support for the hypothesis that the gut immune system is involved in the pathogenesis of T1D.

Chlamydial infertility in women: Diagnosis, epidemiology and immune response

This project primarily focused on the development of a novel, non-invasive peptide-based ELISA to diagnose C. trachomatis-related infertility in women. In this study, an in silico approach was applied to identify and design novel peptide antigens that are specific to women with C. trachomatis-related infertility. The serological assay was developed such that it could potentially replace invasive techniques for early infertility investigation. The thesis further investigated the sero-prevalence of chlamydial infertility in women from a low-resource, high prevalence setting. In addition, the thesis also identified immune markers that were associated with the C. trachomatis-related infertility in women.

Characterization and immune response of a protein produced by a cDNA clone of foot and mouth disease virus, type Asia 1 63/72

A 0.9 kb double stranded cDNA of foot and mouth disease virus (FMDV) Type Asia 1, 63/72 was cloned in an expression vector, pUR222. A protein of 38 kd was produced by the clone which reacted with the antibodies raised against the virus. A 20 kd protein which may be derived from the 38 kd protein contained the antigenic epitopes of the protein VP1 of the virus. Injection of 10-20 micrograms of the partially purified 38 and 20 kd proteins or a lysate of cells containing 240 micrograms of the proteins elicited high titers of FMDV specific antibodies in guinea pigs and cattle respectively. Also, at these concentrations, the proteins protected 5 of 8 guinea pigs and 3 of 8 cattle when challenged with a virulent virus.

Growth kinetics and immune response of chimeric foot-and-mouth disease virus serotype `O' produced through replication competent mini genome of serotype Asia 1, 63/72, in BHK cell lines

Regular vaccinations with potent vaccine, in endemic countries and vaccination to live in non-endemic countries are the methods available to control foot-and-mouth disease. Selection of candidate vaccine strain is not only cumbersome but the candidate should grow well for high potency vaccine preparation. Alternative strategy is to generate an infectious cDNA of a cell culture-adapted virus and use the replicon for development of tailor-made vaccines. We produced a chimeric `O' virus in the backbone of Asia 1 and studied its characteristics. The chimeric virus showed high infectivity titre (>10(10)) in BHK 21 cell lines, revealed small plague morphology and there was no cross reactivity with antiserum against Asia I. The virus multiplies rapidly and reaches peak at 12 h post infection. The vaccine prepared with this virus elicited high antibody titres.

Fourteen gene GBM prognostic signature identifies association of immune response pathway and mesenchymal subtype with high risk group

Background: Recent research on glioblastoma (GBM) has focused on deducing gene signatures predicting prognosis. The present study evaluated the mRNA expression of selected genes and correlated with outcome to arrive at a prognostic gene signature. Methods: Patients with GBM (n = 123) were prospectively recruited, treated with a uniform protocol and followed up. Expression of 175 genes in GBM tissue was determined using qRT-PCR. A supervised principal component analysis followed by derivation of gene signature was performed. Independent validation of the signature was done using TCGA data. Gene Ontology and KEGG pathway analysis was carried out among patients from TCGA cohort. Results: A 14 gene signature was identified that predicted outcome in GBM. A weighted gene (WG) score was found to be an independent predictor of survival in multivariate analysis in the present cohort (HR = 2.507; B = 0.919; p < 0.001) and in TCGA cohort. Risk stratification by standardized WG score classified patients into low and high risk predicting survival both in our cohort (p = <0.001) and TCGA cohort (p = 0.001). Pathway analysis using the most differentially regulated genes (n = 76) between the low and high risk groups revealed association of activated inflammatory/immune response pathways and mesenchymal subtype in the high risk group. Conclusion: We have identified a 14 gene expression signature that can predict survival in GBM patients. A network analysis revealed activation of inflammatory response pathway specifically in high risk group. These findings may have implications in understanding of gliomagenesis, development of targeted therapies and selection of high risk cancer patients for alternate adjuvant therapies.

Inoculation of Triatoma Virus (Dicistroviridae: Cripavirus) elicits a non-infective immune response in mice

Background: Dicistroviridae is a new family of small, non-enveloped, +ssRNA viruses pathogenic to both beneficial arthropods and insect pests. Little is known about the dicistrovirus replication mechanism or gene function, and any knowledge on these subjects comes mainly from comparisons with mammalian viruses from the Picornaviridae family. Due to its peculiar genome organization and characteristics of the per os viral transmission route, dicistroviruses make good candidates for use as biopesticides. Triatoma virus (TrV) is a pathogen of Triatoma infestans (Hemiptera: Reduviidae), one of the main vectors of the human trypanosomiasis disease called Chagas disease. TrV was postulated as a potential control agent against Chagas' vectors. Although there is no evidence that TrV nor other dicistroviruses replicate in species outside the Insecta class, the innocuousness of these viruses in humans and animals needs to be ascertained. Methods: In this study, RT-PCR and ELISA were used to detect the infectivity of this virus in Mus musculus BALB/c mice. Results: In this study we have observed that there is no significant difference in the ratio IgG2a/IgG1 in sera from animals inoculated with TrV when compared with non-inoculated animals or mice inoculated only with non-infective TrV protein capsids. Conclusions: We conclude that, under our experimental conditions, TrV is unable to replicate inmice. This study constitutes the first test to evaluate the infectivity of a dicistrovirus in a vertebrate animal model.

The innate immune response to grass carp hemorrhagic virus (GCHV) in cultured Carassius auratus blastulae (CAB) cells

Virus infection of mammalian cells activates an innate antiviral immune response characterized by production of interferon (IFN) and the subsequent transcriptional upregulation of IFN-stimulated genes (ISGs) by the JAK-STAT signaling pathway. Here, we report that a fish cell line, crucian carp (Carassius auratus L.) blastulae embryonic (CAB) cells, can produce IFN activity and then form an antiviral state after infection with UV-inactivated grass carp hemorrhagic virus (GCHV), a double-stranded (ds) RNA virus. From UV-inactivated GCHV-infected CAB cells, 15 pivotal genes were cloned and sequenced, and all of them were shown to be involved in IFN antiviral innate immune response. These IFN system genes include the dsRNA signal sensing factor TLR3, IFN, IFN signal transduction factor STAT1, IFN regulatory factor IRF7, putative IFN antiviral effectors Mx1, Mx2, PKR-like, Viperin, IFI56, and other IFN stimulated genes (ISGs) IFI58, ISG15-1, ISG15-2, USP18, Gig1 and Gig2. The identified fish IFN system genes were highly induced by active GCHV, UV-inactivated GCHV, CAB IFN or poly(I).poly(C), and showed similar expression patterns to mammals. The data indicate that an IFN antiviral innate immune response similar to that in mammals exists in the UV-inactivated GCHV-infected CAB cells, and the IFN response contributes to the formation of an antiviral state probably through JAK-STAT signaling pathway. This study provides strong evidence for existence of IFN antiviral innate immune response in fish, and will assist in elucidating the origin and evolution of vertebrate IFN system. (c) 2006 Elsevier Ltd. All rights reserved.