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m.d. farrar, t.r. whitehead, j. lan, p. dilger, r. thorpe, k.t. holland and s.r. carding. 2005.The aim of this work was to engineer a gut commensal bacterium,Bacteroidesovatus, to produce and secrete a biologically active cytokine in a regulated manner as a basis for novel immunotherapies for chronic gut disorders.Bacteroides ovatuswas engineered to produce murine interleukin-2 (MuIL2) intracellularly in response to xylan in culture media by inserting the MuIL2 gene into the xylanase operon of the organism. A second strain was engineered to secrete MuIL2 by addingBacteroides fragilisenterotoxin secretion signal sequence to the protein. The recombinant strains produced MuIL2 only in the presence of xylan as determined by ELISA of cell lysates and culture supernatants. The IL2-dependent cell line CTLL-2 was used to demonstrate that MuIL2 produced by bothB. ovatusstrains was biologically active. This activity could be blocked by an anti-IL2 neutralizing antibody. The xylan-inducible nature of this system was demonstrated by RT-PCR.Bacteroides ovatuswas successfully engineered to produce and secrete biologically active MuIL2 in a xylan-inducible manner.The production and secretion of a biologically active mammalian protein by a member of the gut microflora could lead to the development of new long-term immunotherapies for inflammatory gut diseases. [ABSTRACT FROM AUTHOR]

We analyse molecular and phenotypic evolution in a group of taxonomically problematic Indomalayan pitvipers, the Trimeresurus sumatranus group. Mitochondrial DNA sequencing provides a well-resolved phylogeny, with each species representing a distinct lineage. Multivariate morphological analysis reveals a high level of phenotypic differentiation, which is congruent between the sexes but does not reflect phylogenetic history. An adaptive explanation for the observed pattern of differentiation is supported by independent contrasts analysis, which shows significant correlations between current ecology and the characters that most account for the variation between taxa, including those that are presently used to identify the species. Reduced precipitation and altitude, and increased temperature, are correlated with higher numbers of scales on the head, body and tail. It is hypothesized that scale number plays an important role in heat and water exchange by influencing the area of exposed of interstitial skin, and that colour pattern variation reflects selection pressures involving camouflage and thermoregulation. Ecological convergence in traits used for classification is found to have important implications for species identification where taxa are distributed over varying environments. [ABSTRACT FROM AUTHOR]

SUMMARY We have screened for spontaneous anticytokine autoantibodies in patients with infections, neoplasms and autoimmune diseases, because of their increasingly reported co-occurrence. We tested for both binding and neutralizing autoantibodies to a range of human cytokines, including interleukin-1alpha (IL-1α ), IL-1β , IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, IL-18, interferon-alpha2 (IFN-α 2), IFN-ω , IFN-β , IFN-γ , tumour necrosis factor alpha (TNF-α ), transforming growth factor beta-1 (TGF-β 1) and granulocyte-macrophage colony stimulating factor (GM-CSF), in plasmas or sera. With two notable exceptions described below, we found only occasional, mostly low-titre, non-neutralizing antibodies, mainly to GM-CSF; also to IL-10 in pemphigoid. Strikingly, however, high-titre, mainly IgG, autoantibodies to IFN-α 2, IFN-ω and IL-12 were common at diagnosis in patients with late-onset myasthenia gravis (LOMG+ ), thymoma (T) but no MG (TMG– ) and especially with both thymoma and MG together (TMG+ ). The antibodies recognized other closely related type I IFN-α subtypes, but rarely the distantly related type I IFN-β , and never (detectably) the unrelated type II IFN-γ . Antibodies to IL-12 showed a similar distribution to those against IFN-α 2, although prevalences were slightly lower; correlations between individual titres against each were so modest that they appear to be entirely different specificities. Neither showed any obvious correlations with clinical parameters including thymoma histology and HLA type, but they did increase sharply if the tumours recurred. These antibodies neutralized their respective cytokine in bioassays in vitro ; although they persisted for years severe infections were surprisingly uncommon, despite the immunosuppressive therapy also used in most cases. These findings must hold valuable clues to autoimmunizing mechanisms in... [ABSTRACT FROM AUTHOR]

Serum samples from 557 individuals participating in studies from four separate lowland and highland populations in Papua New Guinea exhibited consistently false-positive results for human T lymphotropic virus (HTLV) type 1 (100%) and human immunodeficiency virus (HIV) type 1 (50%) antibody in direct antiglobulin and agglutination assays. All serum samples werenegative in competitive ELISAs and radioimmunoassays for HTLV-1 and HIV-1; selected samples of reactive sera were negative in an HTLV-2 competitive ELISA. Immunofluorescent antibody tests using HTLV-1 infected cells correlated poorly with ELISA results. None of the sera from Papua New Guinea neutralized vesicular stomatitis virus pseudotypes of HTLV-1. By Western blot analysis, only three serum samples were weakly reactive to HTLV-1 gag proteins. These studies suggest there is as yet no firm evidence of HTLV-1, HTLV-2,or HIV-1 infection in Papua New Guinea, although there may be a low prevalence. [ABSTRACT FROM PUBLISHER]

In this study, the development of neutralizing und non-neutralizing GM-CSF antibodies and the clinical consequences related to the induction of these antibodies were analysed in 20 patients with metastatic colorectal carcinoma receiving a combination therapy of Escherichia coli-derived GM-CSF and a colon carcinoma-reactive MoAb in the absence of any concomitant chemotherapy. The recombinant human GM-CSF was administered subcutancously for 10 days every month for 4 months. Following the first cycle of treatment, no GM-CSF antibodies were detected, but during subsequent therapy, 19 of the 20 patients studied developed GM-CSF binding antibodies. However, only a proportion (40%) of the 19 antibody-positive patients developed antibodies that neutralized the biological activity of GM-CSF in an in vitro bioassay. The presence of GM-CSF neutralizing antibodies was associated with a significant reduction in GM-CSF-induced expansion of leucocytes, neutrophils and eosinophils. Such clinical effects were not apparent in patients with non-neutralizing antibodies. Further characterization of sera from patients with neutralizing antibodies showed that, in most cases, the antibodies neutralized the biological activity of GM-CSF preparations derived using different expression systems (Chinese hamster ovary cells and yeast), suggesting that these antibodies may have the potential to cross-react with endogenously produced GM-CSF, These effects should be considered before therapeutic use of cytokines, particularly in patients who are not immunosuppressed, and therefore capable of mounting an effective immune response. Our results indicate that assessment of production of neutralizing antibodies induced during cytokine therapy can be used to predict diminished clinical response to further therapy. [ABSTRACT FROM AUTHOR]

Although the immunopathology of most autoimmune diseases has been well defined, the mechanisms responsible for the breakdown of self-tolerance and which lead to the development of systemic and organ-specific autoaggression are still unclear. Evidence has accumulated which supports a role for a disregulated production of cytokines by leucocytes and possibly other cells in the pathogenesis of some autoimmune diseases. However, due to the complexity and heterogeneity of cytokine effects in the regulation of the immune response, it is difficult to determine whether abnormalities in the patterns of cytokine production are primary or secondary to the pathological process. Confusion is also caused by the fact that the biological activities of cytokines are multiple and often overlapping, and consequently it is difficult to focus on a unique effect of any one cytokine. Characterization of the potential and actual involvement of cytokines is important not only for a better understanding of the pathogenesis of autoimmune conditions, but particularly because of the implications for the development of immunotherapeutic strategies for the prevention and treatment of the diseases. [ABSTRACT FROM AUTHOR]

Cytokines are known to play an important role in autoimmunity and have been suggested to be involved in the pathogenesis of insulin-dependent diabetes (IDDM), In the present study we have measured IL-1, IL-2, IL-4, IL-6, interferon-gamma (IFN-γ) and tumour necrosis factor (TNF) (using both immunoassays and bioassays) in sera from 50 patients affected by IDDM at the time of clinical diagnosis and 51 age and sex matched controls. Detectable levels of IL-1, IL-2, IL-6 and IFN-γ were found in the serum of a small percentage of subjects and were not significantly different between patients and controls, IL-4 was detectable in a higher number of both patients and controls and circulating TNF-α (> 1 U/ml) was found in a percentage of patients (24%) significantly higher than controls (P<0.01), Raised levels of TNF-α were detectable using an immunoenzymatic assay whereas TNF bioactivity in these samples was negligible. We conclude that the presence of immunoreactive TNF-α in the patient's sera may reflect an increased localized production of this cytokine at pancreatic level. However, the measurement in serum of other cytokines does not add information on the role that they may play in the pathogenesis of IDDM. [ABSTRACT FROM AUTHOR]

A patient with a history of at least 10 years 'common' variable hypogammaglobulinaemia seroconverted to HIV-I and became hypergammaglobulinaemic. The HIV isolated from his blood did not polyclonally activate B cells from normal donors. The mechanism of the hypergammaglobulinaemia is discussed. [ABSTRACT FROM AUTHOR]

Murine monoclonal antibodies have been produced against a single human IgE myeloma preparation. A single fusion yielded six different monoclonal antibodies which bound strongly to the immunogen. Two of these failed to react with 'normal' human IgE or with a second IgE paraprotein. The other four antibodies reacted with normal and myelomaderived IgE and were found to be specific for antibodies of this class. Two of these antibodies were potent reagents for the detection of IgE by two site immunoradiometric assay, immunoblotting, and radioallergosorbent test. The other two antibodies were considerably less potent reagents when used in these assay systems. Our findings suggest that care should be taken in selection of suitable monoclonal antibodies for immunochemical study of allergens and sera from allergic patients. [ABSTRACT FROM AUTHOR]