concanavalin-a and Lyme-Disease

Rapid diagnosis of Lyme borreliosis has been carried out on chemically modified porous polyethylene sinter bodies. Photografting of 2-propenol on sinter body's surface was performed as a first step, introducing active hydroxyl groups as a result of polyalcohol formation. The hydroxyl groups were used for further immobilization and could be linked via 3-aminopropyltriethoxysilane (APTES) to polysaccharides like mannan. Prone to coupling, mannan was activated using N, N'-disuccinimidyl carbonate (DSC) to allow smooth reaction with the primary amine groups of the silane layer. In a final preparation step, a recombinant fusion protein consisting of the mannan-binding domain of the lectin Concanavalin A (ConA) and a specific Borrelia surface antigen was immobilized by self-organization on the mannan surface. The fusion protein was used as biological interface structure. This strategy is highly efficient and resulted in a defined orientation of the antigen part of the fusion protein. Rapid and convenient differentiation could be then established between Borrelia-negative and a -positive serum even in 1000-fold diluted samples and detection of Lyme borreliosis in a rather early stage is likely. Furthermore, this generic strategy can be easily transferred to other bacterial or viral antigen structures.

Topics: Antigens, Bacterial; Borrelia; Concanavalin A; Lyme Disease; Mannans; Particle Size; Polyethylene; Porosity; Recombinant Fusion Proteins; Surface Properties

We recently described the inhibition of host B lymphocytes by Ixodes ricinus tick saliva. In this study, we characterized the factor responsible for this activity and examined the modulation of lipopolysaccharide (LPS)- and Borrelia burgdorferi outer surface protein (Osp)-induced proliferation of naive murine B lymphocytes by an enriched fraction of this factor. The B-lymphocyte inhibitory activity was destroyed by trypsin treatment, indicating that a proteinaceous factor was responsible for this activity. The removal of glutathione-S-transferase (GST) from tick salivary glands extracts (SGE) showed that this B-cell inhibitory protein (BIP) was not a GST. Gel filtration liquid chromatography indicated that BIP has a native molecular weight of approximately 18,000. An enrichment protocol, using a combination of anion-exchange and reverse-phase liquid chromatography, was established. BIP-enriched fractions did not suppress T-cell proliferation. Delayed addition of BIP-enriched fractions, up to 7 hr after LPS addition, inhibited the proliferation of isolated B cells. BIP-enriched fractions dramatically inhibited both OspA- and OspC-induced proliferation of isolated B cells. These results strongly suggest that BIP may facilitate B. burgdorferi transmission by preventing B-cell activation, and also highlights the potential of BIP as a therapeutic agent in B-cell maladies.

Topics: Animals; Antigens, Bacterial; Antigens, Surface; B-Lymphocytes; Bacterial Outer Membrane Proteins; Bacterial Vaccines; Borrelia burgdorferi; Cell Division; Concanavalin A; Female; Immune Tolerance; Ixodes; Lipopolysaccharides; Lipoproteins; Lyme Disease; Lymphocyte Activation; Lymphokines; Molecular Weight; Saliva; Spleen

Tick-borne pathogens would appear to be vulnerable to vertebrate host immune responses during the protracted duration of feeding required by their vectors. However, tick salivary components deposited during feeding may inhibit hemostasis and induce immunosuppression. The mode of action and the nature of immunosuppressive salivary components remains poorly described. We determined that saliva from the main vector of the agent of Lyme disease, Ixodes dammini, profoundly inhibited splenic T cell proliferation in response to stimulation with concanavalin A or phytohemagglutin, in a dose-dependent manner. In addition, interleukin 2 secretion by the T cells was markedly diminished by saliva. Tick saliva also profoundly suppressed nitric oxide production by macrophages stimulated with lipopolysaccharide. Finally, we analyzed the molecular basis for the immunosuppressive effects of saliva and discovered that the molecule in saliva responsible for our observations was not PGE2, as hypothesized by others, but rather, was a protein of 5,000 mol wt or higher.

Topics: Animals; Concanavalin A; Dinoprostone; Female; Interleukin-2; Lyme Disease; Lymphocyte Activation; Macrophages; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Nitric Oxide; Saliva; Suppressor Factors, Immunologic; T-Lymphocytes; Ticks