Targeted silencing of DNA-specific B cells combined with partial plasma cell depletion displays additive effects on delaying disease onset in lupus-prone mice.

Targeting autoreactive B lymphocytes at any stage of their differentiation could yield viable therapeutic strategies for treating autoimmunity. All currently used drugs, including the most recently introduced biological agents, lack target specificity. Selective silencing of double-stranded DNA-specific B cells in animals with spontaneous lupus has been achieved previously by the administration of a chimeric antibody molecule that cross-links their DNA-reactive B cell immunoglobulin receptors with inhibitory FcγIIb (CD32) receptors. However, long-lived plasmacytes are resistant to this chimeric antibody as well as to all conventional treatments. Bortezomib (a proteasome inhibitor) depletes most plasma cells and has been shown recently to suppress disease activity in lupus mice. We hypothesized that the co-administration of non-toxic doses of bortezomib, that partially purge long-lived plasma cells, together with an agent that selectively silences DNA-specific B cells, should have additive effects in an autoantibody-mediated disease. Indeed, our data show that the simultaneous treatment of lupus-prone MRL/lpr mice with suboptimal doses of bortezomib plus the chimeric antibody resulted in the prevention or the delayed appearance of the disease manifestations as well as in a prolonged survival. The effect of the combination therapy was significantly stronger than that of the respective monotherapies and was comparable to that observed after cyclophosphamide administration.

Target silencing of disease-associated B-lymphocytes by chimeric molecules in SCID model of pristane-induced autoimmunity.

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the generation of autoantibodies against a diverse array of self-antigens. The B cells producing immunoglobulin G (IgG) antibodies to double-stranded DNA appear to play a main role in the disease progression. Their specific elimination is a reasonable mechanism for effective therapy of SLE. The presently used approaches for silencing autoreactive disease-associated B cells are nonspecific and more precise therapies are needed. We have previously constructed a chimeric protein molecule consisting of several DNA-mimotope peptides coupled to a rat monoclonal anti-mouse CD32 (FcγRIIb) antibody. The mineral oil pristane induces a lupus-like syndrome in non-autoimmune mice leading to the development of glomerulonephritis and lupus-associated autoantibodies. In the present paper, using a pristane-induced autoimmune model in SCID mice, we analyzed the ability of the chimeric antibody to suppress selectively the autoreactive B lymphocytes by cross-linking B-cell surface immunoglobulin receptors with the inhibitory IgG FcγRIIb receptors. Treatment with DNA-like chimeric molecules inhibited B- and T-cell proliferation, restricted the number of anti-DNA antibody-producing cells and suppressed the generation of IgG anti-DNA antibodies. In contrast, phosphate buffered saline (PBS)-injected control mice experienced an increase of disease-associated antibody levels and developed glomerulonephritis similar to pristane-treated donor Balb/c mice.

Effects of nutrients on the production of AK-111-81 macrolide antibiotic by Streptomyces hygroscopicus.

The influence of different carbon and nitrogen sources on the production of AK-111-81 nonpolyenic macrolide antibiotic by Streptomyces hygroscopicus 111-81 was studied. Substitution of glucose with lactose or glycerol significantly affected maximal antibiotic AK-111-81 productivity as the growth rate was close to that of the basal fermentation medium. Addition of ammonium succinate to the fermentation medium markedly increased the antibiotic productivity as the growth rate was low. Divalent ions as Mn2+, Cu2+, Fe2+ stimulated AK-111-81 antibiotic biosynthesis. These results allow us to develop a new fermentation medium showing 6-fold increase of AK-111-81 antibiotic formation compared with the basal fermentation medium.

A new antibiotic, TH818, and its properties.

Antibiotic TH818 was isolated and purified from a culture broth of Streptomyces fulvoviolaceus 818 by extraction and reversed-phase liquid chromatography. The antibiotic TH818 possesses high molecular mass and unique structure, constructed from carbohydrates, amino acids and lipids. The hydrolysis experiments showed the presence of six fatty acids (hexadecanoic, hexadecenoic, octadecanoic, iso-nonadecanoic, docosanoic and pentacosanoic acids), four sugars (glucose, galactose, N -acetylglucosamine and N -acetylgalactosamine), and seven amino acids (threonine, alanine, glutamic acid, glycine, serine and two unidentified). TH818 has a broad spectrum of anti-microbial activity against Gram-positive and Gram-negative bacteria, yeasts and fungi.

Physiological and antagonistic potential of actinomycetes from loquat rhizosphere.

The microflora of Eriobotrya japonica (loquat) was studied by the serial dilutions-spread plate method. A variety of bacteria, fungi and actinomycetes was detected. The genus Steptomyces dominated in the population (83%). A great species diversity was found among Streptomycetes. Some of the actinomycetes possessed antibacterial activity (47%), others produced different exo-enzymes.

Fatty acid composition of Streptomyces hygroscopicus strains producing antibiotics.

The fatty acid composition from mycelia of Streptomyces hygroscopicus strains was studied. A significant proportion of C18:2 was found in cultures. High levels of C16:0, iso-C18:0 and C18:1 were also detected in all S. hygroscopicus strains. The different representatives of S. hygroscopicus had almost the same proportion of unsaturated fatty acids. Certain shifts in the amount of iso, anteiso and straight-chain fatty acids in some cultures were revealed. This might be explained by the adaptation capability of strains belonging to one species to form a variety of available fatty acids determined by particular cell membrane composition favouring certain antibiotic biosynthesis.

[Interaction of actinomycetes in relation to an intensification of protease biosynthesis]. / Vzaimodeistvie vnutri assotsiatsii aktinomitsetov v sviazi s intensifikatsiei biosinteza proteaz.

This work was aimed at studying the interactions during the growth of Actinomyces rimosus producing proteases and Actinomyces violocinereus which did not synthesize secreted proteolytic enzymes. The production of proteases in the association of the actinomycetes was shown to be stimulated by metabolic products released by A. violocinereus into the surrounding medium. The stimulating agent from the cultural broth of this culture accelerated differentiation of the mycelium of the first hyphal generation in A. rimosus, decelerated spore formation of the second hyphal generation, inhibited the growth rate, and increased the rate of protease accumulation as well as the productivity of the synthesis.

[Characteristics of the ultrastructural organization of Actinomyces rimosus and Actinomyces violocinereus in monocultures and in an association producing extracellular proteases]. / Osobennosti ul'trastrukturnoi organizatsii Actinomyces rimosus i Actinomyces violocinereus v monokul'turakh i v assotsiatsii, produtsiruiushchei vnekletochnye proteazy.

The ultrastructural organization of Actinomyces rimosus and Actinomyces violocinereus was compared in monocultures and in associations. The cells of the two species can be discriminated by certain cytological characteristics. A rimosus predominated under the studied conditions and periods of growth. This organism had growth processes disordered (intrahyphal growth). A. violocinereus was characterized by the following processes in the association: peptidoglycan hypersynthesis, formation of calloses of the cell wall which occurred in parallel to hypertrophy of mesosomes, a loss of the capability to form capsules, and delayed spore formation. The reduced synthesis of granular and fibrillar material indicated that these products were not associated with exoprotease. The enzymatic activity was higher and could be detected in earlier in the association than in the monoculture of A. rimosus.

[Optimization of the composition of the medium for thrombolytic enzyme biosynthesis in pure and mixed cultures]. / Optimizatsiia sostava sredy dlia biosinteza tromboliticheskikh fermentov v chistoi i smeshannykh kul'turakh.

The medium for the biosynthesis of exoproteases with the thrombolytic action in monocultures and mixed cultures of actinomycetes was optimized in several steps using the method of complete and fractional factor experiments. When Actinomyces rimosus was cultivated on the selected medium, the productivity of the mycelium rose, the fibrinolytic activity increased three-fold, and the caseinolytic activity became 2.2 times greater. During mixed cultivation of A. rimosus and A. violaceus on this medium, fibrinolysis increased six times comparing with the A. rimosus monoculture growth on the original medium while caseinolysis became only 3.7 times greater, which increased the specificity for fibrin twice.