Suppressors of Cytokine Signaling (SOCS)1 and SOCS3 Proteins Are Mediators of Interleukin-10 Modulation of Inflammatory Responses Induced by Chlamydia muridarum and Its Major Outer Membrane Protein (MOMP) in Mouse J774 Macrophages.

The immunopathology of chlamydial diseases is exacerbated by a broad-spectrum of inflammatory mediators, which we reported are inhibited by IL-10 in macrophages. However, the chlamydial protein moiety that induces the inflammatory mediators and the mechanisms by which IL-10 inhibits them are unknown. We hypothesized that Chlamydia major outer membrane protein (MOMP) mediates its disease pathogenesis, and the suppressor of cytokine signaling (SOCS)1 and SOCS3 proteins are mediators of the IL-10 inhibitory actions. Our hypothesis was tested by exposing mouse J774 macrophages to chlamydial stimulants (live Chlamydia muridarum and MOMP) with and without IL-10. MOMP significantly induced several inflammatory mediators (IL-6, IL-12p40, CCL5, CXCL10), which were dose-dependently inhibited by IL-10. Chlamydial stimulants induced the mRNA gene transcripts and protein expression of SOCS1 and SOCS3, with more SOCS3 expression. Notably, IL-10 reciprocally regulated their expression by reducing SOCS1 and increasing SOCS3. Specific inhibitions of MAPK pathways revealed that p38, JNK, and MEK1/2 are required for inducing inflammatory mediators as well as SOCS1 and SOCS3. Chlamydial stimulants triggered an M1 pro-inflammatory phenotype evidently by an enhanced nos2 (M1 marker) expression, which was skewed by IL-10 towards a more M2 anti-inflammatory phenotype by the increased expression of mrc1 and arg1 (M2 markers) and the reduced SOCS1/SOCS3 ratios. Neutralization of endogenously produced IL-10 augmented the secretion of inflammatory mediators, reduced SOCS3 expression, and skewed the chlamydial M1 to an M2 phenotype. Inhibition of proteasome degradation increased TNF but decreased IL-10, CCL5, and CXCL10 secretion by suppressing SOCS1 and SOCS3 expressions and dysregulating their STAT1 and STAT3 transcription factors. Our data show that SOCS1 and SOCS3 are regulators of IL-10 inhibitory actions, and underscore SOCS proteins as therapeutic targets for IL-10 control of inflammation for Chlamydia and other bacterial inflammatory diseases.

Cu(II) Complexes with FomA Protein Fragments of Fusobacterium Nucleatum Increase Oxidative Stress and Malondialdehyde Level.

An explanation of carcinogenesis processes may certainly contribute to the prevention and development of novel methods for cancer treatment. In this paper, we considered the probable relationship between the presence of Fusobacterium nucleatum in the colon and its possible influence on the development of colorectal cancer. For this purpose, intracellular and/or extracellular generation of reactive oxygen species (ROS) by mouse colon carcinoma cells (CT26) was stimulated by two fragments of FomA adhesin from F. nucleatum and their complexes with copper(II): Cu(II)-Ac-KGHGNG-NH2 (1Cu) and Cu(II)-Ac-PTVHNE-NH2 (2Cu). Incubation of the cells with copper complexes was followed with ICP-MS technique. The overall generation of ROS was shown by means of fluorescence spectroscopy with two proper probes, whereas identification of ROS was achieved by the spin trapping technique and electron paramagnetic resonance measurements. As a result, an abundant production of the hydroxyl radicals, both inside and outside the cells, was observed upon the stimulation of the CT26 cells with the copper complexes. Clearly both compounds induced strong oxidation stress which triggered a radicals' cascade that finally resulted in the pronounced lipid peroxidation. The latter was evidenced with the measured level of malondialdehyde, a biomarker of the peroxidation process. By applying N-acetylcysteine antioxidant to the studied system, the free radical mechanism of the lipid peroxidation process was confirmed. Hypothetically this mechanism can lead to colon cell damage and further cancerogenesis processes.

Brucella abortus-activated microglia induce neuronal death through primary phagocytosis.

Inflammation has long been implicated as a contributor to pathogenesis in neurobrucellosis. Many of the associated neurocognitive symptoms of neurobrucellosis may be the result of neuronal dysfunction resulting from the inflammatory response induced by Brucella abortus infection in the central nervous system. In this manuscript, we describe an immune mechanism for inflammatory activation of microglia that leads to neuronal death upon B. abortus infection. B. abortus was unable to infect or harm primary cultures of mouse neurons. However, when neurons were co-cultured with microglia and infected with B. abortus significant neuronal loss occurred. This phenomenon was dependent on TLR2 activation by Brucella lipoproteins. Neuronal death was not due to apoptosis, but it was dependent on the microglial release of nitric oxide (NO). B. abortus infection stimulated microglial proliferation, phagocytic activity and engulfment of neurons. NO secreted by B. abortus-activated microglia induced neuronal exposure of the "eat-me" signal phosphatidylserine (PS). Blocking of PS-binding to protein milk fat globule epidermal growth factor-8 (MFG-E8) or microglial vitronectin receptor-MFG-E8 interaction was sufficient to prevent neuronal loss by inhibiting microglial phagocytosis without affecting their activation. Taken together, our results indicate that B. abortus is not directly toxic to neurons; rather, these cells become distressed and are killed by phagocytosis in the inflammatory surroundings generated by infected microglia. Neuronal loss induced by B. abortus-activated microglia may explain, in part, the neurological deficits observed during neurobrucellosis.

B7-H3 augments the inflammatory response and is associated with human sepsis.

B7-H3, a new member of the B7 superfamily, acts as both a T cell costimulator and coinhibitor, and thus plays a key role in the regulation of T cell-mediated immune responses. However, it is unclear whether B7-H3 is involved in the innate immune monocyte/macrophage-mediated inflammatory response. In this paper, we show that, although B7-H3 alone failed to stimulate proinflammatory cytokine release from murine macrophages, it strongly augmented both LPS- and bacterial lipoprotein-induced NF-kappaB activation and inflammatory response. This occurred in both a TLR4- and TLR2-dependent manner. Blockage of B7-H3 in vivo attenuated LPS-induced proinflammatory cytokine release and endotoxic shock-related lethality. Furthermore, we found that patients diagnosed with sepsis, in contrast to healthy individuals, exhibited significant levels of raised plasma soluble B7-H3 (sB7-H3) and that this level correlated with the clinical outcome and levels of plasma TNF-alpha and IL-6. In addition, a putative receptor for B7-H3 was detected on monocytes and peritoneal macrophages from septic patients but not on monocytes from healthy donors. Stimulation of human monocytes with LPS and inflammatory cytokines led to a substantial release of sB7-H3. Taken together, our data indicate that significantly elevated plasma sB7-H3 in septic patients may predict a poor outcome. Furthermore, we demonstrate that B7-H3 functions as a costimulator of innate immunity by augmenting proinflammatory cytokine release from bacterial cell wall product-stimulated monocytes/macrophages and may contribute positively to the development of sepsis.

Molecular and virulence characteristics of an outer membrane-associated RTX exoprotein in Pasteurella pneumotropica.

BACKGROUND: Pasteurella pneumotropica is a ubiquitous bacterium that is frequently isolated from laboratory rodents and causes various clinical symptoms in immunodeficient animals. Currently two RTX toxins, PnxIA and PnxIIA, which are similar to hemolysin-like high-molecular-weight exoproteins are known in this species. In this study, we identified and analyzed a further RTX toxin named PnxIIIA and the corresponding type I secretion system. RESULTS: The RTX exoprotein, PnxIIIA, contains only a few copies of the RTX repeat-like sequence and 3 large repeat sequences that are partially similar to the outer membrane protein found in several prokaryotes. Recombinant PnxIIIA protein (rPnxIIIA) was cytotoxic toward J774A.1 mouse macrophage cells, whereas cytotoxicity was attenuated by the addition of anti-CD11a monoclonal antibody. rPnxIIIA could bind to extracellular matrices (ECMs) and cause hemagglutination of sheep erythrocytes. Binding was dependent on the 3 large repeat sequences in PnxIIIA. Protein interaction analyses indicated that PnxIIIA is mainly localized in the outer membrane of P. pneumotropica ATCC 35149 in a self-assembled oligomeric form. PnxIIIA is less cytotoxic to J774A.1 cells than PnxIA and PnxIIA. CONCLUSIONS: The results implicate that PnxIIIA is located on the cell surface and participates in adhesion to ECMs and enhanced hemagglutination in the rodent pathogen P. pneumotropica.

Cytotoxic effects of Kingella kingae outer membrane vesicles on human cells.

Kingella kingae is an emerging pathogen causing osteoarticular infections in pediatric patients. Electron microscopy of K. kingae clinical isolates revealed the heterogeneously-sized membranous structures blebbing from the outer membrane that were classified as outer membrane vesicles (OMVs). OMVs purified from the secreted fraction of a septic arthritis K. kingae isolate were characterized. Among several major proteins, K. kingae OMVs contained virulence factors RtxA toxin and PilC2 pilus adhesin. RtxA was also found secreted as a soluble protein in the extracellular environment indicating that the bacterium may utilize different mechanisms for the toxin delivery. OMVs were shown to be hemolytic and possess some leukotoxic activity while high leukotoxicity was detected in the non-hemolytic OMV-free component of the secreted fraction. OMVs were internalized by human osteoblasts and synovial cells. Upon interaction with OMVs, the cells produced increased levels of human granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 6 (IL-6) suggesting that these cytokines might be involved in the signaling response of infected joint and bone tissues during natural K. kingae infection. This study is the first report of OMV production by K. kingae and demonstrates that OMVs are a complex virulence factor of the organism causing cytolytic and inflammatory effects on host cells.

Effect of Leptospira interrogans outer membrane proteins LipL32 on HUVEC.

Leptospira cause disease through a toxin-mediated process by inducing vascular injury, particularly a small-vessel vasculitis. Breakdown of vessel endothelial cell integrity may increase vessel permeability which is correlated with the changes of tight junction and/or apoptosis in vessel endothelial cells. The specific toxin responsible remains unidentified. In this study, we amplified outer membrane protein LipL32 from the genome of Leptospira interrogans serovar Lai, and it was subcloned in pET32a(+) vector to express thioredoxin(Trx)-LipL32 fusion protein in Escherichia coli BL21(DE3). The protein was expressed and purified, and Trx-LipL32 was administered to culture with human umbilical vein endothelial cells (HUVEC) to elucidate the role of leptospiral outer membrane proteins in vessel endothelial cell. The purified recombinant protein was capable to increase the permeability of HUVECs. And the protein was able to decrease the expression of ZO-1 and induce F-actin in HUVECs display thickening and clustering. Moreover, apoptosis of HUVEC was significantly accelerated. But the fusion partner had no effect in these regards. It is possible that LipL32 is involved in the vessel lesions.

OmpA-like protein Loa22 from Leptospira interrogans serovar Lai is cytotoxic to cultured rat renal cells and promotes inflammatory responses.

Leptospirosis renal disease is one of the common clinical manifestations of leptospirosis, including acute renal failure and tubulointerstitial nephritis. Outer membrane protein A-like protein Loa22 is a lipoprotein from Leptospira interrogans and has been suggested to be a corresponding virulence factor. However, the role of Loa22 in leptospiral nephropathy is not yet understood. In the present study, we constructed a vector and artificially expressed Loa22 in Escherichia coli BL21(DE)pLysS cells. After extensive purification, along with a GST tag protein control, Loa22 protein was used to test the cytotoxicity in cultured rat proximal tubule cells (NRK52E) and examine its effects on the induction of inflammatory responses. Using morphological examination, 2,3-bis(2-methoxy-4- nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H-tetrazoium hydrixide absorbance, lactate dehydrogenase assays and an analysis of apoptosis via flow cytometry, it was found that Loa22 protein mediates a direct cytotoxic effect on NRK52E cells in a dose-dependent manner. Using real-time PCR, western blotting and immunofluorescence, it was found that Loa22 protein upregulates the expression of toll-like receptor 2 (TLR2), induces nitric oxide synthase and promotes the production of nitric oxide (NO) and monocyte chemoattractant protein-1 (MCP-1) by NRK52E cells. Additionally, using a TLR2 blocking antibody, it was found that enhanced NO and MCP-1 production by NRK52E cells after Loa22 stimulation requires the activation of TLR2. Collectively, our data suggested that Loa22 is a critical virulence factor of L. interrogans and is involved in the leptospiral nephropathy through mediating direct cytotoxicity and enhancing inflammatory responses.

Group A streptococcal M-protein specific antibodies and T-cells drive the pathology observed in the rat autoimmune valvulitis model.

Acute rheumatic fever (ARF) and rheumatic heart disease (RHD) are autoimmune mediated diseases triggered by group A streptococcal (GAS) infections. Molecular mimicry between GAS M-proteins and host tissue proteins has been proposed as the mechanism that initiates autoreactive immune responses in ARF/RHD. However, the individual role of antibodies and T-cells specific for GAS M-proteins in the pathogenesis of autoimmune carditis remains under-explored. The current study investigated the role of antibodies and T-cells in the development of carditis in the Lewis rat autoimmune valvultis (RAV) model by transferring serum and/or splenic T-cells from rats previously injected with GAS recombinant M5 protein. Here we report that serum antibodies alone and serum plus in vitro expanded rM5-specific T-cells from hyperimmune rats were capable of transferring carditis to naïve syngeneic animals. Moreover, the rats that received combined serum and T-cells developed more severe carditis. Recipient rats developed mitral valvulitis and myocarditis and showed prolongation of P-R intervals in electrocardiography. GAS M5 protein-specific IgG reactivity and T-cell recall response were also demonstrated in recipient rats indicating long-term persistence of antibodies and T-cells following transfer. The results suggest that both anti-GAS M5 antibodies and T-cells have differential propensity to induce autoimmune mediated carditis in syngeneic rats following transfer. The results highlight that antibodies and effector T-cells generated by GAS M protein injection can also independently home into cardiac tissue to cross-react with tissue proteins causing autoimmune mediated immunopathology.

Retinoic acid pre-treatment down regulates V. cholerae outer membrane vesicles induced acute inflammation and enhances mucosal immunity.

Bacterial outer membrane vesicles have been extensively investigated and considered as a next generation vaccine. Recently, we have demonstrated that the cholera pentavalent outer membrane vesicles (CPMVs) immunogen induced adaptive immunity and had a strong protective efficacy against the circulating V. cholerae strains in a mouse model. In this present study, we are mainly focusing on reducing outer membrane vesicle (OMV) -mediated toxicity without altering its antigenic property. Therefore, we have selected All-trans Retinoic Acid (ATRA), active metabolites of vitamin A, which have both anti-inflammatory and mucosal adjuvant properties. Pre-treatment of ATRA significantly reduced CPMVs induced TLR2 mediated pro-inflammatory responses in vitro and in vivo. Furthermore, we also found ATRA pre-treatment significantly induced mucosal immune response and protective efficacy after two doses of oral immunization with CPMVs (75µg). This study can help to reduce OMV based vaccine toxicity and induce better protective immunity where children and men suffered from malnutrition mainly in developing countries.

Nontoxic outer membrane vesicles efficiently increase the efficacy of an influenza vaccine in mice and ferrets.

In this study, we developed a further-modified outer membrane vesicle (fmOMV) from the ΔmsbB/ΔpagP mutant of Escherichia coli transformed with the plasmid, pLpxF, in order to use it as an adjuvant for pandemic H1N1 (pH1N1) influenza vaccine. We evaluated the efficacy of the pH1N1 influenza vaccine containing the fmOMV in animal models as compared to the commercial adjuvants, alum or AddaVaxTM. The fmOMV-adjuvanted pH1N1 influenza vaccine induced a significant increase in the humoral immunity; however, this effect was less than that of the AddaVaxTM. The fmOMV-adjuvanted vaccine displayed pronounced an enhanced protective efficacy with increased T cell immune response and reduced the viral load in the lungs of the infected mice after challenging them with a lethal dose of the homologous virus. Moreover, it resulted in a significantly higher cross-protection against heterologous virus challenge than that of the pH1N1 vaccine with alum or with no adjuvants. In ferrets, the fmOMV-adjuvanted vaccine elicited a superior antibody response based on the HI titer and efficiently protected the animals from the lethal viral challenges. Taken together, the nontoxic fmOMV could be a promising adjuvant for inducing robust T cell priming into the pH1N1 vaccine and might be broadly applicable to the development of preventive measures against influenza virus infection.

Escherichia coli Braun Lipoprotein (BLP) exhibits endotoxemia - like pathology in Swiss albino mice.

The endotoxin lipopolysaccharide (LPS) promotes sepsis, but bacterial peptides also promote inflammation leading to sepsis. We found, intraperitoneal administration of live or heat inactivated E. coli JE5505 lacking the abundant outer membrane protein, Braun lipoprotein (BLP), was less toxic than E. coli DH5α possessing BLP in Swiss albino mice. Injection of BLP free of LPS purified from E. coli DH5α induced massive infiltration of leukocytes in lungs and liver. BLP activated human polymorphonuclear cells (PMNs) ex vivo to adhere to denatured collagen in serum and polymyxin B independent fashion, a property distinct from LPS. Both LPS and BLP stimulated the synthesis of platelet activating factor (PAF), a potent lipid mediator, in human PMNs. In mouse macrophage cell line, RAW264.7, while both BLP and LPS similarly upregulated TNF-α and IL-1ß mRNA; BLP was more potent in inducing cyclooxygenase-2 (COX-2) mRNA and protein expression. Peritoneal macrophages from TLR2-/- mice significantly reduced the production of TNF-α in response to BLP in contrast to macrophages from wild type mice. We conclude, BLP acting through TLR2, is a potent inducer of inflammation with a response profile both common and distinct from LPS. Hence, BLP mediated pathway may also be considered as an effective target against sepsis.

Random mutagenesis identifies a C-terminal region of YopD important for Yersinia type III secretion function.

A common virulence mechanism among bacterial pathogens is the use of specialized secretion systems that deliver virulence proteins through a translocation channel inserted in the host cell membrane. During Yersinia infection, the host recognizes the type III secretion system mounting a pro-inflammatory response. However, soon after they are translocated, the effectors efficiently counteract that response. In this study we sought to identify YopD residues responsible for type III secretion system function. Through random mutagenesis, we identified eight Y. pseudotuberculosis yopD mutants with single amino acid changes affecting various type III secretion functions. Three severely defective mutants had substitutions in residues encompassing a 35 amino acid region (residues 168-203) located between the transmembrane domain and the C-terminal putative coiled-coil region of YopD. These mutations did not affect regulation of the low calcium response or YopB-YopD interaction but markedly inhibited MAPK and NFκB. [corrected] activation. When some of these mutations were introduced into the native yopD gene, defects in effector translocation and pore formation were also observed. We conclude that this newly identified region is important for YopD translocon function. The role of this domain in vivo remains elusive, as amino acid substitutions in that region did not significantly affect virulence of Y. pseudotuberculosis in orogastrically-infected mice.

Phase 1 testing of detoxified LPS/group B meningococcal outer membrane protein vaccine with and without synthetic CPG 7909 adjuvant for the prevention and treatment of sepsis.

BACKGROUND: Gram-negative bacteria (GNB) are a leading cause of nosocomial infection and sepsis. Increasing multi-antibiotic resistance has left clinicians with fewer therapeutic options. Antibodies to GNB lipopolysaccharide (LPS, or endotoxin) have reduced morbidity and mortality as a result of infection and are not subject to the resistance mechanisms deployed by bacteria against antibiotics. In this phase 1 study, we administered a vaccine that elicits antibodies against a highly conserved portion of LPS with and without a CpG oligodeoxynucleotide (ODN) TLR9 agonist as adjuvant. METHODS: A vaccine composed of the detoxified LPS (dLPS) from E. coli O111:B4 (J5 mutant) non-covalently complexed to group B meningococcal outer membrane protein (OMP). Twenty healthy adult subjects received three doses at 0, 29 and 59 days of antigen (10 µg dLPS) with or without CPG 7909 (250 or 500 µg). Subjects were evaluated for local and systemic adverse effects and laboratory findings. Anti-J5 LPS IgG and IgM antibody levels were measured by electrochemiluminesence. Due to premature study termination, not all subjects received all three doses. RESULTS: All vaccine formulations were well-tolerated with no local or systemic events of greater than moderate severity. The vaccine alone group achieved a ≥ 4-fold "responder" response in IgG and IgM antibody in only one of 6 subjects. In contrast, the vaccine plus CPG 7909 groups appeared to have earlier and more sustained (to 180 days) responses, greater mean-fold increases, and a higher proportion of "responders" achieving ≥ 4-fold increases over baseline. CONCLUSIONS: Although the study was halted before all enrolled subjects received all three doses, the J5dLPS/OMP vaccine, with or without CpG adjuvant, was safe and well-tolerated. The inclusion of CpG increased the number of subjects with a ≥ 4-fold antibody response, evident even after the second of three planned doses. A vaccine comprising J5dLPS/OMP antigen with CpG adjuvant merits further investigation. CLINICAL TRIALS REGISTRATION: ClinicalTrials.gov Identifier: NCT01164514.

Harpin-induced hypersensitive cell death is associated with altered mitochondrial functions in tobacco cells.

Mitochondria play important roles in animal apoptosis and are implicated in salicylic acid (SA)-induced plant resistance to viral pathogens. In a previous study, we demonstrated that SA induces rapid inhibition of mitochondrial electron transport and oxidative phosphorylation in tobacco cells. In the present study, we report that plant programmed cell death induced during pathogen elicitor-induced hypersensitive response (HR) is also associated with altered mitochondrial functions. Harpin, an HR elicitor produced by Erwinia amylovora, induced inhibition of ATP synthesis in tobacco cell cultures. Inhibition of ATP synthesis occurred almost immediately after incubation with harpin and preceded hypersensitive cell death induced by the elicitor. Diphenylene iodonium, an inhibitor of the oxidative burst, did not block harpin-induced inhibition of ATP synthesis or cell death, suggesting that oxidative burst was not the direct cause for these two harpin-induced processes. Unlike SA, harpin had no significant effect on total respiratory O2 uptake of treated cells. However, respiration of harpin-treated tobacco cells became very sensitive to the alternative oxidase inhibitors salicyl-hydroxamic acid and n-propyl gallate. Thus, harpin treatment resulted in reduced capacity of mitochondrial cytochrome pathway electron transport, which could lead to the observed inhibition of ATP synthesis. Given the recently demonstrated roles of mitochondria in apoptosis, this rapid inhibition of mitochondrial functions may play a role in harpin-induced hypersensitive cell death.

Porphyromonas gingivalis surface components induce interleukin-1 release and tyrosine phosphorylation in macrophages.

The aim of the present study was to characterize the responses of macrophages to surface antigens of Porphyromonas gingivalis. Native fimbriae, full-length recombinant fimbrillin, and a lectin-like 12-kDa antigen all stimulated BALB/c peritoneal macrophages to secrete interleukin (IL)-1 beta. The antigens induced similar patterns of tyrosine phosphorylation; proteins in approximately 35-46 kDa range of undetermined identities were phosphorylated in the macrophages. The abilities of the surface antigens to induce IL-1 beta were markedly attenuated by tyrosine kinase inhibitors. This inhibition correlated with inhibition of the induced phosphorylation of specific macrophage proteins at tyrosine. The data suggest that tyrosine kinase(s) plays an important role in the regulatory intracellular signaling mechanisms by which P. gingivalis surface antigens can mediate certain responses in macrophages.

Some biological properties of outer membrane proteins of Hafnia.

Outer membrane proteins (OMP) isolated from four antigenically different strains of Hafnia alvei were tested for the toxicity, pyrogenicity, ability to induce Shwartzman reaction as well as for their influence on the leukocyte system. LD50 doses for the studied preparations determined on inbred mice were 18, 20, 28 and 34 mg/kg. These differences in the toxicity of the preparations were reflected in manifestation of Shwartzman reaction; more toxic preparations induced strongest necrohemorrhagic changes at the site of injection. The OMP preparations injected intravenously to rabbits caused moderate increase of body temperature. They induced changes in the number of leukocytes in the animals comparable with those of other preparates of bacterial origin.

Some biological properties of outer membrane proteins of Shigella.

Outer membrane proteins (OMP) derived from two antigenically different representatives of Shigella (Sh. flexneri 3a and Sh. sonnei phase I) were tested for the toxicity, pyrogenicity, ability to induce Shwartzman reaction as well as for their influence on the leukocyte system. LD50 dose determined on mice was 28 mg/kg for OMP of Sh. flexneri and 23 mg/kg for OMP of Sh. sonnei. Both preparations injected intravenously to rabbits caused moderate increase of body temperature, expressed by the value 1.8 degrees C. Intravenous administration of protein preparations to rabbits, induced at first leukopenia and then transient leukocytosis. When injected subcutaneously in the dose of 500 micrograms and after 24 h intravenously in the dose 100 micrograms, they produced slight hemorrhagic changes at the site of administration.

[The protective properties of a porin from the outer membrane of Yersinia pseudotuberculosis]. / Protektivnye svoistva porina iz vneshnei membrany Yersinia pseudotuberculosis.

The protective properties of the species-specific pore-forming polypeptide of Y. pseudotuberculosis outer membrane (porin) were studied. The study showed that 80-90% of mice immunized with porin survived after challenge with 10-30 LD50 of Y. pseudotuberculosis, serovars 1 and 3. The LD50 of the preparation exceeded its ED50 more than 100-fold. Immunization made in two injections was more effective than immunization in one injection. The immunization of the animals with porin led to an increase in the activity of peritoneal exudate macrophages with respect to Y. pseudotuberculosis, serovars 1 and 3.

[Components of the outer membrane of Yersinia pseudotuberculosis and their role in the pathogenesis of pseudotuberculosis]. / Komponenty naruzhnoi membrany Yersinia pseudotuberculosis i ikh rol' v patogeneze psevdotuberkuleza.

The pore-forming protein, lipopolysaccharide-protein complex and lipopolysaccharide of Y. pseudotuberculosis outer membrane have been shown to participate in the penetration of the bacteria into the cells of the macroorganism, to produce a toxic effect on these cells and to enhance the ingesting activity of macrophages in small doses, while suppressing it in large doses. When introduced parenterally, protein induces a more pronounced clinical picture of specific reactive hepatitis in experimental animals and greater changes in their kidneys than the lipopolysaccharide--protein complex.