Direct and monocyte-induced innate immune response of human lung epithelial cells to Brucella abortus infection

Although Brucella frequently infects humans through inhalation, its interaction with pulmonary cells has been overlooked. We examined whether human lung epithelial cells produce proinflammatory mediators in response to Brucella infection. Infection with smooth or rough strains of Brucella abortus induced the secretion of IL-8 and GM-CSF by the bronchial epithelial cell lines Calu-6 and 16HBE14o-, but not by the alveolar epithelial cell line A549. Infected Calu-6 cells also produced low levels of MCP-1. Since monocyte-derived cytokines may induce chemokine secretion in epithelial cells, cocultures of human monocytes (THP-1 cell line) and respiratory epithelial cells were used to study such interaction. IL-8 and MCP-1 levels in B. abortus-infected THP-1:A549 and THP-1:Calu-6 cocultures, and MCP-1 levels in THP-1:16HBE14o- cocultures, were higher than those detected in infected epithelial monocultures. Conditioned medium from infected monocytes induced the secretion of IL-8 and/or MCP-1 by A549 and Calu-6 cells, and these effects were mainly mediated by IL-1 (in A549 cells) or TNF-α (in Calu-6 cells). Conversely, culture supernatants from Brucella-infected bronchial epithelial cells induced MCP-1 production by monocytes, an effect largely mediated by GM-CSF. This study shows that human lung epithelial cells mount a proinflammatory response to Brucella, either directly or after interaction with Brucella-infected monocytes.     

The role of toll-like receptors in immune tolerance induced by Helicobacter pylori infection

Helicobacter pylori (H. pylori) is a gram-negative, microaerobic bacterium that colonizes the gastric mucosa in about half of the world's population. H. pylori infection can lead to various diseases. Chronic infection by H. pylori exposes the gastric mucosa to bacterial components such as lipopolysaccharide (LPS), outer membrane vesicles (OMVs), and several toxic proteins. Infected with H. pylori activates the release of pro-inflammatory factors and triggers inflammatory responses that damage the gastric mucosa. As the only microorganism that permanently colonizes the human stomach, H. pylori can suppress host immunity to achieve long-term colonization. Toll-like receptors (TLRs) play a crucial role in T-cell activation, promoting innate immune responses and immune tolerance during H. pylori infection. Among the 10 TLRs found in humans, TLR2, TLR4, TLR5, and TLR9 have been thoroughly investigated in relation to H. pylori-linked immune regulation. In the present review, we provide a comprehensive analysis of the various mechanisms employed by different TLRs in the induction of immune tolerance upon H. pylori infection, which will contribute to the research of pathogenic mechanism of H. pylori.     

Positional cloning of Lps, and the general role of toll-like receptors in the innate immune response

In mice (and by inference, in all mammals), a single pathway exists to serve lipopolysaccharide (LPS) signal transduction, and as such, allelic mutations at a single locus entirely abolish responses to LPS in C3H/HeJ and C57BL/10ScCr mice. Positional cloning of this locus, known as Lps, revealed that mutations of the Toll-like receptor 4 gene (Tlr4) are responsible for endotoxin resistance. A quick succession of studies have shown Tlr4 to be the critical transmembrane component of the LPS signal transduction complex. As LPS sensing by Tlr4 depends on physical contact between the two molecules, Tlr4 is a direct interface with the microbial world. Eight other molecules with strong similarity to Tlr4 are presently known in mammals, and taking Tlr4 as a model, all may be guessed to participate in the early detection of invasive pathogens. Acting together, the Toll-like receptors may be assumed to present macrophages with a comprehensive "picture" of the micobial world, and thus comprise the principal sensing molecules utilized by cells of the innate immune system.     

The role of scavenger receptors in the innate immune system

Akey aspect of the innate immune system is the ability to discriminate between self and infectious nonself. This is achieved through pattern recognition receptors which directly recognise molecular epitopes expressed by microbes. Scavenger receptors (SRs) have been studied primarily due to their ability to bind and internalise modified lipoproteins, suggesting an important role in foam cell formation and the pathogenesis of atherosclerosis. However, the ability of some SRs to function as pattern recognition receptors through their binding of a wide variety of pathogens indicates a potential role in host defence. This review will detail our current understanding of the function of SRs in innate immunity, and in the initiation of aquired immune responses.     

Host innate immune receptors and beyond: making sense of microbial infections

The complexity of the immune system mirrors its manifold mechanisms of host-microbe interactions. A relatively simplified view was posited after the identification of host innate immune receptors that their distinct mechanisms of sensing "microbial signatures" create unique molecular switches to trigger the immune system. Recently, more sophisticated and cooperative strategies for these receptors have been revealed during receptor-ligand interactions, trafficking, and intra- and intercellular signaling, in order to deal with a diverse range of microbes. Continued mapping of the complex networks of host-microbe interactions may improve our understanding of self/non-self discrimination in immunity and its intervention.     

Brucella abortus DNA is a major bacterial agonist to activate the host innate immune system

Immunity against Brucella abortus depends on the recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs). Signaling pathways triggered by Brucella DNA involves TLR9, AIM2 and possibly STING and MAVS. Herein, we review the advances in B. abortus DNA sensing by host innate immune receptors and the progress in this field.     

Experimental Brucella abortus infection in wolves

Four juvenile male wolves (Canis lupus) each received an oral dose of 1.6-1.7 x 10(12) colony-forming units of Brucella abortus biovar 1 isolated from a bison (Bison bison) in Wood Buffalo National Park (Canada), and two others served as negative controls. Infected wolves did not show clinical signs of disease but did develop high Brucella antibody titers. Small numbers of B. abortus were excreted sporadically in feces until day 50 postinoculation (PI). Very small numbers of the bacterium were isolated from urine of only one wolf late on the same day that it was infected, and very small numbers of colonies of B. abortus were obtained from buccal swabs of three wolves for up to 48 hr PI. Two infected wolves euthanized 6 mo after the start of the experiment had no lesions, and colonies of B. abortus were isolated from thymus and most major lymph nodes. The other two infected wolves euthanized 12 mo after the start of the experiment had no lesions, and smaller numbers of brucellae were recovered from fewer lymph nodes compared with the wolves killed 6 mo earlier. The sporadic excretion of very small numbers of brucellae by the wolves was insignificant when compared with the infective dose for cattle. Brucella abortus, brucellosis, Canis lupus, pathogenesis, serology, wolf.     

Significance of the functional activity of toll-like receptors and other receptors of the innate immune system of the kidney physiology

The kidney needs to defend against microbial pathogens in order to maintain normal structure and function. This is achieved through innate and adaptive components of the immune system. For a long time, immunologists were concentrating on the adaptive immune system, which, as a result, was studied in detail; at the same time, the significance of the innate immune system was underestimated. This gap was partly filled in the recently, when the key role of the innate immune system in fighting microorganisms and in activating and regulating the adaptive immune system was convincingly established. In the first part of the present article, the sense apparatus of the innate immune system (the so-called pattern-recognition receptors) will be reviewed; particular attention will be paid to the toll-like receptors (TLRs), which bear the main burden of microorganism recognition. Signalling pathways that are activated by TLRs and result in the activation of effector mechanisms will also be reviewed. In the second part of the review, we will analyse available data on how these mechanisms of the innate immune system secure defence and normal functioning of the kidney.     

Toll样受体（TLR）介导的天然免疫间的相互调节

模式识别受体（PRR）在宿主细胞识别与抵御微生物病原体中起到了重要作用。Toll样受体（TLR）是研究比较清楚的一类PRR,可以识别多种病原体成份,启动天然免疫反应。此外,近来发现了几类其他模式识别受体,如C型凝集素受体（CLR）,核苷酸寡聚结合域（NOD）样受体（NLR）和视黄酸诱导基因I（RIG—I）样受体（RLR）,表明机体的天然免疫反应受到多种机制的精密调控。本文着重综述TLR与其他PRR在识别病原体和介导天然免疫信号通路间的相互关系。     

The role of the vacB gene in the pathogenesis of Brucella abortus

Brucella species are important zoonotic pathogens affecting a wide variety of mammals. Therefore, the identification of new Brucella virulence factors is of great interest in understanding bacterial pathogenesis and immune evasion. In this study, we have identified Brucella abortus vacB gene that presents 2343 nucleotides and 781 amino acids and it shows 39% identity with Shigella flexneri vacB gene that encodes an exoribonuclease RNase R involved in bacterial virulence. Further, we have inactivated Brucella vacB by gene replacement strategy generating a deletion mutant strain. In order to test the role of Brucella vacB in pathogenesis, BALB/c and interferon regulatory factor-1 (IRF-1) knockout (KO) mice received Brucella vacB mutant, the virulent parental strain 2308 or the vaccine strain RB51 and the bacterial CFU numbers in spleens and mous survival were monitored. Our results demonstrated that the B. abortus DeltavacB mutant and the wild type strain 2308 showed similar CFU numbers in BALB/c mice. Additionally, IRF-1 KO mice that received either the vacB mutant or S2308 strain died in 12-14 days postinfection; in contrast, all animals that received the RB51 vaccine strain survived for 30 days postinoculation. In summary, this study reports that the vacB gene in B. abortus has no impact on bacterial pathogenesis.     

Central role of MyD88-dependent dendritic cell maturation and proinflammatory cytokine production to control Brucella abortus infection

Brucella abortus is a facultative intracellular bacterium that infects humans and domestic animals. The enhanced susceptibility to virulent B. abortus observed in MyD88 knockout (KO) mice led us to investigate the mechanisms involved in MyD88-dependent immune responses. First, we defined the role of MyD88 in dendritic cell (DC) maturation. In vitro as well as in vivo, B. abortus-exposed MyD88 KO DCs displayed a significant impairment on maturation as observed by expression of CD40, CD86, and MHC class II on CD11c+ cells. In addition, IL-12 and TNF-alpha production was totally abrogated in MyD88 KO DCs and macrophages. Furthermore, B. abortus-induced IL-12 production was found to be dependent on TLR2 in DC, but independent on TLR2 and TLR4 in macrophages. Additionally, we investigated the role of exogenous IL-12 and TNF-alpha administration on MyD88 KO control of B. abortus infection. Importantly, IL-12, but not TNF-alpha, was able to partially rescue host susceptibility in MyD88 KO-infected animals. Furthermore, we demonstrated the role played by TLR9 during virulent B. abortus infection. TLR9 KO-infected mice showed 1 log Brucella CFU higher than wild-type mice. Macrophages and DC from TLR9 KO mice showed reduced IL-12 and unaltered TNF-alpha production when these cells were stimulated with Brucella. Together, these results suggest that susceptibility of MyD88 KO mice to B. abortus is due to impaired DC maturation and lack of IL-12 synthesis. Additionally, DC activation during Brucella infection plays an important regulatory role by stimulating and programming T cells to produce IFN-gamma.     

The effects of superovulation on Brucella abortus infection in the bovine uterus

Uterine flushings were collected three times at predetermined intervals from 11 mixed-breed beef cows and cultured for Brucella abortus . Prior to sampling, all cows had aborted fetuses from which brucellae had been isolated. Initial collections were made between 21 and 34 days following abortion. The second flushing was conducted at the onset of injections used for inducing superovulation and the third flushing was conducted 6 to 8 days after the ensuing estrus. The latter two flushes were conducted between 60 and 120 days following abortion. Brucellae were isolated from uterine flushings collected from 6 of the 11 cows on the initial round of sampling. Cultures of all subsequent uterine flushings collected before and after injections for superovulation were negative. It was concluded that the superovulatory treatment is not likely to reactivate the release of brucellae into the uterine lumen during the period when embryos are normally collected.     

The role of the innate immune system of the liver in the control of HBV and HCV

Hepatitis B virus (HBV) and Hepatitis C virus (HCV) infection are among the most frequent causes of chronic liver disease worldwide. As recent studies suggested that Toll like receptor (TLR)-based therapies may represent a promising approach in the treatment of HBV infection, we have studied the role of the local innate immune system of the liver as possible mediator of this effect. Murine non-parenchymal liver cells (NPC; Kupffer cells, KC; sinusoidal endothelial cells, LSEC) were isolated from C57/BL6 and stimulated by TLR 1-9 agonists. Supernatants were harvested and assayed for their antiviral activity against HBV in HBV-Met cells and HCV in the murine HCV replicon cell line MH1. Only supernatants from TLR 3 and -4 stimulated KC and TLR 3 stimulated LSEC were able to potently suppress HBV and HCV replication. By using neutralizing antibodies we could demonstrate that the TLR 3- but not the TLR 4 mediated effect is exclusively mediated through IFN-β. Our data indicate that TLR 3 and -4 mediated stimulation of NPC leads to production of IFN-β which can potently suppress HBV and HCV replication. This is of relevance for the local control of viral hepatitis infection by the innate immune system of the liver, the development of novel TLR-based therapeutic approaches and sheds new light on the viral crosstalk between HCV (TLR 3 stimulator) and HBV.     

Experimental infection of captive axis deer with Brucella abortus

Four captive-raised axis deer, Axis axis (Erxleben), which were negative serologically to Brucella were inoculated with 1 X 10(8) virulent Brucella abortus biotype 1 organisms (Texas #221 isolate) administered bilaterally into the conjunctival sac. Sera collected from each deer prior to inoculation and 30 days post-inoculation (PI) were examined for Brucella antibodies by the buffered Brucella antigen (card), the rivanol precipitation, the standard tube agglutination, and the cold complement fixation tube serologic tests. All four axis deer converted serologically as determined by all tests at 30 days PI. Brucella abortus biotype 1 was isolated from 26 of 32 tissue samples collected at necropsy and also from milk from the lactating female.     

The role of dendritic cells in the innate immune system

Dendritic cells (DCs) are bone-marrow-derived leucocytes that are specialised antigen-presenting cells capable of stimulating a primary T-lymphocyte response to specific antigen. In this chapter we discuss the role DCs play in the innate response acting as a critical link with the adaptive response and the influence of the innate response on dendritic cells.     

Immunologic consequences of Francisella tularensis live vaccine strain infection: role of the innate immune response in infection and immunity

Francisella tularensis (Ft), a Gram-negative intracellular bacterium, is the etiologic agent of tularemia. Although attenuated for humans, i.p. infection of mice with <10 Ft live vaccine strain (LVS) organisms causes lethal infection that resembles human tularemia, whereas the LD50 for an intradermal infection is >10(6) organisms. To examine the immunological consequences of Ft LVS infection on the innate immune response, the inflammatory responses of mice infected i.p. or intradermally were compared. Mice infected i.p. displayed greater bacterial burden and increased expression of proinflammatory genes, particularly in the liver. In contrast to most LPS, highly purified Ft LVS LPS (10 microg/ml) was found to be only minimally stimulatory in primary murine macrophages and in HEK293T cells transiently transfected with TLR4/MD-2/CD14, whereas live Ft LVS bacteria were highly stimulatory for macrophages and TLR2-expressing HEK293T cells. Despite the poor stimulatory activity of Ft LVS LPS in vitro, administration of 100 ng of Ft LVS LPS 2 days before Ft LVS challenge severely limited both bacterial burden and cytokine mRNA and protein expression in the absence of detectable Ab at the time of bacterial challenge, yet these mice developed a robust IgM Ab response within 2 days of infection and survived. These data suggest that prior administration of Ft LVS LPS protects the host by diminishing bacterial burden and blunting an otherwise overwhelming inflammatory response, while priming the adaptive immune response for development of a strong Ab response.