ArticleLiterature Review

Exotoxins and endotoxins: Inducers of inflammatory cytokines

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Abstract

Endotoxins and exotoxins are among the most potent bacterial inducers of cytokines. During infectious processes, the production of inflammatory cytokines including tumor necrosis factor (TNF), interleukin-1β (IL-1β), gamma interferon (IFNγ) and chemokines orchestrates the anti-infectious innate immune response. However, an overzealous production, leading up to a cytokine storm, can be deleterious and contributes to mortality consecutive to sepsis or toxic shock syndrome. Endotoxins of Gram-negative bacteria (lipopolysaccharide, LPS) are particularly inflammatory because they generate auto-amplificatory loops after activation of monocytes/macrophages. LPS and numerous pore-forming exotoxins also activate the inflammasome, the molecular platform that allows the release of mature IL-1β and IL-18. Among exotoxins, some behave as superantigens, and as such activate the release of cytokines by T-lymphocytes. In most cases, pre-exposure to exotoxins enhances the cytokine production induced by LPS and its lethality, whereas pre-exposure to endotoxin usually results in tolerance. In this review we recall the various steps, which, from the very early discovery of pyrogenicity induced by bacterial products, ended to the discovery of the endogenous pyrogen. Furthermore, we compare the specific characteristics of endotoxins and exotoxins in their capacity to induce inflammatory cytokines.

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... The most plausible explanation for increased amounts of interleukins in semen samples of the GO and LO group may partially lie in an active recognition of bacterial proteins, which results in the activation of the sperm TLRs. This process initiates a series of reactions that leads to the cleavage of caspase 1, essential for IL-1 secretion [60]. Moreover, Fraczek et al. [5,50] indicated that pro-inflammatory cytokines promote ROS production to a level at which significant peroxidative damage occurs to the sperm membrane. ...
... A promising alternative could lie in the supplementation of antimicrobial peptides that are part of the innate immune system. The use of such molecules presents with a significantly lower risk for resistance development since these induce bacterial cell lysis in a non-specific manner [9,60]. ...
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This study focused on the identification of naturally occurring bacteria in the reproductive fluid and impact on the quality of ejaculates obtained from the turkey breed British United Turkeys (BUT) Big 6 (n = 60). We determined possible relationships between the bacterial load and advanced sperm quality parameters that are important for effective artificial insemination and high fertility, as well as the concentration of selected antimicrobial proteins and pro-inflammatory markers of turkey semen. Sperm motility was assessed with computer-assisted sperm analysis (CASA), while the membrane and acrosome integrity were examined with smearing and staining methods. Reactive oxygen species (ROS) generation was quantified via luminometry, sperm DNA fragmentation was evaluated using the TUNEL assay, and the JC-1 assay was applied to evaluate the mitochondrial membrane potential. Cell lysates were prepared to investigate the extent of lipid and protein oxidation. Furthermore, levels of interleukins 1 and 6 (IL-1, IL-6), C-reactive protein, cathelicidin, and β-defensin were quantified in the seminal plasma using the ELISA method. The most dominant species identified by the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry was Escherichia coli, Proteus mirabilis, Staphylococcus lentus, and Citrobacter braakii. The bacterial load had a negative effect on the sperm motility (p < 0.001), as well as membrane (p < 0.05) and acrosome integrity (p < 0.01). A strong positive relationship between the bacterial load and DNA fragmentation (p < 0.001) was detected as well. Positive associations were recorded between the increasing presence of bacteria, ROS overgeneration (p < 0.001), and a subsequent oxidative damage to the proteins (p < 0.001) and lipids (p < 0.01). It was revealed that the antimicrobial peptides β-defensin (p < 0.001) and cathelicidin (p < 0.001) had a positive relationship with the motility. In contrast, pro-inflammatory markers, such as IL-1 (p < 0.001) and IL-6 (p < 0.001), had a negative impact on the motion behavior of turkey spermatozoa. Our results suggest that the semen quality may be notably affected by the bacterial quantity as well as quality. It seems that bacteriospermia is associated with inflammatory processes, oxidative stress, sperm structural deterioration, and a subsequent risk for a failed artificial insemination in turkey breeding.
... LPS is a component in the outer wall of gramnegative bacteria [36]. Injecting LPS into animals can produce endotoxin shock pathological changes, and endotoxic shock is one of the most serious manifestations of sepsis. ...
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Sepsis is a type of systemic inflammation response syndrome that leads to organ function disorders. Currently, there is no specific medicine for sepsis in clinical practice. Lipopolysaccharide (LPS) is an important endotoxin that causes sepsis. Here, we report an effective two-drug combination therapy to treat LPS-induced liver and kidney injury in endotoxic rats. Ulinastatin (UTI) and Thrombomodulin (TM) are biological macromolecules extracted from urine. In our study, combination therapy significantly improved LPS-induced liver and kidney pathological structure and functional injury, and significantly improved the survival rate of endotoxic rats. Results of TUNEL staining and Western blot showed that UTI combined with TM inhibited the excessive apoptosis of liver and kidney cells caused by LPS. The drug combination also promoted the proliferation of liver and kidney cells, reduced the levels of pro-inflammatory factors interleukin (IL)-6, IL-1β, tumor or necrosis factor (TNF)-α and nitric oxide, and down-regulated the expression of High Mobility Group Box 1 (HMGB1), Toll-like receptor (TLR) 4 and Nuclear Factor (NF)-κB phosphorylation to inhibit inflammation. In addition, the combination of UTI and TM also promoted the production of a variety of antioxidant enzymes in the tissues and inhibited the production of lipid peroxidation malondialdehyde (MDA) to enhance antioxidant defenses. Our experiments also proved that UTI combined with TM did not reduce the anticoagulant effect of TM. These results suggested that UTI combined with TM can improve endotoxin-induced liver and kidney damage and mortality by inhibiting liver and kidney cell apoptosis, promoting proliferation, and inhibiting inflammation and oxidative injury.
... Lipopolysaccharide (LPS) is one of the most effective CRS inducers. It can activate monocytes/macrophages to release cytokines, including tumor necrosis factor (TNF-α), interleukin-1β (IL-1β), IL-6, chemokines, and other inflammatory cytokines (Cavaillon, 2018). Not only that, LPS can induce sepsis and toxic shock syndrome in mice. ...
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... In addition to these so-called endotoxins, extracellular toxic proteins known as exotoxins, are produced and secreted by pathogens via specific secretion machinery. Unlike the extensive systemic damage of endotoxins that is mainly based on an adverse immune reaction, exotoxins typically target local tissues and are often restricted to particular cell types or receptors (Cavaillon, 2018). For other virulence factors, their identification and in-depth characterization is fundamental to the development of specific diagnostics and treatment tools. ...
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Bacterial fish pathogens are one of the key challenges in the aquaculture industry, one of the fast-growing industries worldwide. These pathogens rely on arsenal of virulence factors such as toxins, adhesins, effectors and enzymes to promote colonization and infection. Translocation of virulence factors across the membrane to either the extracellular environment or directly into the host cells is performed by single or multiple dedicated secretion systems. These secretion systems are often key to the infection process. They can range from simple single-protein systems to complex injection needles made from dozens of subunits. Here, we review the different types of secretion systems in Gram-negative bacterial fish pathogens and describe their putative roles in pathogenicity. We find that the available information is fragmented and often descriptive, and hope that our overview will help researchers to more systematically learn from the similarities and differences between the virulence factors and secretion systems of the fish-pathogenic species described here.
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This study investigated the effects of dietary Flos populi extract (FPE) on the growth, antioxidation capability, innate immune response, and disease resistance in gibel carp. A total of 480 fish were fed with five different diets containing 0, 0.5, 1.0, 1.5, or 2.0 g kg⁻¹ FPE (designated as control, D0.5, D1.0, D1.5, or D2.0 groups) for 45 days. The fish were challenged with A. hydrophila after the feeding trial. Compared with the control, the feed efficiency (FE), weight gain (WG), final body weight (FBW), and specific growth rate (SGR) were significantly improved in groups D1.0 and D1.5. Dietary FPE significantly increased serum superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) activities, as well as glutathione (GSH) content. The contents of protein carbonyl (PCC) and malondialdehyde (MDA) in serum decreased significantly. Additionally, FPE supplementation in diets resulted in significant improvement in serum lysozyme (LZM) and myeloperoxidase (MPO) activities, as well as immunoglobulin M (IgM) and complement 3 (C3) concentrations. The hepatic antioxidant enzymes (CAT and SOD) activities increased, whereas content of MDA decreased in fish treated with dietary FPE than those of control both pre- and post-challenged. After 12 h-challenge, an obvious downregulation of hepatic Kelch-like-ECH-associated protein 1 (Keap1), splenic tumor necrosis factor-α (TNF-α), interleukin (IL)-8, IL-1β, and toll-like receptor 2 (TLR2) mRNA levels was observed in fish treated with dietary FPE, whereas hepatic Nrf2 transcription level was upregulated compared to the control. Furthermore, compared to group D0.5, higher relative percent survival (RPS) was observed in gibel carp fed dietary 1.0–2.0 g/kg FPE. Our results reveal that FPE supplemented diet has a stimulatory effect on antioxidant capacity and nonspecific immune response, along with improved growth performance and enhanced resistance against A. hydrophila infection in juvenile gibel carp.
... The toxins produced by pathogenic bacteria can result in life-threatening infections. There are two categories of bacterial toxins, endotoxins and exotoxins, which are the leading causes of cytokine outbreaks induced by a bacterial infection (Cavaillon, 2018). Neutralizing toxins helps to disarm pathogens and is a direct way to treat symptoms of infection, because of toxins' destructive nature and the vital role they play in different pathogenic mechanisms (Cegelski et al., 2008;Rasko & Sperandio, 2010). ...
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Despite the enormous success of antibiotics in antimicrobial therapy, the rapid emergence of antibiotic resistance and the complexity of the bacterial infection microenvironment make traditional antibiotic therapy face critical challenges against resistant bacteria, antitoxin, and intracellular infections. Consequently, there is a critical need to design antimicrobial agents that target infection microenvironment and alleviate antibiotic resistance. Cell membrane-coated nanoparticles (CMCNPs) are biomimetic materials that can be obtained by wrapping the cell membrane vesicles directly onto the surface of the nanoparticles (NPs) through physical means. Incorporating the biological functions of cell membrane vesicles and the superior physicochemical properties of NPs, CMCNPs have shown great promise in recent years for targeting infections, neutralizing bacterial toxins, and designing bacterial infection vaccines. This review highlights topics where CMCNPs present great value in advancing the treatment of bacterial infections, including drug delivery, detox-ification, and vaccination. Lastly, we discuss the future hurdles and prospects of translating this technique into clinical practice, providing a comprehensive review of the technological developments of CMCNPs in the treatment of bacterial infections.
... Exotoxins are secreted by organisms, mainly diffusion proteins secreted by Gram-positive bacteria into the surrounding medium. Endotoxin is a structural component of organisms and an inducer of inflammatory cytokines [131], as shown in Figure 10. The most common endotoxin that workers are exposed to is a lipopolysaccharide. ...
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The application of cutting fluid in the field of engineering manufacturing has a history of hundreds of years, and it plays a vital role in the processing efficiency and surface quality of parts. Among them, water-based cutting fluid accounts for more than 90% of the consumption of cutting fluid. However, long-term recycling of water-based cutting fluid could easily cause deterioration, and the breeding of bacteria could cause the cutting fluid to fail, increase manufacturing costs, and even endanger the health of workers. Traditional bactericides could improve the biological stability of cutting fluids, but they are toxic to the environment and do not conform to the development trend of low-carbon manufacturing. Low-carbon manufacturing is inevitable and the direction of sustainable manufacturing. The use of nanomaterials, transition metal complexes, and physical sterilization methods on the bacterial cell membrane and genetic material could effectively solve this problem. In this article, the mechanism of action of additives and microbial metabolites was first analyzed. Then, the denaturation mechanism of traditional bactericides on the target protein and the effect of sterilization efficiency were summarized. Further, the mechanism of nanomaterials disrupting cell membrane potential was discussed. The effects of lipophilicity and the atomic number of transition metal complexes on cell membrane penetration were also summarized, and the effects of ultraviolet rays and ozone on the destruction of bacterial genetic material were reviewed. In other words, the bactericidal performance, hazard, degradability, and economics of various sterilization methods were comprehensively evaluated, and the potential development direction of improving the biological stability of cutting fluid was proposed.
... Increase of endogenous reactive oxygen species (ROS) is one of the important factors that damage the body. Although ROS benefit physiological processes, such as removal of infectious pathogens, healing, and repair of tissues (Tovar-y-Romo et al., 2016;Cavaillon, 2018), nevertheless, excessive levels can damage tissues (Yu et al., 2015;Scialo et al., 2017). When exposed to ionizing radiation for treatment, it does not only direct toward tumor cells but also affects the surrounding microenvironment (Haimovitz-Friedman et al., 2020). ...
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Background: Oxidative stress induced by radiation causes variable expression of antioxidant enzymes in a tissue-specific manner. Testicular tissues carry out the complex process of spermatogenesis, and studies indicate that testicular damages due to irradiation require long-term recovery before complete resumption. Ionizing radiation also causes oxidative stress in tissues, leading to testicular damage. Aims and Objectives: This study measured differential expression of antioxidant enzymes following administration of C. borivilianum root extract (CRB) in response to irradiation-induced oxidative stress. The activity of various important endogenous enzymatic defense systems was evaluated and correlated for strength of association. Materials and method: Two forms of C. borivilianum (CB) extracts [CB alone and CB-silver nanoparticles (AgNPs)] were administered at a dose of 50 mg/kg body weight to Swiss albino male mice for 7 consecutive days. After that, they were irradiated with 6 Gy irradiation and further used to study various parameters of antioxidant enzymes. Results: Results indicate a significant increase in the level of glutathione (GSH) and the activity of GSH-related antioxidant enzymes in irradiated mice treated with CRE and CRE-AgNPs (silver nanoparticles biosynthesized using C. borivilianum root extract) in comparison to non-pretreated ones (groups I and II). Reciprocal elevation was observed in related enzymes, that is, glutathione S-transferase activity (GST), glutathione reductase (GR), and glutathione peroxidase activity (GPx). Elevation in the activity of catalase (CAT) and superoxide dismutase (SOD) was also evident in both the irradiated groups pretreated with CRE-AgNPs. However, expression of CAT in the CRE-treated irradiated group was similar to that of the non-treated irradiated group. Higher association among CAT-SOD, CAT-GPx, and GR-GST was observed. Conclusion: Overall, it was observed that testicular cells post-irradiation in all groups go through intense oxidative stress; however, groups pretreated with CRE or CRE-AgNPs indicated better toleration and resumption of antioxidant capacity. CRE or CRE-AgNPs pretreated non-irradiated groups mostly remained within the control range indicating stimulated expression of antioxidants.
... A primary designation in the classification of gram-negative bacteria is the presence of lipopolysaccharide (LPS), instead of LTA, as part of the outer cell membrane. LPS, also known as endotoxin, is one of the most potent immunostimulants (Alexander and Rietschel, 2001) and consists of a glycan polymer, oligosaccharide core, and membrane-anchor lipid (Alexander and Rietschel, 2001;Cavaillon, 2018). Physiological recognition of the lipid component by the immune system causes pro-inflammatory cytokine activity (Alexander and Rietschel, 2001;Liu et al., 2018;Monguio-Tortajada et al., 2018;Tawfik et al., 2020). ...
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Cell-based mathematical models have previously been developed to simulate the immune system in response to pathogens. Mathematical modeling papers which study the human immune response to pathogens have predicted concentrations of a variety of cells, including activated and resting macrophages, plasma cells, and antibodies. This study aims to create a comprehensive mathematical model that can predict cytokine levels in response to a gram-positive bacterium, S. aureus by coupling previous models. To accomplish this, the cytokines Tumor Necrosis Factor Alpha (TNF- α ), Interleukin 6 (IL-6), Interleukin 8 (IL-8), and Interleukin 10 (IL-10) are included to quantify the relationship between cytokine release from macrophages and the concentration of the pathogen, S. aureus, ex vivo . Partial differential equations (PDEs) are used to model cellular response and ordinary differential equations (ODEs) are used to model cytokine response, and interactions between both components produce a more robust and more complete systems-level understanding of immune activation. In the coupled cellular and cytokine model outlined in this paper, a low concentration of S. aureus is used to stimulate the measured cellular response and cytokine expression. Results show that our cellular activation and cytokine expression model characterizing septic conditions can predict ex vivo mechanisms in response to gram-negative and gram-positive bacteria. Our simulations provide new insights into how the human immune system responds to infections from different pathogens. Novel applications of these insights help in the development of more powerful tools and protocols in infection biology.
... On the one hand, lipid A has been known to affect the immune system, as it is an immunostimulant capable of inducing a protective, non-specific immune response in the host. On the other hand, depending on the dose and the structure of lipid A, the resulting immune response can lead to the uncontrolled production of inflammatory cytokines that may evoke fatal effects, such as high fever, septic shock, and multiorgan failure [2]. In this context, it is not surprising that several lipid A derivatives (synthetic or produced from bacteria) have been targets for developing effective adjuvants in vaccines and immunotherapy formulations [3,4]. ...
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Lipid A, the membrane-bound phosphoglycolipid component of bacteria, is held responsible for the clinical syndrome of gram-negative sepsis. In this study, the fragmentation behavior of a set of synthetic lipid A derivatives was studied by electrospray ionization multistage mass spectrometry (ESI-MSn), in conjunction with tandem mass spectrometry (MS/MS), using low-energy collision-induced dissociation (CID). Genealogical insight about the fragmentation pathways of the deprotonated 4'-monophosphoryl lipid A structural analogs led to proposals of a number of alternative dissociation routes that have not been reported previously. Each of the fragment ions was interpreted using various possible mechanisms, consistent with the principles of reactions described in organic chemistry. Specifically, the hypothesized mechanisms are: (i) cleavage of the C-3 primary fatty acid leaves behind an epoxide group attached to the reducing sugar; (ii) cleavage of the C-3' primary fatty acid (as an acid) generates a cyclic phosphate connected to the nonreducing sugar; (iii) cleavage of the C-2' secondary fatty acid occurs both in acid and ketene forms; iv) the C-2 and C-2' primary fatty acids are eliminated as an amide and ketene, respectively; (v) the 0,2A2 cross-ring fragment contains a four-membered ring (oxetanose); (vi) the 0,4A2 ion is consecutively formed from the 0,2A2 ion by retro-aldol, retro-cycloaddition, and transesterification; and (vii) formations of H2PO4- and PO3- are associated with the formation of sugar epoxide. An understanding of the relation between 0,2A2 and 0,4A2-type sugar fragments and the different cleavage mechanisms of the two ester-linked primary fatty acids is invaluable for distinguishing lipid A isomers with different locations of a single ester-linked fatty acid (i.e., at C-3 or C-3'). Thus, in addition to a better comprehension of lipid A fragmentation processes in mass spectrometers, our observations can be applied for a more precise elucidation of naturally occurring lipid A structures.
... Tumor necrosis factor α (TNFα) is a multifunctional inflammatory cytokine that is secreted mainly by activated macrophages and plays a crucial role in many processes of functioning of both normal and malignant cells, including survival, apoptosis, and necroptosis, as well as intercellular communication [257]. A violation of the regulation of this complex signaling network is a distinctive feature of a wide range of inflammatory diseases [258], in particular, oncological pathologies [259][260][261][262]. TNFα functions by binding to two receptors (TNFR1 and TNFR2 receptor), which leads to the regulation of a number of cytokines, proteases, and growth factor production [263]. ...
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Chronic inflammation and oxidative stress are the interconnected pathological processes, which lead to cancer initiation and progression. The growing level of oxidative and inflammatory damage was shown to increase cancer severity and contribute to tumor spread. The overproduction of reactive oxygen species (ROS), which is associated with the reduced capacity of the endogenous cell defense mechanisms and/or metabolic imbalance, is the main contributor to oxidative stress. An abnormal level of ROS was defined as a predisposing factor for the cell transformation that could trigger pro-oncogenic signaling pathways, induce changes in gene expression, and facilitate accumulation of mutations, DNA damage, and genomic instability. Additionally, the activation of transcription factors caused by a prolonged oxidative stress, including NF-κB, p53, HIF1α, etc., leads to the expression of several genes responsible for inflammation. The resulting hyperactivation of inflammatory mediators, including TNFα, TGF-β, interleukins, and prostaglandins can contribute to the development of neoplasia. Pro-inflammatory cytokines were shown to trigger adaptive reactions and the acquisition of resistance by tumor cells to apoptosis, while promoting proliferation, invasion, and angiogenesis. Moreover, the chronic inflammatory response leads to the excessive production of free radicals, which further aggravate the initiated reactions. This review summarizes the recent data and progress in the discovery of mechanisms that associate oxidative stress and chronic inflammation with cancer onset and metastasis. In addition, the review provides insights for the development of therapeutic approaches and the discovery of natural substances that will be able to simultaneously inhibit several key oncological and inflammation-related targets.
... Sepsis is a severe clinical syndrome whereby the host cannot control the spread of pathogens within the body [75,76]. Cytokine storms resulting from either endotoxin or exotoxininduced cytokine overproduction can lead to harmful effects (e.g., sepsis, toxic shock syn-drome) [77,78]. Sepsis has a mortality rate of nearly 20% and thus is a major public health problem. ...
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Cytokine storm is a phenomenon characterized by strong elevated circulating cytokines that most often occur after an overreactive immune system is activated by an acute systemic infection. A variety of cells participate in cytokine storm induction and progression, with profiles of cytokines released during cytokine storm varying from disease to disease. This review focuses on pathophysiological mechanisms underlying cytokine storm induction and progression induced by pathogenic invasive infectious diseases. Strategies for targeted treatment of various types of infection-induced cytokine storms are described from both host and pathogen perspectives. In summary, current studies indicate that cytokine storm-targeted therapies can effectively alleviate tissue damage while promoting the clearance of invading pathogens. Based on this premise, “multi-omics” immune system profiling should facilitate the development of more effective therapeutic strategies to alleviate cytokine storms caused by various diseases.
... These are negative regulators of the Janus kinase JAK2/STAT pathway [23]. These mechanisms are activated by inflammatory cytokines in LPS-induced sepsis [24]. ...
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Inflammation induces a wide response of the neuroendocrine system, which leads to modifications in all the endocrine axes. The hypothalamic–growth hormone (GH)–insulin-like growth factor-1 (IGF-1) axis is deeply affected by inflammation, its response being characterized by GH resistance and a decrease in circulating levels of IGF-1. The endocrine and metabolic responses to inflammation allow the organism to survive. However, in chronic inflammatory conditions, the inhibition of the hypothalamic–GH–IGF-1 axis contributes to the catabolic process, with skeletal muscle atrophy and cachexia. Here, we review the changes in pituitary GH secretion, IGF-1, and IGF-1 binding protein-3 (IGFBP-3), as well as the mechanism that mediated those responses. The contribution of GH and IGF-1 to muscle wasting during inflammation has also been analyzed.
... The use of gelatin in biomedicine is somehow restricted due to its high affinity to endotoxin contamination. Endotoxins are large and complex lipopolysaccharides that are localised at the outer membrane of Gram-negative bacteria [14,15] and are often associated with contamination of medical devices. Specifically, as endotoxins possess a high thermal stability [16] and are hard to destroy with conventional sterilisation conditions [17], residual endotoxins constitute the most significant pyrogen in parenteral drugs and medical devices [18,19] and frequently lead to post-operative complications, such as delayed tissue regeneration and homeostasis, implant failure and septic shock [20][21][22][23]. ...
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In the medical device sector, bloom index and residual endotoxins should be controlled, as they are crucial regulators of the device’s physicochemical and biological properties. It is also imperative to identify a suitable crosslinking method to increase mechanical integrity, without jeopardising cellular functions of gelatin-based devices. Herein, gelatin preparations with variable bloom index and endotoxin levels were used to fabricate non-crosslinked and polyethylene glycol succinimidyl glutarate crosslinked gelatin scaffolds, the physicochemical and biological properties of which were subsequently assessed. Gelatin preparations with low bloom index resulted in hydrogels with significantly (p < 0.05) lower compression stress, elastic modulus and resistance to enzymatic degradation, and significantly higher (p < 0.05) free amine content than gelatin preparations with high bloom index. Gelatin preparations with high endotoxin levels resulted in films that induced significantly (p < 0.05) higher macrophage clusters than gelatin preparations with low endotoxin level. Our data suggest that the bloom index modulates the physicochemical properties, and the endotoxin content regulates the biological response of gelatin biomaterials. Although polyethylene glycol succinimidyl glutarate crosslinking significantly (p < 0.05) increased compression stress, elastic modulus and resistance to enzymatic degradation, and significantly (p < 0.05) decreased free amine content, at the concentration used, it did not provide sufficient structural integrity to support cell culture. Therefore, the quest for the optimal gelatin crosslinker continues.
... Chewing of SLT not only exposes the buccal mucosa but also its ingestion can lead to serious consequences. Bacterial toxins are inflammagens known to induce cytokine storm; the endotoxins trigger TLR4/MD2 expressed on the surface of monocytes, dendritic cells, and macrophages via CD14, whereas exotoxins induce the release of T-lymphocytes derived cytokines (Cavaillon 2018). Gram-positive and Gram-negative bacteria express hemolysins which are protein toxins that enable pore formation in biological membranes thereby lysing eukaryotic cells (Braun and Focareta 1991). ...
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Smokeless tobacco product (STP) consumption is a significant public health threat across the globe. STPs are not only a storehouse of carcinogens and toxicants but also harbor microbes that aid in the conversion of tobacco alkaloids to carcinogenic tobacco-specific nitrosamines (TSNAs), thereby posing a further threat to the health of its consumers. The present study analyzed the bacterial diversity of popular dry and loose STPs by 16S rRNA gene sequencing. This NGS-based investigation revealed four dominant phyla Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria and identified 549 genera, Prevotella, Bacteroides, and Lactobacillus constituting the core bacteriome of these STPs. The most significantly diverse bacteriome profile was displayed by the loose STP Mainpuri kapoori. The study further predicted the functional attributes of the prevalent genera by Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) algorithm. Genes encoding for nitrate and nitrite reduction and transport enzymes, antibiotic resistance, multi-drug transporters and efflux pumps, secretion of endo- and exotoxin, and other pro-inflammatory molecules were identified. The loose STPs showed the highest level of nitrogen metabolism genes which can contribute to the synthesis of TSNAs. This study reveals the bacteriome of Indian domestic loose STPs that stagger behind in manufacturing and storage stringencies. Our results raise an alarm that the consumption of STPs harboring pathogenic genera can potentially lead to the onset of several oral and systemic diseases. Nevertheless, an in-depth correlation analysis of the microbial diversity of STPs and their elicit impact on consumer health is warranted. Graphical abstract Key points • Smokeless tobacco harbors bacteria that aid in synthesis of carcinogenic nitrosamines. • Most diverse bacteriome profile was displayed by loose smokeless tobacco products. • Pathogenic genera in these products can harm the oral and systemic health of users.
... A variety of inflammatory cytokines are sequentially produced after bacterial infection (called the 'cytokine cascade'), beginning with the production of pro-inflammatory cytokines such as IL-1, TNF-α, and IL-12 [50]. In addition, anti-inflammatory cytokines including IL-4 and TGF-β, which can resolve inflammation caused by pro-inflammatory cytokines, are produced at the lesion. ...
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Despite advances in medicine, mortality due to sepsis has not decreased. Pulsed electromagnetic field (PEMF) therapy is emerging as an alternative treatment in many inflammation-related diseases. However, there are few studies on the application of PEMF therapy to sepsis. In the current study, we examined the effect of PEMF therapy on a mouse model of lipopolysaccharide (LPS)-induced septic shock. Mice injected with LPS and treated with PEMF showed higher survival rates compared with the LPS group. The increased survival was correlated with decreased levels of pro-inflammatory cytokine mRNA expression and lower serum nitric oxide levels and nitric oxide synthase 2 mRNA expression in the liver compared with the LPS group. In the PEMF + LPS group, there was less organ damage in the liver, lungs, spleen, and kidneys compared to the LPS group. To identify potential gene targets of PEMF treatment, microarray analysis was performed, and the results showed that 136 genes were up-regulated, and 267 genes were down-regulated in the PEMF + LPS group compared to the LPS group. These results suggest that PEMF treatment can dramatically decrease septic shock through the reduction of pro-inflammatory cytokine gene expression. In a clinical setting, PEMF may provide a beneficial effect for patients with bacteria-induced sepsis and reduce septic shock-induced mortality.
... It prolongs the presence of M1 macrophages in chronic wounds in two ways. P. aeruginosa produces LPS and binds to the TLR4 receptor complex in macrophages to promote the secretion of inflammatory cytokines (131). Moreover, the type III Secretion System (T3SS) virulence structure is a common trait among all P. aeruginosa clinical isolates, which functions as a conduit to directly translocate effector toxins into the target cells and result in inflammation (132). ...
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Persistent hyper-inflammation is a distinguishing pathophysiological characteristic of chronic wounds, and macrophage malfunction is considered as a major contributor thereof. In this review, we describe the origin and heterogeneity of macrophages during wound healing, and compare macrophage function in healing and non-healing wounds. We consider extrinsic and intrinsic factors driving wound macrophage dysregulation, and review systemic and topical therapeutic approaches for the restoration of macrophage response. Multidimensional analysis is highlighted through the integration of various high-throughput technologies, used to assess the diversity and activation states as well as cellular communication of macrophages in healing and non-healing wound. This research fills the gaps in current literature and provides the promising therapeutic interventions for chronic wounds.
... Because the alteration of the physiological equilibrium of gut microbiota induced by prophylactic antibiotic therapy can affect the efficacy of chemotherapy and immunotherapy, a careful approach should be considered with regard to prophylactic antibiotics for cancer patients. Furthermore, intestinal barrier dysfunction is associated with increased levels of systemic LPS-positive bacterial EVs that induce the secretion of pro-inflammatory mediators (IL-6, IL-8, MCP-1 and MIP-1α) by peripheral blood mononuclear cells [126] with consequent increased risk of sepsis or toxic shock syndrome [127] . Therefore, the use of selective antibiotic therapy to target specific harmful bacterial species and spare those associated with the effectiveness of cancer therapy along with a concomitant administration of probiotics, prebiotics to rebalance the gut microbiota should be taken in consideration [128] . ...
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... The endotoxin of gram-negative bacteria (lipopolysaccharide, LPS) is one of the most potent bacterial inducers of cytokine release, including the pro-inflammatory cytokines TNF-α, IL-1β and IL-6 [43], and gene expression of various other pro-inflammatory markers and factors. Consistent with these data, LPS increased the expression of Cd40 in both WT and KO cultures. ...
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... Endotoxins are highly immunogenic, which could trigger the inflammatory response. An overexpression of cytokines leads to a septic shock and consecutive death (Cavaillon, 2018). Phage preparation should be necessarily purified to ensure the low level of the endotoxin and other bacterial contamination. ...
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... Thus, pro-inflammatory macrophages are potently activated by LPS, leading to the production of a vast number of inflammatory mediators, including oxygen and nitrogen intermediators, prostaglandins, and cytokines (MarcCavaillon 2018). ...
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The human embryonal kidney 293 cell (HEK-293) is a widely used expression host for transient gene expression. The genes or plasmids used for the transient transfections are usually propagated and extracted from the gram-negative bacterium Escherichia coli, the workhorse for molecular biologists. As a gram-negative bacterium E. coli has an outer membrane (OM) containing lipopolysaccharides (LPS) or endotoxins. LPS are very potent inducers of inflammatory cytokines in the body. In early research phases DNA intended for transient transfections is not routinely checked for LPS-levels. In this study we addressed the question whether LPS has an impact on the cultivation and production of a recombinant antibody. At high concentrations the presence of LPS has a detrimental impact on cell viability and recombinant protein expression. But low LPS concentrations are tolerated and might even enhance protein expression levels.
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Sex and age have distinct influences and roles in behavior and immune reactivity; yet, most studies use adult male rodents with little attention to middle age, a time associated with key physiological transitions in both sexes. Thus, this study investigated sex differences during middle age in behavior, immune response to lipopolysaccharide (LPS), and glucose regulation in C57BL/6 mice with GFP-tagged monocytes/microglia. Behaviorally, males performed better in tests of motor function (Open Field [OF], Grip Strength, Sticker Removal, Gait, and Pole tests) and displayed less depressive- and anxiety-like behaviors across multiple mood tests (OF, Elevated Zero Maze, Sucrose Preference, and Swim test). However, females performed better in tests of cognition (Barnes Maze and Novel Object Recognition). Following behavioral assessment, mice were given LPS to characterize sex-dependent inflammagen responses. Females displayed greater sickness behavior in the OF, higher levels of peripheral cytokines, and subtle neuroinflammation in the cortex, striatum, and hippocampus. A separate middle-aged cohort was used for glucose tolerance and insulin sensitivity testing. Both sexes had excessive blood glucose rebound after insulin challenge, but displayed differences following glucose administration, where males had higher baseline glucose and females remained hyperglycemic. This study suggests that during middle-age male mice have better emotional regulation and motor function, but not cognitive ability than females. Further, males are less sensitive than females to the acute effects of LPS peripherally and centrally, but both sexes showed sex-specific impairments in blood glucose regulation. Overall, it appears that middle age is an important transition point with multiple sex differences, some of which are unique to this stage of life.
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This experiment was conducted to investigate the effects of different supplementation levels of palygorskite and probiotics complex (Pal-Pro) on laying performance, hatching performance, egg quality, plasma antioxidative status, and immune response of broiler breeders. A total of 432 Xuefeng black-bone chickens (42-week-old) were randomly assigned into four treatments with six replicates of 18 hens. The broiler breeders were fed the basal diet supplemented with 0, 250, 500, and 750 mg/kg Pal-Pro for 8 weeks. The Pal-Pro supplementation linearly or quadratically increased (p < .05) laying rate, fertilisation rate, hatchability of fertile eggs, heath hatched chicks rate, eggshell thickness, yolk index, total antioxidant capacity, activities of total superoxide dismutase, catalase, glutathione peroxidase, and contents of immunoglobulins, interleukin-2, and tumour necrosis factor-α in plasma. With Pal-Pro supplementation increased, feed convention ratio during 5–8 and 1–8 week, embryo mortality, malondialdehyde linearly decreased (p < .05). This finding suggested that dietary supplementation with 750 mg/kg Pal-Pro administration could improve the antioxidative status and immunoglobulins contents in plasma of broiler breeders, which may contribute to an increase in broiler’s laying rate, hatching performance, egg quality, such as eggshell thickness and yolk index. • HIGHLIGHTS • Dietary supplementation of Pal-Pro decreased the feed conversion ratio and embryo mortality of broiler breeders. • Dietary supplementation of Pal-Pro improved the laying rate, fertility, hatchability of fertile eggs, and yolk index of broiler breeders. • Dietary supplementation of Pal-Pro enhanced the antioxidant capacity and the level of immunoglobulins in the plasma.
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The aim of this study was to investigate the protective effect of trans-anethole (TA) on lipopolysaccharide-induced acute liver inflammation model of chickens by determining the levels of inflammatory mediators in serum and liver, relative mRNA expression and protein expression of inflammation-related genes in NF-κB signaling pathway. A total of 160 one-day-old male chickens (Arbor Acres) were assigned into four treatments with eight replicates of five birds each. On d 20, the control group was intraperitoneally injected with sterile saline and the other groups were injected with lipopolysaccharide (LPS) (5 mg/kg body weight). There were no significant differences in average daily gain (ADG), average daily feed intake (ADFI) and feed conversion ratio (FCR) among groups. However, compared with the control group, the LPS group significantly increased (P < 0.01) the serum levels of interleukin-6 (IL-6), interleukin-1beta (IL-1β), tumor necrosis factor-alpha (TNF-α), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) and decreased (P < 0.01) the interleukin-10 (IL-10) level. TA attenuated (P < 0.01) these increases in IL-1β, TNF-α, ALT and AST levels and improved (P < 0.01) the IL-10 level. In liver, the groups fed with TA had lower (P < 0.01) concentrations of IL-6 and TNF-α as well as higher (P < 0.05) concentration of IL-10. Furthermore, TA down-regulated (P < 0.05) the mRNA expression levels of nuclear factor kappa B p65 (NF-κB p65) and TNF-α, also up-regulated (P < 0.05) IL-10 and inhibitor of NF-κB alpha (IκBα) upon LPS challenge. In protein level, supplementation of 600 mg/kg of TA down-regulated (P < 0.05) and up-regulated (P < 0.05) the protein expression of NF-κB p65 and IκBα, respectively. The present findings suggest that TA could alleviate the acute liver inflammation induced by LPS via blocking the activation of NF-κB and the 600 mg/kg of TA plays more fruitful role in protecting broilers against LPS stimulus.
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Rheumatoid arthritis (RA) is a systemic autoimmune disease which could not only cause inflammatory reaction, but also cause cartilage and bone damage. The clinical treatment is mainly aimed at anti-inflammatory and immunotherapy. Targeting both at inflammation and bone damage repairing will be great significance for RA therapy. Triptolide has high efficiency of anti-inflammatory and anti-immune. And reported it had the effect of bone damage repairing on RA rats. But serious toxicity limits its development and application. In this study we prepared lipid cubic crystal nanoparticles gels of triptolide (TP-LCNPS-GEL) to treat rheumatoid arthritis by transdermal delivery. Temperature sensitivity and microstructure of TP-LCNPS-GEL was characterized. TP-LCNPS-GEL greatly improved percutaneous permeability, prolong biological half-life, average retention time and bioavailability of triptolide. The animal experiment results show that TP-LCNPS-GEL enhanced the effects on anti-inflammatory and bone protection of triptolide with low toxicity. All results indicated that TP-LCNPS-GEL as a transdermal delivery preparation is an excellent choice for rheumatoid arthritis.
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More than 40 years ago ethyl nitrosoеurea was identified as a powerful mutagen for mammalian germ cells resulting in random point mutations in gamete DNA. This feature allowed the use of this mutagen for genetic studies on the mechanisms of various pathological and physiological processes in model organisms. In our study genome-wide mutagenesis in C3H mice by ethyl nitrosourea followed in generation F3 by selection of animals resistant to acute lethal hepatotoxicity caused by a combination of E. coli lipopolysaccharide (LPS) and D-galactosamine (D-gal). Tumor necrosis factor (TNF) is known to be a critical mediator of this pathology. Exposure to D-galactosamine increases sensitivity of hepatocytes to TNF leading to their necrosis and/or apoptosis. After double LPS/D-gal screening in F3 several mice resistant to LPS/D-gal-induced hepatotoxicity were identified, and became the founders of the corresponding “mutant” families. Using outcrossing to C57BL6 background followed by intercrossing, generations F5 and F7 were obtained. Among families of mutant animals only one family showed the resistance to the combination of LPS and D-gal, but sensitivity to TNF-D-galactosamine. This phenotype showed approximately Mendelian inheritance consistent with the recessive mutation hypothesis. This latter fact was confirmed by the sensitivity of mice from “heterozygous generations” (F4 and F6) to lethal LPS/Dgal hepatotoxicity. Primary bone marrow macrophages obtained from half of the mutant mice showed significantly reduced levels of TNF after LPS stimulation in vitro. At the same time, the serum TNF levels 1 hour after the administration of a non-lethal LPS dose did not differ in the mutant family mice and wild-type mice. These results implicate a recessive mutation either in innate TLR4-mediated signaling pathway, including proteins associated with LPS transfer, adapter molecules, components of kinase signaling cascades, transcription factors, or in enzymes involved in regulation of TLR4 cascades, such as components of the ubiquitin cycle, or in genomic regulatory sequences that control the expression of one of these genes, including the tnf gene.
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It has become evident that the care of patients with COVID-19 does not end at the time of negative SARS-CoV-2 detection, as the number of patients with post-acute COVID-19 syndrome increases with an ever-increasing total infected patient population. This case report shows the possibility of alleviating the gastrointestinal symptoms of post-acute COVID-19 syndrome via microbiota-targeted nutritional intervention.
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The present introductory chapter presents the general overview of structure and functions of female reproductive system, reproductive organs (ovary, fallopian tubes, uterus), the processes of ovarian folliculogenesis, ovulation, luteogenesis and luteolysis, oocyte maturation, state of fallopian tubes, and uterus. The hormonal and intracellular regulators and factors controlling these functions and the most common reproductive disorders are outlined too.
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Background Sepsis often results in acute lung injury (ALI). Dexmedetomidine (Dex) was reported to protect cells and organs due to its direct cellular effects. This study aims to investigate the role of vagus nerves on Dex induced lung protection in lipopolysaccharide (LPS)-induced ALI rats. Methods The bilateral cervical vagus nerve of male Sprague-Dawley rats was sectioned or just exposed as sham surgery. After LPS administration, Dex antagonist yohimbine (YOH) and/or Dex was injected intraperitoneally to rats with or without vagotomy. The severity of ALI was determined with survival curve analysis and lung pathological scores. The plasma concentrations of interleukin 1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), catecholamine and acetylcholine were measured with enzyme-linked immunosorbent assay. Results The median survival time of LPS-induced ALI rats was prolonged by Dex (22 h, 95% CI, [24.46, 92.20]) vs. 14 h, 95% CI, [14.60, 89.57] of the LPS control group, P < 0.05), and the ALI score was reduced by Dex (6.5, 95% CI, [5.23, 8.10] vs. 11.5, 95% CI, [10.23, 13.10] in the LPS group, P < 0.01). However, these protective effects were significantly decreased by either YOH administration or vagotomy. Dex decreased LPS-induced IL-1β, TNF-α, and catecholamine but increased acetylcholine in blood serum; these effects of Dex was partially abolished by vagotomy. Conclusions Our data suggested that Dex increased vagal nerve tone that partially contributed to its anti-inflammatory and lung-protective effects. The indirect anti-inflammation and direct cytoprotection of Dex are likely through high vagal nerve tone and α2-adrenoceptor activation, respectively.
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Background: Sepsis-associated encephalopathy (SAE) is frequently encountered in sepsis and is often accompanied by neuroimaging findings indicating ischemia, hemorrhage, and edema. Posterior reversible encephalopathy syndrome (PRES) has been vastly underrecognized in previously reported cohorts of patients with sepsis and SAE. Our aim was to determine the prevalence and distinguishing clinical, neuroimaging, and electroencephalography features of PRES in SAE. Methods: In this prospective observational study, patients with radiologically identified PRES were selected from a consecutively enrolled cohort of 156 patients with SAE and assessed for neurological outcome using the extended Glasgow Outcome Scale for 12 months. Patients with SAE and PRES and other types of brain lesions were compared in terms of clinical and diagnostic workup features. Results: Fourteen of 156 patients (8.9%) were determined to be radiologically compatible with PRES, whereas 48 patients displayed other types of acute brain lesions. Patients with PRES often showed lesions in atypical regions, including frontal lobes, the corpus callosum, and the basal ganglia. Source of infection was mostly gram-negative bacteria originating from pneumonia or intraabdominal infections. Patients with PRES were not different from other patients with SAE with brain lesions in terms of features of sepsis and neurological outcome. However, patients with PRES showed increased prevalence of seizures and intraabdominal source of infection. Conclusions: PRES is highly prevalent in SAE, often encompasses unusual brain regions, and usually presents with generalized seizures. Patients with SAE and PRES do not appear to have distinguishing clinical and diagnostic workup features. However, generalized seizures may serve as warning signs for presence of PRES in patients with SAE.
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The inflammasomes are intracellular protein complexes that play an important role in innate immune sensing. Activation of inflammasomes leads to activation of caspase-1 and maturation and secretion of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18. In certain myeloid cells, this activation can also lead to an inflammatory cell death (pyroptosis). Inflammasome sensor proteins have evolved to detect a range of microbial ligands and bacterial exotoxins either through direct interaction or by detection of host cell changes elicited by these effectors. Bacterial exotoxins activate the inflammasomes through diverse processes, including direct sensor cleavage, modulation of ion fluxes through plasma membrane pore formation, and perturbation of various host cell functions. In this review, we summarize the findings on some of the bacterial exotoxins that activate the inflammasomes.
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Subtilase cytotoxin (SubAB) is the prototype of a recently emerged family of AB5 cytotoxins produced by Shiga-toxigenic Escherichia coli (STEC). Its mechanism of action involves highly specific A-subunit-mediated proteolytic cleavage of the essential endoplasmic reticulum (ER) chaperone BiP. Our previous in vivo studies showed that intraperitoneal injection of purified SubAB causes a major redistribution of leukocytes and elevated leukocyte apoptosis in mice, as well as profound splenic atrophy. In the current study, we investigated selected chemokine and proinflammatory cytokine responses to treatment with SubAB, a nontoxic derivative (SubAA272B), or Shiga toxin 2 (Stx2) in human macrophage (U937), brain microvascular endothelial (HBMEC), and colonic epithelial (HCT-8) cell lines, at the levels of secreted protein, cell-associated protein, and gene expression. Stx2 treatment upregulated expression of chemokines and cytokines at both the protein and mRNA levels. In contrast, SubAB induced significant decreases in secreted interleukin-8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) in all three tested cell lines and a significant decrease in secreted IL-6 in HBMECs. The downregulation of secreted chemokines or cytokines was not observed in SubAA272B-treated cells, indicating a requirement for BiP cleavage. The downregulation of secreted chemokines and cytokines by SubAB was not reflected at the mRNA and cell-associated protein levels, suggesting a SubAB-induced export defect.
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Administration of non-lethal doses of lipopolysaccharide (LPS) to experimental animals and humans results for a short period of time in a state of hyporesponsiveness to a second LPS challenge. This phenomenon, known as endotoxin tolerance, has been reproduced in vitro using human monocytes, rendered endotoxin-tolerant following a first incubation with LPS. A further activation by LPS was manifested by a dramatically reduced production of tumor necrosis factor α (TNFα). We report this low responsiveness of LPS pretreated monocytes as an endotoxin non-specific phenomenon. Indeed, TNFα release upon further activation with either killed Gram-positive bacteria (Staphylococci, Streptococci) or zymosan was also significantly diminished. This was not the case when phorbol myristate acetate (PMA) was used as a second triggering agent, suggesting that the monocyte hyporesponsiveness due to LPS does not affect all activation pathways, particularly that of protein kinase C. On the other hand, both PMA and zymosan pretreatment could reduce a further activation of monocytes by LPS. We investigated whether the first signal(s) delivered by LPS, could be mimicked by some of the LPS-induced cytokines. Preincubation of monocytes with either interleukin-1 (IL-1), IL-10 or transforming growth factor β (TGFβ) lower the TNFα production upon further activation with LPS. None of the cytokines alone was as efficient as the LPS molecule, but high levels of tolerization were obtained with combination of IL-1, IL-10 and TGFβ. Neither IL-6, IL-8 nor TNFα led to hyporeactive cells. Our data indicate that endotoxin tolerance is not an LPS-specific phenomenon and that more than one cytokine can contribute to render human monocytes hyporeactive to further activation by LPS.
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SUMMARY This review begins with a discussion of the large family of Staphylococcus aureus and beta-hemolytic streptococcal pyrogenic toxin T lymphocyte superantigens from structural and immunobiological perspectives. With this as background, the review then discusses the major known and possible human disease associations with superantigens, including associations with toxic shock syndromes, atopic dermatitis, pneumonia, infective endocarditis, and autoimmune sequelae to streptococcal illnesses. Finally, the review addresses current and possible novel strategies to prevent superantigen production and passive and active immunization strategies.
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Caspase-1 activation by inflammasome scaffolds comprised of intracellular nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) and the adaptor ASC is believed to be essential for production of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18 during the innate immune response. Here we show, with C57BL/6 Casp11 gene-targeted mice, that caspase-11 (also known as caspase-4) is critical for caspase-1 activation and IL-1β production in macrophages infected with Escherichia coli, Citrobacter rodentium or Vibrio cholerae. Strain 129 mice, like Casp11(-/-) mice, exhibited defects in IL-1β production and harboured a mutation in the Casp11 locus that attenuated caspase-11 expression. This finding is important because published targeting of the Casp1 gene was done using strain 129 embryonic stem cells. Casp1 and Casp11 are too close in the genome to be segregated by recombination; consequently, the published Casp1(-/-) mice lack both caspase-11 and caspase-1. Interestingly, Casp11(-/-) macrophages secreted IL-1β normally in response to ATP and monosodium urate, indicating that caspase-11 is engaged by a non-canonical inflammasome. Casp1(-/-)Casp11(129mt/129mt) macrophages expressing caspase-11 from a C57BL/6 bacterial artificial chromosome transgene failed to secrete IL-1β regardless of stimulus, confirming an essential role for caspase-1 in IL-1β production. Caspase-11 rather than caspase-1, however, was required for non-canonical inflammasome-triggered macrophage cell death, indicating that caspase-11 orchestrates both caspase-1-dependent and -independent outputs. Caspase-1 activation by non-canonical stimuli required NLRP3 and ASC, but caspase-11 processing and cell death did not, implying that there is a distinct activator of caspase-11. Lastly, loss of caspase-11 rather than caspase-1 protected mice from a lethal dose of lipopolysaccharide. These data highlight a unique pro-inflammatory role for caspase-11 in the innate immune response to clinically significant bacterial infections.
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Bacterial superantigens are Gram-positive exotoxins that induce proinflammatory cytokine release in vitro, cause lethal shock in vivo, and can be detected in the bloodstream of critically ill patients. They also have a powerful priming effect on the TLR4 agonist LPS. The aim of this study was to investigate the relationship between superantigens and the TLR2 agonist bacterial lipoprotein (BLP). Priming of human monocytes or PBMCs with superantigens significantly enhanced proinflammatory cytokine TNF-α and IL-6 release in response to BLP stimulation. The priming effect of superantigens could be blocked by inhibiting p38 MAPK during the priming phase as opposed to NF-κB or ERK inhibition. This was consistent with higher expression of the phosphorylated p38 after superantigen priming and BLP or LPS stimulation. C57BL/6 mice with superantigen priming (10 μg/mouse) when challenged with BLP (600 μg/mouse) exhibited substantially higher mortality (100%) compared with mice without superantigen priming (zero). Mice given superantigen alone did not demonstrate any signs of illness. Mice challenged with both superantigen and BLP had significantly higher levels of serum TNF-α and IL-6 compared with those of mice challenged with either agent alone. Depletion of the monocyte/macrophage subpopulation significantly reduced the mortality rate from 100 to 20% in superantigen-primed, BLP-challenged C57BL/6 mice, with a 5- to 10-fold decrease in serum TNF-α and IL-6. Our results demonstrate that bacterial superantigens enhance the in vitro proinflammatory cytokine release and in vivo lethality of BLP. This novel finding may help to explain the massive proinflammatory cytokine release seen in superantigen-mediated septic shock.
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Staphylococcal enterotoxin (SE) -induced toxic shock is triggered by inflammatory cytokine signal amplification after SE binding to major histocompatibility complex class II molecules on antigen-presenting cells and T-cell receptors. Identifying host cellular elements contributing to this pro-inflammatory signal amplification is critical for developing a strategy for therapeutic intervention. Myeloid differentiation primary-response protein 88 (MyD88) is an intracellular signalling adaptor protein primarily known for mediating pro-inflammatory cytokine responses. We investigated the role of MyD88 in staphylococcal enterotoxin A (SEA) -treated cell cultures and mouse models of toxic shock. Our results demonstrated that elevated levels of tumour necrosis factor-alpha, interferon-gamma, interleukin-1alpha/beta (IL-1alpha/beta), IL-2 and IL-6 production correlated with up-regulation of MyD88 after treatment of spleen cells and mice with SEA alone or in combination with lipopolysaccharide (LPS). The SEA-induced lethality was also observed in (LPS-independent) D-galactosamine-sensitized mice. While LPS potentiated SEA-induced cytokine responses, D-galactosamine treatment had no additive effect. Most importantly, our results demonstrated that MyD88(-/-) mice were resistant to SEA-induced toxic shock and had reduced pro-inflammatory cytokine responses. These results suggest that SEA-induced lethality is primarily dependent on MyD88. Our findings offer an important insight on potential therapeutic treatment of SEA-induced toxic shock targeting MyD88.
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Bacteremia is recognized as a critical condition that influences the outcome of sepsis. Although large-scale surveillance studies of bacterial species causing bacteremia have been published, the pathophysiological differences in bacteremias with different causative bacterial species remain unclear. The objective of the present study is to investigate the differences in pathophysiology and the clinical course of bacteremia caused by different bacterial species. We reviewed the medical records of all consecutive patients admitted to the general intensive care unit (ICU) of a university teaching hospital during the eight-year period since introduction of a rapid assay for interleukin (IL)-6 blood level to routine ICU practice in May 2000. White blood cell count, C-reactive protein (CRP), IL-6 blood level, and clinical course were compared among different pathogenic bacterial species. The 259 eligible patients, as well as 515 eligible culture-positive blood samples collected from them, were included in this study. CRP, IL-6 blood level, and mortality were significantly higher in the septic shock group (n = 57) than in the sepsis group (n = 127) (P < 0.001). The 515 eligible culture-positive blood samples harbored a total of 593 isolates of microorganisms (Gram-positive, 407; Gram-negative, 176; fungi, 10). The incidence of Gram-negative bacteremia was significantly higher in the septic shock group than in the sepsis group (P < 0.001) and in the severe sepsis group (n = 75, P < 0.01). CRP and IL-6 blood level were significantly higher in Gram-negative bacteremia (n = 176) than in Gram-positive bacteremia (n = 407) (P < 0.001, <0.0005, respectively). The incidence of Gram-negative bacteremia was significantly higher in bacteremic ICU patients with septic shock than in those with sepsis or severe sepsis. Furthermore, CRP and IL-6 levels were significantly higher in Gram-negative bacteremia than in Gram-positive bacteremia. These findings suggest that differences in host responses and virulence mechanisms of different pathogenic microorganisms should be considered in treatment of bacteremic patients, and that new countermeasures beyond conventional antimicrobial medications are urgently needed.
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Vertebrates vary in resistance and resilience to infectious diseases, and the mechanisms that regulate the tradeoff between these often opposing protective processes are not well understood. Variability in the sensitivity of species to the induction of damaging inflammation in response to equivalent pathogen loads (resilience) complicates the use of animal models that reflect human disease. We found that induction of proinflammatory cytokines from macrophages in response to inflammatory stimuli in vitro is regulated by proteins in the sera of species in inverse proportion to their in vivo resilience to lethal doses of bacterial lipopolysaccharide over a range of 10,000-fold. This finding suggests that proteins in serum rather than intrinsic cellular differences may play a role in regulating variations in resilience to microbe-associated molecular patterns between species. The involvement of circulating proteins as key molecules raises hope that the process might be manipulated to create better animal models and potentially new drug targets.
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TNF, IL-1, and IL-6 are integral components of the cytokine cascade released in the response to inflammatory stimuli such as LPS. IL-8 is produced both in response to LPS as well as TNF and IL-1. The early, local production of TNF and IL-1 may therefore contribute to the subsequent expression of IL-8. This hypothesis was tested using LPS-stimulated human whole blood as an ex vivo model of local cytokine production. The production of TNF, IL-1 alpha, IL-1 beta, IL-6, and IL-8 was found to be responsive to a wide range of LPS concentrations (0.1 ng/ml-10 micrograms/ml). These cytokines were first detected between 1 to 4 h post-LPS stimulation, and reached plateau levels after 6 to 12 h. IL-8, however, also displayed a secondary wave of production, with the levels again increasing between 12 to 24 h. The IL-8 present in the plasma after LPS stimulation was biologically active, as assessed by neutrophil chemotaxis. In further studies, addition of anti-TNF and anti-IL-1 neutralizing antibodies, alone and in combination, to LPS-stimulated blood resulted in nearly complete ablation of the secondary phase of IL-8 synthesis at both the levels of protein and mRNA, while leaving the first, LPS-mediated phase of IL-8 synthesis unaffected. This model of cytokine production in human whole blood may reflect the sequence of events in a localized environment of inflammation where both a primary stimulus and the induced early cytokine mediators may serve to elicit multiple, temporally distinct phases of IL-8 production.
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Because mice are more resistant than humans to the pathogenic effects of bacterial toxins, we used D-Galactosamine- (D-Gal) sensitized mice as a model system to evaluate potential toxic shock symptoms triggered by the superantigen staphylococcal enterotoxin B (SEB). We show that similar to endotoxin (lipopolysaccharide) [LPS], the exotoxin SEB causes lethal shock within 8 h in D-Gal-sensitized mice, inducing 100% and about 50% lethality with 20 and 2 micrograms SEB, respectively. The lethal shock triggered by the superantigen SEB is mediated by T cells, a conclusion based on the observation that T cell repopulation of SCID mice conferred sensitivity to SEB. Since CSA also conferred protection, the role of T cell-derived lymphokines in mediating lethal shock was evaluated. Within 30-60 min after SEB injection, serum tumor necrosis factor (TNF) levels peaked, followed immediately by interleukin-2 (IL-2). Serum-borne lymphokines were detected well in advance of signs of T cell activation, as assessed by IL-2 receptor expression of SEB-reactive V beta 8+ T cells. Passive immunization with anti-TNF-alpha/beta-neutralizing monoclonal antibody also conferred protection, indicating that it is TNF which is critical for initiating toxic shock symptoms. Taken together, this study defines basic differences between endotoxin (LPS)- and exotoxin (SEB)-mediated lethal shock, in that the former is mediated by macrophages and the latter by T cells. Yet the pathogenesis distal to the lymphokine/cytokine-producing cells appears surprisingly similar in that TNF represents a key mediator in inducing shock.
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We investigated the ability of staphylococcal enterotoxins A and B, exfoliative toxins A and B, and toxic shock syndrome toxin 1 to activate macrophages. All of the toxins tested had the potential to stimulate tumoricidal activity in peritoneal macrophages from lipopolysaccharide-responsive C3HeB/FeJ mice. In contrast, none of the toxins activated cytotoxicity in lipopolysaccharide-unresponsive macrophages from C3H/HeJ mice. We also studied toxin stimulation of monokine secretion. Staphylococcal enterotoxin A, toxic shock syndrome toxin 1, and both exfoliative toxins triggered C3HeB/FeJ macrophages to secrete tumor necrosis factor alpha, but enterotoxin B induced only marginal amounts of tumor necrosis factor. All of the toxins used stimulated interleukin-6 production by macrophages from both strains of mice. Nitric oxide is produced in response to the exfoliative toxins only by the lipopolysaccharide-responsive macrophages. These results suggest that macrophages respond differently to several staphylococcal exotoxins.
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The production by monocytes of interleukin-1 alpha (IL-1 alpha), interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF alpha) in intensive care unit (ICU) patients with sepsis syndrome (n = 23) or noninfectious shock (n = 6) is reported. Plasma cytokines, cell-associated cytokines within freshly isolated monocytes and LPS-induced in vitro cytokine production were assessed at admission and at regular intervals during ICU stay. TNF alpha and IL-6 were the most frequently detected circulating cytokines. Despite the fact that IL-1 alpha is the main cytokine found within monocytes upon in vitro activation of cells from healthy individuals, it was very rarely detected within freshly isolated monocytes from septic patients, and levels of cell-associated IL-1 beta were lower than those of TNF alpha. Cell-associated IL-1 beta and TNF alpha were not correlated with corresponding levels in plasma. Upon LPS stimulation, we observed a profound decrease of in vitro IL-1 alpha production by monocytes in all patients, and of IL-1 beta, IL-6, and TNF alpha in septic patients. This reduced LPS-induced production of cytokines was most pronounced in patients with gram-negative infections. Finally, monocytes from survival patients, but not from nonsurvival ones recovered their capacity to produce normal amounts of cytokines upon LPS stimulation. In conclusion, our data indicate an in vivo activation of circulating monocytes during sepsis as well as in noninfectious shock and suggest that complex regulatory mechanisms can downregulate the production of cytokines by monocytes during severe infections.
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Sepsis and trauma are both leading causes of death in the United States and represent major public health challenges. Murine models have largely been used in sepsis and trauma research to better understand the pathophysiological changes that occur after an insult and to develop potential life-saving therapeutic agents. Mice are favorable subjects for this type of research given the variety of readily available strains including inbred, outbred, and transgenic strains. In addition, they are relatively easy to maintain and have a high fecundity. However, pharmacological therapies demonstrating promise in preclinical mouse models of sepsis and trauma often fail to demonstrate similar efficacy in human clinical trials, prompting considerable criticism surrounding the capacity of murine models to recapitulate complex human diseases like sepsis and traumatic injury. Fundamental differences between the two species include, but are not limited to, the divergence of the transcriptomic response, the mismatch of temporal response patterns, differences in both innate and adaptive immunity, and heterogeneity within the human population in comparison to the homogeneity of highly inbred mouse strains. Given the ongoing controversy, this narrative review aims to not only highlight the historical importance of the mouse as an animal research model but also highlight the current benefits and limitations of the model as it pertains to sepsis and trauma. Lastly, this review will propose future directions that may promote further use of the model.
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In the article "The staphylococcal enterotoxins and their relatives" by Philippa Marrack and John Kappler (11 May, p. 705), figures 1, 2, and 3 were incorrectly printed. The color portions of figures 2 and 3 should have been included in figure 1. The correct figure 1 is printed below. See image in PDF file
Article
Toxic shock syndrome (TSS) is caused by staphylococcal and streptococcal superantigens (SAgs) that provoke a swift hyperinflammatory response typified by a cytokine storm. The precipitous decline in the host's clinical status and the lack of targeted therapies for TSS emphasize the need to identify key players of the storm's initial wave. Using a humanized mouse model of TSS and human cells, we herein demonstrate that SAgs elicit in vitro and in vivo IL-17A responses within hours. SAg-triggered human IL-17A production was characterized by remarkably high mRNA stability for this cytokine. A distinct subpopulation of CD4+ effector memory T (TEM) cells that secrete IL-17A, but not IFN-γ, was responsible for early IL-17A production. We found mouse "TEM-17" cells to be enriched within the intestinal epithelium and among lamina propria lymphocytes. Furthermore, interfering with IL-17A receptor signaling in human PBMCs attenuated the expression of numerous inflammatory mediators implicated in the TSS-associated cytokine storm. IL-17A receptor blockade also abrogated the secondary effect of SAg-stimulated PBMCs on human dermal fibroblasts as judged by C/EBP δ expression. Finally, the early IL-17A response to SAgs was pathogenic because in vivo neutralization of IL-17A in humanized mice ameliorated hepatic and intestinal damage and reduced mortality. Together, our findings identify CD4+ TEM cells as a key effector of TSS and reveal a novel role for IL-17A in TSS immunopathogenesis. Our work thus elucidates a pathogenic, as opposed to protective, role for IL-17A during Gram-positive bacterial infections. Accordingly, the IL-17-IL-17R axis may provide an attractive target for the management of SAg-mediated illnesses.
Article
Pathogenic Yersinia, including Y. pestis, the agent of plague in humans, and Y. pseudotuberculosis, the related enteric pathogen, deliver virulence effectors into host cells via a prototypical type III secretion system to promote pathogenesis. These effectors, termed Yersinia outer proteins (Yops), modulate multiple host signaling responses. Studies in Y. pestis and Y. pseudotuberculosis have shown that YopM suppresses infection-induced inflammasome activation; however, the underlying molecular mechanism is largely unknown. Here we show that YopM specifically restricts the pyrin inflammasome, which is triggered by the RhoA-inactivating enzymatic activities of YopE and YopT, in Y. pseudotuberculosis-infected macrophages. The attenuation of a yopM mutant is fully reversed in pyrin knockout mice, demonstrating that YopM inhibits pyrin to promote virulence. Mechanistically, YopM recruits and activates the host kinases PRK1 and PRK2 to negatively regulate pyrin by phosphorylation. These results show how a virulence factor can hijack host kinases to inhibit effector-triggered pyrin inflammasome activation.
Article
Interleukin-1β (IL-1β) is a key proinflammatory cytokine that drives antimicrobial immune responses. IL-1β is aberrantly activated in autoimmune diseases, and IL-1β inhibitors are used as therapeutic agents to treat patients with certain autoimmune disorders. Review of postmarketing surveillance of patients receiving IL-1β inhibitors found a disproportionate reporting of invasive infections by group A Streptococcus (GAS). IL-1β inhibition increased mouse susceptibility to GAS infection, but IL-1β was produced independent of canonical inflammasomes. Newly synthesized IL-1β has an amino-terminal prodomain that blocks signaling activity, which is usually proteolytically removed by caspase-1, a protease activated within the inflammasome structure. In place of host caspases, the secreted GAS cysteine protease SpeB generated mature IL-1β. During invasive infection, GAS isolates may acquire pathoadaptive mutations eliminating SpeB expression to evade detection by IL-1β. Pharmacological IL-1β inhibition alleviates this selective pressure, allowing invasive infection by nonpathoadapted GAS. Thus, IL-1β is a sensor that directly detects pathogen-associated proteolysis through an independent pathway operating in parallel with host inflammasomes. Because IL-1β function is maintained across species, yet cleavage by caspases does not appear to be, detection of microbial proteases may represent an ancestral system of innate immune regulation.
Chapter
Endotoxins isolated from Gram negative bacteria can induce profound physiological modifications and trigger the immune system. Endotoxins are lipopolysaccharides (LPS) which consist of a polysaccharide chain (PS) covalently linked to a hydrophobic moiety (Lipid A) via a 2-keto-3-deoxy-octonic acid residue(s) (KDO). The classical endotoxic activities (e.g. local Shwartzman reaction, pyrogenicity, lethality…) as well as the immunological effects of endotoxins (adjuvanticity, mitogenicity, polyclonal activation of immunoglobulin secreting cells…) are mediated by the Lipid A (1,2). However, it has also been shown that the polysaccharide moiety of some LPS act as an adjuvant (3,4); protect mice against lethal irradiation (5,6); induce the production of colony stimulating factors (5–8); induce a “transfer tumor resistance” in serum of BCG-infected mice (9); and mediate B-cell proliferation (10). LPS can also stimulate complex positive and negative immunoregulatory circuits in which monocytes/macrophages play a central role (10–14).
Article
Burkholderia cenocepacia is an opportunistic pathogen of the cystic fibrosis lung that elicits a strong inflammatory response. B. cenocepacia employs a type VI secretion system (T6SS) to survive in macrophages by disarming Rho-type GTPases, causing actin cytoskeletal defects. Here, we identified TecA, a non-VgrG T6SS effector responsible for actin disruption. TecA and other bacterial homologs bear a cysteine protease-like catalytic triad, which inactivates Rho GTPases by deamidating a conserved asparagine in the GTPase switch-I region. RhoA deamidation induces caspase-1 inflammasome activation, which is mediated by the familial Mediterranean fever disease protein Pyrin. In mouse infection, the deamidase activity of TecA is necessary and sufficient for B. cenocepacia-triggered lung inflammation and also protects mice from lethal B. cenocepacia infection. Therefore, Burkholderia TecA is a T6SS effector that modifies a eukaryotic target through an asparagine deamidase activity, which in turn elicits host cell death and inflammation through activation of the Pyrin inflammasome. Burkholderia cenocepacia is highly proinflammatory and causes severe lung infection in cystic fibrosis patients. Aubert et al. identify a type VI effector, TecA, in B. cenocepacia that disrupts macrophage actin cytoskeleton by deamidating Rho GTPases. The action of TecA is detected by the Pyrin inflammasome, which triggers inflammation in mice.
Article
Three years ago, at a meeting of this body in Baltimore, a paper was read upon a similar subject by a distinguished surgeon. In that paper, based upon a personal experience of nine cases, three of which were carcinoma and a large proportion of the remainder types of sarcoma, which I had myself regarded as unfavorable, the broad generalization was made that the method was of no value, and the prophecy was ventured that in one year from that date it would be forgotten and buried in that oblivion that has furnished a quiet resting-place for so many of the brilliant cancer cures that have from time to time dotted the medical history of the past.
Article
Oligonucleotide and complementary DNA microarrays are being used to subclassify histologically similar tumours, monitor disease progress, and individualize treatment regimens. However, extracting new biological insight from high-throughput genomic studies of human diseases is a challenge, limited by difficulties in recognizing and evaluating relevant biological processes from huge quantities of experimental data. Here we present a structured network knowledge-base approach to analyse genome-wide transcriptional responses in the context of known functional interrelationships among proteins, small molecules and phenotypes. This approach was used to analyse changes in blood leukocyte gene expression patterns in human subjects receiving an inflammatory stimulus (bacterial endotoxin). We explore the known genome-wide interaction network to identify significant functional modules perturbed in response to this stimulus. Our analysis reveals that the human blood leukocyte response to acute systemic inflammation includes the transient dysregulation of leukocyte bioenergetics and modulation of translational machinery. These findings provide insight into the regulation of global leukocyte activities as they relate to innate immune system tolerance and increased susceptibility to infection in humans.
Article
We had previously shown that the specific binding of endotoxin (lipopolysaccharide, LPS) to human monocytes in the presence of serum was mediated by the polysaccharide moiety of the LPS molecule. CD14 has been identified as the main receptor for endotoxin on monocytes/macrophages. In the present report we demonstrate that the CD14 molecule exhibits lectin-like properties. Anti-CD14 monoclonal antibodies inhibited the binding of [3H]-radiolabeled Neisseria meningitidis LPS as efficiently as the homologous unlabeled LPS. Rough Escherichia coli LPS (Rc- and Re-types) could also inhibit the binding of [3H]-LPS to a similar extent, whereas lipid A had no or very weak inhibitory activity. This suggests a major contribution of the inner-core region within the LPS and particularly the Kdo sugars. The lectin-like nature of CD14 was assessed with polyanionic sugars as well as with uncharged polysaccharides. The relative efficiencies in competition were dextran sulfate > fucoidan > mannan > polygalacturonic acid = heparan sulfate ≥ heparin ≥ chondroitin sulfate. Candida albicans phospholipomannan was far more active in the competition experiment than the mannan, indicating that, besides the osidic residues, anionic charges and/or fatty acids may contribute to the interaction with the CD14 molecule. Binding of polysaccharide to CD14 was not sufficient to trigger TNFα and IL-6 production since phospholipomannan and dextran sulfate were unable to induce cytokine release. Taken together, these results demonstrate that the binding of [3H]-LPS to CD14 involves the contribution of sugars and suggest that the signals for cytokine production require additional interactions.
Article
We report on the genetic effect onin vivoproduction of tumor necrosis factor (TNF)-α induced by lipopolysaccharides (LPS) using various congenic mouse strains. B10.A, B10.A(3R), B10.AQR, B10.A(5R), and B10.S(7R) produced significantly high TNF-α compared with B10.BR, B10.S, C57BL/10, B10.A(2R), B10.A(4R), B10.G, B10.DA(80NS), and B10.RIII(71NS). This suggests that LPS-induced TNF-α production is genetically controlled by H-2. Mice with the same alleles on K, A, E, or S loci produced various (high or low) levels of TNF-α, thus indicating that regulatory genes are located outside these loci. All strains with H-2Ddproduced significantly high levels of TNF-α, but strains with other alleles in the H-2D locus produced low levels. Thus, TNF-α production appears to be genetically linked to H-2D itself or H-2D linked genes and the allele d is linked to a high responder gene. This was the case with the A background. C3H/HeN (H-2k), however, showed a high TNF-α production, suggesting the presence of anoth