interleukin-8 and Dysentery--Bacillary

A rapid rise in resistance to conventional antibiotics for Shigella spp. has created a problem in treating shigellosis. Hence, there is an urgent need for new and non-conventional anti-bacterial agents. The aim of this study is to show how Asiatic acid, a plant-derived compound, inhibits the intracellular growth of Shigella flexneri.. Shigella flexneri sensitive and resistant strains were used for checking antimicrobial activity of Asiatic acid by gentamicin protection assay. Asiatic acid inhibited the intracellular growth of all strains. Gene expression analysis showed antimicrobial peptide (AMP) up-regulation by Asiatic acid in intestinal cells. Further western blot analysis showed that ERK, p38, and JNK are activated by Asiatic acid. ELISA was performed to check IL-8, IL-6, and cathelicidin secretion. The antibacterial effect of Asiatic acid was further verified in an in vivo mouse model.. The reason behind the antibacterial activities of Asiatic acid is probably over-expression of antimicrobial peptide genes. Besides, direct antimicrobial activities, antimicrobial peptides also carry immunomodulatory activities. Here, Asiatic acid increased IL-6 and IL-8 secretion to induce inflammation. Overall, Asiatic acid up-regulates antimicrobial peptide gene expression and inhibits intracellular S. flexneri growth. Moreover, Asiatic acid reduced bacterial growth and recovered intestinal tissue damages in in vivo mice model.

Topics: Animals; Anti-Bacterial Agents; Antimicrobial Peptides; Dysentery, Bacillary; Gene Expression; Interleukin-6; Interleukin-8; Mice; Microbial Sensitivity Tests; Shigella; Shigella flexneri

Recently, we have demonstrated, immunization of adult female mice with outer membrane vesicles (OMVs) of Shigella boydii type 4 protected their offspring passively from shigellosis. In our present study, we have advanced our research by formulating multi-serotype outer membrane vesicles (MOMVs), mixing the OMVs of Shigella dysenteriae 1 Δstx, Shigella flexneri 2a, 3a and 6, S. boydii type 4 and Shigella sonnei to achieve a broad spectrum protection against shigellosis. Adult mice were immunized orally with 50 μg of MOMVs, four times at weekly intervals. Immunological parameters were observed at various time points, before, during and after immunization, in adult mice. Passive protection was examined in their offspring by measuring protective efficacy and studying intestinal colonization, after challenging with various Shigella strains. Immunized dams exhibited a consistent broad spectrum antibody response. 3-4 day-old offspring of immunized dams showed significant long term passive protection against wild type S. flexneri 2a, 3a, and 6, S. boydii type 2 and S. dysenteriae 1. Their stomach extracts, essentially containing mother's milk, have also exhibited significant levels of anti-MOMVs immunoglobulins. In conclusion, MOMVs formulation represents an easy, safe immunization strategy that was found suitable to provide complete passive protection to the neonatal mice against all four serogroups of Shigellae. It could be exploited for the development of a novel non-living vaccine against human shigellosis in near future.

Topics: Animals; Animals, Newborn; Antibodies, Bacterial; Antibody Specificity; Bacterial Outer Membrane Proteins; Cell Membrane; Dysentery, Bacillary; Female; Gastrointestinal Contents; Immunity, Cellular; Immunity, Maternally-Acquired; Immunoglobulins; Interleukin-8; Macrophages; Mice; Phagocytosis; Shigella dysenteriae; Shigella flexneri; Shigella sonnei; Shigella Vaccines

Intestinal pathogens use the host's excessive inflammatory cytokine response, designed to eliminate dangerous bacteria, to disrupt epithelial gut wall integrity and promote their tissue invasion. We sought to develop a non-antibiotic-based approach to prevent this injury. Molecular docking studies suggested that glycosylated dendrimers block the TLR4-MD-2-LPS complex, and a 13.6 kDa polyamidoamine (PAMAM) dendrimer glucosamine (DG) reduced the induction of human monocyte interleukin (IL)-6 by Gram-negative bacteria. In a rabbit model of shigellosis, PAMAM-DG prevented epithelial gut wall damage and intestinal villous destruction, reduced local IL-6 and IL-8 expression, and minimized bacterial invasion. Computational modelling studies identified a 3.3 kDa polypropyletherimine (PETIM)-DG as the smallest likely bioactive molecule. In human monocytes, high purity PETIM-DG potently inhibited Shigella Lipid A-induced IL-6 expression. In rabbits, PETIM-DG prevented Shigella-induced epithelial gut wall damage, reduced local IL-6 and IL-8 expression, and minimized bacterial invasion. There was no change in β-defensin, IL-10, interferon-β, transforming growth factor-β, CD3 or FoxP3 expression. Small and orally delivered DG could be useful for preventing gut wall tissue damage in a wide spectrum of infectious diarrhoeal diseases.

Topics: Administration, Oral; Animals; Bacterial Translocation; Dendrimers; Diarrhea; Disease Models, Animal; Dysentery, Bacillary; Gastrointestinal Agents; Gastrointestinal Tract; Glucosamine; Immunologic Factors; Interleukin-6; Interleukin-8; Rabbits; Shigella

Topics: Animals; Dendrimers; Dysentery, Bacillary; Gastrointestinal Agents; Gastrointestinal Tract; Interleukin-6; Interleukin-8

Rab GTPases are frequent targets of vacuole-living bacterial pathogens for appropriate trafficking of the vacuole. Here we discover that bacterial effectors including VirA from nonvacuole Shigella flexneri and EspG from extracellular Enteropathogenic Escherichia coli (EPEC) harbor TBC-like dual-finger motifs and exhibits potent RabGAP activities. Specific inactivation of Rab1 by VirA/EspG disrupts ER-to-Golgi trafficking. S. flexneri intracellular persistence requires VirA TBC-like GAP activity that mediates bacterial escape from autophagy-mediated host defense. Rab1 inactivation by EspG severely blocks host secretory pathway, resulting in inhibited interleukin-8 secretion from infected cells. Crystal structures of VirA/EspG-Rab1-GDP-aluminum fluoride complexes highlight TBC-like catalytic role for the arginine and glutamine finger residues and reveal a 3D architecture distinct from that of the TBC domain. Structure of Arf6-EspG-Rab1 ternary complex illustrates a pathogenic signaling complex that rewires host Arf signaling to Rab1 inactivation. Structural distinctions of VirA/EspG further predict a possible extensive presence of TBC-like RabGAP effectors in counteracting various host defenses.

Topics: ADP-Ribosylation Factors; Amino Acid Sequence; Animals; Autophagy; Dysentery, Bacillary; Enteropathogenic Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Fibroblasts; GTPase-Activating Proteins; Interleukin-8; Mice; Models, Molecular; Molecular Sequence Data; Protein Structure, Tertiary; Sequence Alignment; Shigella flexneri; Virulence; Virulence Factors

This study focused on identifying possible new options derived from natural sources for the treatment of bacterial infections. Several natural products were investigated for their potential in modulating Shigella-host-cell interactions. The proliferation of Shigella sonnei was effectively inhibited inside HEp-2 cells in the presence of 4-methoxycinnamic acid and propolin D. Propolin D also significantly reduced the apoptosis of infected macrophage-like U937 cells and moderately reduced the secretion of interleukin (IL)-1β and IL-18, which probably resulted from the inhibition of invasion plasmid antigen B secretion by this compound. Further characterization showed that propolin D did not prevent escape of Shigella from phagocytic vacuoles, as evidenced by actin-based motility and by the fact that addition of chloroquine did not further reduce the number of intracellular c.f.u. The role of propolin D in modulating autophagy could not be established under the experimental conditions used. As these compounds had no direct anti-Shigella activity in vitro, it was concluded that these compounds modulated Shigella-host-cell interactions by targeting yet-to-be defined mechanisms that provide benefits to host cells.

Topics: Apoptosis; Cell Survival; Cinnamates; Dysentery, Bacillary; Flavonoids; Hep G2 Cells; Humans; Interleukin-1beta; Interleukin-8; Microbial Sensitivity Tests; Shigella sonnei; U937 Cells

Shigella dysenteriae Type 1 dysentery is a major cause of morbidity and mortality in children from less developed and developing countries. The present study explores the hypothesis that lactobacilli protect the host cell during S. dysenteriae Type 1 infection and its mechanism of action.. Caco-2 cells incubated for 1h with Lactobacillus rhamnosus or Lactobacillus acidophilus at the multiplicity of infection of 100, either alone or in combination followed by addition of Shigella at the same multiplicity of infection for 5h served as treatment groups. Cells incubated with Shigella without lactobacilli addition served as infected cells. At the end of experimental period, cells were processed suitably to enumerate adherent and internalized Shigella. Reverse transcription-polymerase chain reaction was performed to assess mRNA expression of interleukin-8 and tumour necrosis factor-alpha. Immunoblot for heat shock protein-70 and cytotoxicity assay were performed.. Pretreatment with the combination of lactobacilli significantly (p<0.05) prevented adherence and internalization of Shigella coupled with reduced expression of tumour necrosis factor-alpha and interleukin-8 in host cells.. L. rhamnosus and L. acidophilus, synergistically offered better protection during S. dysenteriae Type 1 infection by efficiently inhibiting adherence and internalization of Shigella coupled with inhibition of pro-inflammatory response.

Topics: Bacterial Adhesion; Caco-2 Cells; Coculture Techniques; Dysentery, Bacillary; Host-Pathogen Interactions; HSP70 Heat-Shock Proteins; Humans; Immunity, Innate; Interleukin-8; Lacticaseibacillus rhamnosus; Lactobacillus acidophilus; Shigella dysenteriae; Tumor Necrosis Factor-alpha

The lack of a standardized laboratory animal model that mimics key aspects of human shigellosis remains a major obstacle to addressing questions about pathogenesis, screening therapeutics, and evaluation of vaccines.. We characterized a piglet model for Shigella dysenteriae type 1.. Piglets developed acute diarrhea, anorexia, and dehydration, which could often be fatal, with symptom severity depending on age and dose. Bacteria were apparent in the lumen and on the surface epithelium throughout the gut initially, but severe mucosal damage and bacterial cellular invasion were most profound in the colon. Detached necrotic colonocytes were present in the lumen, with inflammatory cells outpouring from damaged mucosa. High levels of interleukin (IL)-8 and IL-12 were followed by high levels of other proinflammatory cytokines. Elevated levels of tumor necrosis factor-alpha, IL-1beta, IL-6, and IL-10 were detected in feces and in gut segments from infected animals. Bacteria were present inside epithelial cells and within colonic lamina propria. In contrast, an isogenic strain lacking Shiga toxin induced similar but milder symptoms, with moderate mucosal damage and lower cytokine levels.. We conclude that piglets are highly susceptible to shigellosis, providing a useful tool with which to compare vaccine candidates for immunogenicity, reactogenicity, and response to challenge; investigate the role of virulence factors; and test the efficacy of microbial agents.

Topics: Animals; Case-Control Studies; Colony Count, Microbial; Cytokines; Disease Models, Animal; Dysentery, Bacillary; Euthanasia, Animal; Feces; Gastroenteritis; Gastrointestinal Tract; Interleukin-12; Interleukin-8; Microscopy, Electron; Shigella dysenteriae; Swine

Many bacterial pathogens utilize a type III secretion system to deliver multiple effector proteins into host cells. Here we found that the type III effectors, NleE from enteropathogenic E. coli (EPEC) and OspZ from Shigella, blocked translocation of the p65 subunit of the transcription factor, NF-kappaB, to the host cell nucleus. NF-kappaB inhibition by NleE was associated with decreased IL-8 expression in EPEC-infected intestinal epithelial cells. Ectopically expressed NleE also blocked nuclear translocation of p65 and c-Rel, but not p50 or STAT1/2. NleE homologues from other attaching and effacing pathogens as well OspZ from Shigella flexneri 6 and Shigella boydii, also inhibited NF-kappaB activation and p65 nuclear import; however, a truncated form of OspZ from S. flexneri 2a that carries a 36 amino acid deletion at the C-terminus had no inhibitory activity. We determined that the C-termini of NleE and full length OspZ were functionally interchangeable and identified a six amino acid motif, IDSY(M/I)K, that was important for both NleE- and OspZ-mediated inhibition of NF-kappaB activity. We also established that NleB, encoded directly upstream from NleE, suppressed NF-kappaB activation. Whereas NleE inhibited both TNFalpha and IL-1beta stimulated p65 nuclear translocation and IkappaB degradation, NleB inhibited the TNFalpha pathway only. Neither NleE nor NleB inhibited AP-1 activation, suggesting that the modulatory activity of the effectors was specific for NF-kappaB signaling. Overall our data show that EPEC and Shigella have evolved similar T3SS-dependent means to manipulate host inflammatory pathways by interfering with the activation of selected host transcriptional regulators.

Topics: Active Transport, Cell Nucleus; Caco-2 Cells; Dysentery, Bacillary; Escherichia coli Infections; Escherichia coli O157; Escherichia coli Proteins; HeLa Cells; Humans; I-kappa B Proteins; Interleukin-8; Intestinal Mucosa; NF-kappa B p50 Subunit; Proto-Oncogene Proteins c-rel; RNA, Messenger; Shigella boydii; Shigella flexneri; STAT1 Transcription Factor; STAT2 Transcription Factor; Transcription Factor RelA; Transcriptional Activation; Virulence; Virulence Factors

The enteroinvasive bacterium Shigella flexneri uses multiple secreted effector proteins to downregulate interleukin-8 (IL-8) expression in infected epithelial cells. Yet, massive IL-8 secretion is observed in Shigellosis. Here we report a host mechanism of cell-cell communication that circumvents the effector proteins and strongly amplifies IL-8 expression during bacterial infection. By monitoring proinflammatory signals at the single-cell level, we found that the activation of the transcription factor NF-κB and the MAP kinases JNK, ERK, and p38 rapidly propagated from infected to uninfected adjacent cells, leading to IL-8 production by uninfected bystander cells. Bystander IL-8 production was also observed during Listeria monocytogenes and Salmonella typhimurium infection. This response could be triggered by recognition of peptidoglycan and is mediated by gap junctions. Thus, we have identified a mechanism of cell-cell communication that amplifies innate immunity against bacterial infection by rapidly spreading proinflammatory signals via gap junctions to yet uninfected cells.

Topics: Caco-2 Cells; Cell Communication; Cell Proliferation; Dysentery, Bacillary; Gap Junctions; HeLa Cells; Humans; Immunity, Innate; Interleukin-8; Listeria monocytogenes; Listeriosis; MAP Kinase Signaling System; Mitogen-Activated Protein Kinases; NF-kappa B; Peptidoglycan; Shigella flexneri

Saccharomyces boulardii is gaining in popularity as a treatment for a variety of diarrheal diseases as well as inflammatory bowel disease. This study was designed to examine the effect of this yeast on infection by Shigella flexneri, a highly infectious and human host-adapted enteric pathogen. We investigated key interactions between the bacteria and host cells in the presence of the yeast in addition to a number of host responses including proinflammatory events and markers. Although the presence of the yeast during infection did not alter the number of bacteria that was able to attach or invade human colon cancer-derived T-84 cells, it did positively impact the tight junction protein zonula occluden-2 and significantly increase the barrier integrity of model epithelia. The yeast also decreased ERK, JNK, and NF-kappaB activation in response to S. flexneri, events likely responsible for the observed reductions in IL-8 secretion and the transepithelial migration of polymorphonuclear leukocytes across T-84 monolayers. These results, suggesting that the yeast allowed for a dampened inflammatory response, were confirmed in vivo utilizing a highly relevant model of human fetal colonic tissue transplanted into scid mice. Furthermore, a cell-free S. boulardii culture supernatant was also capable of reducing IL-8 secretion by infected T-84 cells. These data suggest that although the use of S. boulardii during infection with S. flexneri may alleviate symptoms associated with the inflammatory response of the host, it would not prevent infection.

Topics: Animals; Blotting, Western; Cell Movement; Cells, Cultured; Colon; Colony Count, Microbial; Dysentery, Bacillary; Electric Impedance; Female; Humans; Interleukin-8; Intestinal Mucosa; Mice; Mice, SCID; Neutrophils; NF-kappa B; Phosphorylation; Pregnancy; Saccharomyces; Shigella flexneri; Signal Transduction; Tight Junctions; Transplantation, Heterologous

It has been difficult to evaluate the protective efficacy of vaccine candidates against shigellosis, a major form of bacillary dysentery caused by Shigella spp. infection, because of the lack of suitable animal models. To develop a proper animal model representing human bacillary dysentery, guinea pigs were challenged with virulent Shigella flexneri serotype 2a (strains 2457T or YSH6000) or S. flexneri 5a (strain M90T) by the intrarectal (i.r.) route. Interestingly, all guinea pigs administered these Shigella strains developed severe and acute rectocolitis. They lost approximately 20% of their body weight and developed tenesmus by 24 h after Shigella infection. Shigella invasion and colonization of the distal colon were seen at 24 h but disappeared by 48 h following i.r. infection. Histopathological approaches demonstrated significant damage and destruction of mucosal and submucosal layers, thickened intestinal wall, edema, erosion, infiltration of neutrophils, and depletion of goblet cells in the distal colon. Furthermore, robust expression of IL-8, IL-1beta, and inducible NO synthase mRNA was detected in the colon from 6 to 24 h following Shigella infection. Most importantly, in our new shigellosis model, guinea pigs vaccinated with an attenuated S. flexneri 2a SC602 strain possessing high levels of mucosal IgA Abs showed milder symptoms of bacillary dysentery than did animals receiving PBS alone after Shigella infection. In the guinea pig, administration of Shigella by i.r. route induces acute inflammation, making this animal model useful for assessing the protective efficacy of Shigella vaccine candidates.

Topics: Acute Disease; Animals; Colon; Dysentery, Bacillary; Female; Gene Expression Regulation; Guinea Pigs; Immunoglobulin A; Inflammation; Interleukin-1beta; Interleukin-8; Models, Animal; Nitric Oxide Synthase; Proctocolitis; Shigella flexneri; Shigella Vaccines; Vaccines, Attenuated

Early identification of the pathogen causing acute gastroenteritis in children helps the physicians managing the disease and prevents unnecessary antibiotic treatment. C-reactive protein (CRP), interleukin (IL) 6 and IL-8 play a major role in immune responses and have been studied in a large number of infectious and noninfectious inflammatory diseases. The purpose of this study was to determine the serum IL-6 and IL-8 concentrations early in the course of acute gastroenteritis to see if these cytokines were useful diagnostic markers in differentiating viral from bacterial gastroenteritis.. Interleukin 6, IL-8 and CRP were measured in 18 patients with bacterial gastroenteritis, 21 patients with viral gastroenteritis and 17 healthy children.. Interleukin 6 and CRP concentrations in patients with bacterial gastroenteritis were significantly higher than those in patients with viral gastroenteritis and healthy controls (P < 0.001). IL-8 concentrations in patients with viral and bacterial gastroenteritis were both increased and were not statistically different. IL-6 and IL-8 levels had diagnostic sensitivities of 79% and 50% and specificities of 86% and 67%, respectively. The combination of IL-6 and CRP had a sensitivity of 94%, specificity of 71%, a positive predictive value of 74% and a negative predictive value of 93.75%.. Serum IL-6 may be a useful marker for early differentiation of viral and bacterial gastroenteritis in children, especially in combination with CRP.

Topics: Acute Disease; Biomarkers; C-Reactive Protein; Case-Control Studies; Child, Preschool; Dysentery, Bacillary; Enzyme-Linked Immunosorbent Assay; Female; Gastroenteritis; Humans; Infant; Interleukin-6; Interleukin-8; Male; Rotavirus Infections; Salmonella Infections; Sensitivity and Specificity; Yersinia Infections

The lack of a mouse model of acute rectocolitis mimicking human bacillary dysentery in the presence of invasive Shigella is a major handicap to study the pathogenesis of the disease and to develop a Shigella vaccine. The inability of the mouse intestinal mucosa to elicit an inflammatory infiltrate composed primarily of polymorphonuclear leukocytes (PMN) may be due to a defect in epithelial invasion, in the sensing of invading bacteria, or in the effector mechanisms that recruit the PMN infiltrate. We demonstrate that the BALB/cJ mouse colonic epithelium not only can be invaded by Shigella, but also elicits an inflammatory infiltrate that, however, lacks PMN. This observation points to a major defect of mice in effector mechanisms, particularly the lack of expression of the CXC chemokine, IL-8. Indeed, this work demonstrates that the delivery of recombinant human IL-8, together with Shigella infection of the colonic epithelial surface, causes an acute colitis characterized by a strong PMN infiltrate that, by all criteria, including transcription profiles of key mediators of the innate/inflammatory response and histopathological lesions, mimics bacillary dysentery. This is a major step forward in the development of a murine model of bacillary dysentery.

Topics: Animals; Chemokine CXCL1; Chemokine CXCL2; Chemokines; Chemokines, CXC; Colitis; Colon; Cytokines; Disease Models, Animal; Dysentery, Bacillary; Humans; Immunohistochemistry; Interleukin-8; Intestinal Mucosa; Kinetics; Lipopolysaccharides; Male; Mice; Neutrophil Infiltration; Neutrophils; Peroxidase; Recombinant Proteins; Shigella flexneri; Species Specificity; Transcription, Genetic

Shigellae infect human intestine and cause intense inflammation and destruction of colonic and rectal mucosa. To model the interactions of shigella with human intestine in vivo, we have studied shigella infection in human intestinal xenografts in severe combined immunodeficient mice (SCID-HU-INT mice). Inoculation of shigella into human intestinal xenografts caused severe inflammation and mucosal damage, which was apparent as soon as 4 h following infection. Shigella infection was associated with human intestinal production of interleukin-1B (IL-1B) and IL-8 and a marked neutrophil influx into the graft. Depletion of neutrophils from SCID-HU-INT mice reduced inflammation in the human intestinal xenograft in response to shigella infection but failed to significantly alter tissue damage. However, the number of intracellular bacteria was more than 20-fold higher in the human intestinal xenografts from neutrophil-depleted SCID-HU-INT mice. Infection of human intestinal xenografts with an attenuated vaccine strain of shigella (CVD1203) induced lower levels of IL-1B and IL-8 than wild-type shigella and caused only moderate damage to the intestinal permeability barrier. Our studies establish the SCID-HU-INT mouse as a viable model for studying the interactions between shigella and human intestine and indicate that neutrophils are important for controlling the invasion of human intestine by shigella.

Topics: Animals; Cell Movement; Dysentery, Bacillary; Humans; Interleukin-1; Interleukin-8; Intestines; Mice; Mice, SCID; Models, Animal; Neutrophils; Permeability; Transplantation, Heterologous

In shigellosis, the network of cellular interactions mediated by a balance of pro- and anti-inflammatory cytokines or chemokines is clearly tipped toward acute destructive inflammation of intestinal tissues by the bacterial invader. This work has addressed the role played by interleukin-8 (IL-8) in a rabbit model of intestinal invasion by Shigella flexneri. IL-8, which is largely produced by the epithelial cells themselves, appears to be a major mediator of the recruitment of polymorphonuclear leukocytes (PMNs) to the subepithelial area and transmigration of these cells through the epithelial lining. Neutralization of IL-8 function by monoclonal antibody WS-4 caused a decrease in the amount of PMNs streaming through the lamina propria and the epithelium, thus significantly attenuating the severity of epithelial lesions in areas of bacterial invasion. These findings are in agreement with our previous work (31). In contrast to the PMNs, the bacteria displayed increased transepithelial translocation, as well as overgrowth in the lamina propria and increased passage into the mesenteric blood. By mediating eradication of bacteria at their epithelial entry site, although at the cost of severe epithelial destruction, IL-8 therefore appears to be a key chemokine in the control of bacterial translocation.

Topics: Animals; Bacterial Translocation; Dysentery, Bacillary; Immunohistochemistry; Interleukin-8; Intestinal Mucosa; Neutrophils; Rabbits; Shigella flexneri