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Find video protocols related to scientific articles indexed in Pubmed.
Caspase-8 controls the gut response to microbial challenges by Tnf-?-dependent and independent pathways.
Gut
PUBLISHED: 11-08-2014
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Intestinal epithelial cells (IEC) express toll-like receptors (TLR) that facilitate microbial recognition. Stimulation of TLR ligands induces a transient increase in epithelial cell shedding, a mechanism that serves the antibacterial and antiviral host defence of the epithelium and promotes elimination of intracellular pathogens. Although activation of the extrinsic apoptosis pathway has been described during inflammatory shedding, its functional involvement is currently unclear.
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Gut microbiota: a natural adjuvant for vaccination.
Immunity
PUBLISHED: 09-20-2014
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In this issue of Immunity, Oh et al. (2014) reveal an unappreciated facet of how the microbiota influences immune responses. Immunity to nonadjuvanted vaccines depends on Toll-like-receptor-5-mediated sensing of the microbiota.
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TRIF signaling drives homeostatic intestinal epithelial antimicrobial peptide expression.
J. Immunol.
PUBLISHED: 09-10-2014
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Recent results indicate a significant contribution of innate immune signaling to maintain mucosal homeostasis, but the precise underlying signal transduction pathways are ill-defined. By comparative analysis of intestinal epithelial cells isolated from conventionally raised and germ-free mice, as well as animals deficient in the adaptor molecules MyD88 and TRIF, the TLR3 and TLR4, as well as the type I and III IFN receptors, we demonstrate significant TLR-mediated signaling under homeostatic conditions. Surprisingly, homeostatic expression of Reg3? and Paneth cell enteric antimicrobial peptides critically relied on TRIF and, in part, TLR3 but was independent of IFN receptor signaling. Reduced antimicrobial peptide expression was associated with significantly lower numbers of Paneth cells and a reduced Paneth cell maturation and differentiation factor expression in TRIF mutant compared with wild-type epithelium. This phenotype was not transferred to TRIF-sufficient germ-free animals during cohousing. Low antimicrobial peptide expression in TRIF-deficient mice caused reduced immediate killing of orally administered bacteria but was not associated with significant alterations in the overall composition of the enteric microbiota. The phenotype was rapidly restored in a TRIF-independent fashion after transient epithelial damage. Our results identify TRIF signaling as a truly homeostatic pathway to maintain intestinal epithelial barrier function revealing fundamental differences in the innate immune signaling between mucosal homeostasis and tissue repair.
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Age-dependent enterocyte invasion and microcolony formation by Salmonella.
PLoS Pathog.
PUBLISHED: 09-01-2014
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The coordinated action of a variety of virulence factors allows Salmonella enterica to invade epithelial cells and penetrate the mucosal barrier. The influence of the age-dependent maturation of the mucosal barrier for microbial pathogenesis has not been investigated. Here, we analyzed Salmonella infection of neonate mice after oral administration. In contrast to the situation in adult animals, we observed spontaneous colonization, massive invasion of enteroabsorptive cells, intraepithelial proliferation and the formation of large intraepithelial microcolonies. Mucosal translocation was dependent on enterocyte invasion in neonates in the absence of microfold (M) cells. It further resulted in potent innate immune stimulation in the absence of pronounced neutrophil-dominated pathology. Our results identify factors of age-dependent host susceptibility and provide important insight in the early steps of Salmonella infection in vivo. We also present a new small animal model amenable to genetic manipulation of the host for the analysis of the Salmonella enterocyte interaction in vivo.
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Experimental Colitis Is Exacerbated by Concomitant Infection with Mycobacterium avium ssp. paratuberculosis.
Inflamm. Bowel Dis.
PUBLISHED: 08-22-2014
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Crohn's disease (CD) is a chronic inflammatory disorder of the human gastrointestinal tract. Although genetic, immunological, environmental, and bacterial factors have been implicated, the pathogenesis is incompletely understood. The histopathological appearance of CD strikingly resembles Johne's disease, a ruminant inflammatory bowel disease, caused by Mycobacterium avium ssp. paratuberculosis (MAP), but a causative role of MAP in CD has not been established. In this work, we hypothesized that MAP might exacerbate an already existing intestinal disease.
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Facts, myths and hypotheses on the zoonotic nature of Mycobacterium avium subspecies paratuberculosis.
Int. J. Med. Microbiol.
PUBLISHED: 07-25-2014
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Mycobacterium avium subspecies paratuberculosis (MAP) is the causative agent of paratuberculosis (Johne's disease [JD]), a chronic granulomatous enteritis in ruminants. JD is one of the most widespread bacterial diseases of domestic animals with significant economic impact. The histopathological picture of JD resembles that of Crohn's disease (CD), a human chronic inflammatory bowel disease of still unresolved aetiology. An aetiological relevance of MAP for CD has been proposed. This and the ambiguity of other published epidemiological findings raise the question whether MAP represents a zoonotic agent. In this review, we will discuss evidence that MAP has zoonotic capacity.
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Maturation of the enteric mucosal innate immune system during the postnatal period.
Immunol. Rev.
PUBLISHED: 06-20-2014
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The innate immune system instructs the host on microbial exposure and infection. This information is critical to mount a protective innate and adaptive host response to microbial challenge, but is also involved in homeostatic and adaptive processes that adjust the organism to meet environmental requirements. This is of particular importance for the neonatal host during the transition from the protected fetal life to the intense and dynamic postnatal interaction with commensal and pathogenic microorganisms. Here, we discuss both adaptive and developmental mechanisms of the mucosal innate immune system that prevent inappropriate stimulation and facilitate establishment of a stable homeostatic host-microbial interaction after birth.
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Intestinal mucus affinity and biological activity of an orally administered antibacterial and anti-inflammatory peptide.
Gut
PUBLISHED: 05-10-2014
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Antimicrobial peptides (AMP) provide protection from infection by pathogenic microorganisms and restrict bacterial growth at epithelial surfaces to maintain mucosal homeostasis. In addition, they exert a significant anti-inflammatory activity. Here we analysed the anatomical distribution and biological activity of an orally administered AMP in the context of bacterial infection and host-microbial homeostasis.
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Outer ear canal infection with Rhabditis sp. nematodes in a human.
J. Clin. Microbiol.
PUBLISHED: 03-05-2014
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Here we report the first human case of an outer ear canal infection with a free-living nematode of the genus Rhabditis. Otomicroscopy revealed viable worms in the outer ear canal of a patient suffering from chronic otorrhea and hearing loss. The nematode was identified by microscopy and internal transcribed spacer (ITS)-PCR.
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Interleukin-13-mediated paneth cell degranulation and antimicrobial peptide release.
J Innate Immun
PUBLISHED: 02-19-2014
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Paneth cell-derived enteric antimicrobial peptides significantly contribute to antibacterial host defense and host-microbial homeostasis. Regulation occurs by enzymatic processing and release into the small intestinal lumen, but the stimuli involved are incompletely understood. Here, the capacity of various microbial and immune stimuli to induce antimicrobial peptide release from small intestinal tissue was systematically evaluated using antibacterial activity testing, immunostaining for Paneth cell granules and mass spectrometry. We confirmed the stimulatory activity of the muscarinic receptor agonist carbachol and the nucleotide-binding oligomerization domain ligand muramyl dipeptide. In contrast, no release of antibacterial activity was noted after treatment with the Toll-like receptor ligands poly(I:C), lipopolysaccharide or CpG, and the cytokines interleukin (IL)-15, IL-22, IL-28 and interferon-?. Rapid Paneth cell degranulation and antimicrobial activity release, however, was observed after stimulation with the endogenous mediators IL-4 and IL-13. This process required phosphatidylinositol 3-kinase and was associated with protein kinase B phosphorylation in Paneth cells. Flow cytometric analysis confirmed expression of the IL-13 receptor ?1 on isolated Paneth cells. Our findings identify a novel role of IL-13 as inducer of Paneth cell degranulation and enteric antimicrobial peptide release. IL-13 may thus contribute to mucosal antimicrobial host defense and host microbial homeostasis.
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Norovirus triggered microbiota-driven mucosal inflammation in interleukin 10-deficient mice.
Inflamm. Bowel Dis.
PUBLISHED: 02-04-2014
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Infection may trigger clinically overt mucosal inflammation in patients with predisposition for inflammatory bowel disease. However, the impact of particular enteropathogenic microorganisms is ill-defined. In this study, the influence of murine norovirus (MNV) infection on clinical, histopathological, and immunological features of mucosal inflammation in the IL10-deficient (Il10) mouse model of inflammatory bowel disease was examined.
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Ontogeny of intestinal epithelial innate immune responses.
Front Immunol
PUBLISHED: 01-01-2014
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Emerging evidence indicates that processes during postnatal development might significantly influence the establishment of mucosal host-microbial homeostasis. Developmental and adaptive immunological processes but also environmental and microbial exposure early after birth might thus affect disease susceptibility and health during adult life. The present review aims at summarizing the current understanding of the intestinal epithelial innate immune system and its developmental and adaptive changes after birth.
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Systemic and mucosal immune reactivity upon Mycobacterium avium ssp. paratuberculosis infection in mice.
PLoS ONE
PUBLISHED: 01-01-2014
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Mycobacterium avium ssp. paratuberculosis (MAP) is the cause of Johne's disease, an inflammatory bowel disorder of ruminants. Due to the similar pathology, MAP was also suggested to cause Crohn's disease (CD). Despite of intensive research, this question is still not settled, possibly due to the lack of versatile mouse models. The aim of this study was to identify basic immunologic mechanisms in response to MAP infection. Immune compromised C57BL/6 Rag2-/- mice were infected with MAP intraperitoneally. Such chronically infected mice were then reconstituted with CD4+ and CD8+ T cells 28 days after infection. A systemic inflammatory response, detected as enlargement of the spleen and granuloma formation in the liver, was observed in mice infected and reconstituted with CD4+ T cells. Whereby inflammation in infected and CD4+CD45RB(hi) T cell reconstituted animals was always higher than in the other groups. Reconstitution of infected animals with CD8+ T cells did not result in any inflammatory signs. Interestingly, various markers of inflammation were strongly up-regulated in the colon of infected mice reconstituted with CD4+CD45RB(lo/int) T cells. We propose, the usual non-colitogenic CD4+CD45RB(lo/int) T cells were converted into inflammatory T cells by the interaction with MAP. However, the power of such cells might be not sufficient for a fully established inflammatory response in the colon. Nevertheless, our model system appears to mirror aspects of an inflammatory bowel disease (IBD) like CD and Johne's diseases. Thus, it will provide an experimental platform on which further knowledge on IBD and the involvement of MAP in the induction of CD could be acquired.
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Handle energy resources with care.
Trends Microbiol.
PUBLISHED: 12-02-2013
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While politicians discuss rules to improve the efficacy of global energy consumption and to limit environmental pollution, the mammalian body has numerous mechanisms to efficiently use nutrient resources and appropriately extract energy. A new mechanism has recently been identified by the group of Fredrik Bäckhed in Cell Host & Microbe.
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IFIT2 is an effector protein of type I IFN-mediated amplification of lipopolysaccharide (LPS)-induced TNF-? secretion and LPS-induced endotoxin shock.
J. Immunol.
PUBLISHED: 09-06-2013
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Type I IFN signaling amplifies the secretion of LPS-induced proinflammatory cytokines such as TNF-? or IL-6 and might thus contribute to the high mortality associated with Gram-negative septic shock in humans. The underlying molecular mechanism, however, is ill defined. In this study, we report the generation of mice deficient in IFN-induced protein with tetratricopeptide repeats 2 (Ifit2) and demonstrate that Ifit2 is a critical signaling intermediate for LPS-induced septic shock. Ifit2 expression was significantly upregulated in response to LPS challenge in an IFN-? receptor- and IFN regulatory factor (Irf)9-dependent manner. Also, LPS induced secretion of IL-6 and TNF-? by bone marrow-derived macrophages (BMDMs) was significantly enhanced in the presence of Ifit2. In accordance, Ifit2-deficient mice exhibited significantly reduced serum levels of IL-6 and TNF-? and reduced mortality in an endotoxin shock model. Investigation of the underlying signal transduction events revealed that Ifit2 upregulates Irf3 phosphorylation. In the absence of Irf3, reduced Ifn-? mRNA expression and Ifit2 protein expression after LPS stimulation was found. Also, Tnf-? and Il-6 secretion but not Tnf-? and Il-6 mRNA expression levels were reduced. Thus, IFN-stimulated Ifit2 via enhanced Irf3 phosphorylation upregulates the secretion of proinflammatory cytokines. It thereby amplifies LPS-induced cytokine production and critically influences the outcome of endotoxin shock.
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Preclinical investigations reveal the broad-spectrum neutralizing activity of peptide Pep19-2.5 on bacterial pathogenicity factors.
Antimicrob. Agents Chemother.
PUBLISHED: 01-14-2013
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Bacterial infections are known to cause severe health-threatening conditions, including sepsis. All attempts to get this disease under control failed in the past, and especially in times of increasing antibiotic resistance, this leads to one of the most urgent medical challenges of our times. We designed a peptide to bind with high affinity to endotoxins, one of the most potent pathogenicity factors involved in triggering sepsis. The peptide Pep19-2.5 reveals high endotoxin neutralization efficiency in vitro, and here, we demonstrate its antiseptic/anti-inflammatory effects in vivo in the mouse models of endotoxemia, bacteremia, and cecal ligation and puncture, as well as in an ex vivo model of human tissue. Furthermore, we show that Pep19-2.5 can bind and neutralize not only endotoxins but also other bacterial pathogenicity factors, such as those from the Gram-positive bacterium Staphylococcus aureus. This broad neutralization efficiency and the additive action of the peptide with common antibiotics makes it an exceptionally appropriate drug candidate against bacterial sepsis and also offers multiple other medication opportunities.
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The anti-inflammatory effect of the synthetic antimicrobial peptide 19-2.5 in a murine sepsis model: a prospective randomized study.
Crit Care
PUBLISHED: 01-07-2013
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ABSTRACT: INTRODUCTION: Increasing rates of multi-resistant bacteria are a major problem in the treatment of critically ill patients. Furthermore, conventional antibiotics lead to the release of bacterial derived membrane parts initiating pro-inflammatory cascades with potential harm to the patient. Antimicrobial peptides (AMP) may kill bacteria without releasing pro-inflammatory factors. Thus, we compared three newly developed synthetic anti-lipopolysaccharide peptides (SALPs) with a broader range of efficacy to suppress cytokine release in plasma and CD14 mRNA expression in organ tissue in a murine, polymicrobial sepsis model. METHODS: A randomized, experimental trial was conducted in an animal research facility. Male NMRI mice (n = 90; 8- to 12-weeks old) were randomized to the following six groups: (i) sham operation and parenteral vehicle (NaCl 0.9%) administration (sham); (ii) cecal ligation and puncture (CLP) and vehicle infusion (sepsis-control), (iii) CLP and polymyxin B infusion (polyB), or (iv to vi) CLP and infusion of three different synthetic antimicrobial peptides Peptide 19-2.5 (Pep2.5), Peptide 19-4 (Pep4) or Peptide 19-8 (Pep8). All animals underwent arterial and venous catheterization for hemodynamic monitoring 48 hours prior to CLP or sham-operation. Physical appearance and behavior (activity), plasma cytokine levels, and CD14 mRNA expression in heart, lung, liver, spleen and kidney tissue were determined 24 hours after CLP or sham operation. RESULTS: Only Pep2.5 significantly enhanced the activity after CLP, whereas none of the therapeutic regimens elevated the mean arterial pressure or heart rate. The strongly elevated IL-6, IL-10 and monocyte chemoattractant protein serum levels in septic animals were significantly reduced after Pep2.5 administration (P < 0.001, P < 0.001, and P < 0.001, respectively). Similarly, Pep2.5 significantly reduced the sepsis-induced CD14 mRNA expression in heart (P = 0.003), lung (P = 0.008), and spleen tissue (P = 0.009) but not in kidney and liver. CONCLUSIONS: Structurally variable SALPs exhibit major differences in their anti-inflammatory effect in vivo. Continuous parenteral administration of Pep2.5 is able to reduce sepsis-induced cytokine release and tissue inflammation.
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Generation of mouse small intestinal epithelial cell lines that allow the analysis of specific innate immune functions.
PLoS ONE
PUBLISHED: 01-01-2013
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Cell lines derived from the small intestine that reflect authentic properties of the originating intestinal epithelium are of high value for studies on mucosal immunology and host microbial homeostasis. A novel immortalization procedure was applied to generate continuously proliferating cell lines from murine E19 embryonic small intestinal tissue. The obtained cell lines form a tight and polarized epithelial cell layer, display characteristic tight junction, microvilli and surface protein expression and generate increasing transepithelial electrical resistance during in vitro culture. Significant up-regulation of Cxcl2 and Cxcl5 chemokine expression upon exposure to defined microbial innate immune stimuli and endogenous cytokines is observed. Cell lines were also generated from a transgenic interferon reporter (Mx2-Luciferase) mouse, allowing reporter technology-based quantification of the cellular response to type I and III interferon. Thus, the newly created cell lines mimic properties of the natural epithelium and can be used for diverse studies including testing of the absorption of drug candidates. The reproducibility of the method to create such cell lines from wild type and transgenic mice provides a new tool to study molecular and cellular processes of the epithelial barrier.
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The impact of perinatal immune development on mucosal homeostasis and chronic inflammation.
Nat. Rev. Immunol.
PUBLISHED: 12-09-2011
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The mucosal surfaces of the gut and airways have important barrier functions and regulate the induction of immunological tolerance. The rapidly increasing incidence of chronic inflammatory disorders of these surfaces, such as inflammatory bowel disease and asthma, indicates that the immune functions of these mucosae are becoming disrupted in humans. Recent data indicate that events in prenatal and neonatal life orchestrate mucosal homeostasis. Several environmental factors promote the perinatal programming of the immune system, including colonization of the gut and airways by commensal microorganisms. These complex microbial-host interactions operate in a delicate temporal and spatial manner and have an important role in the induction of homeostatic mechanisms.
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The mammalian intestinal epithelium as integral player in the establishment and maintenance of host-microbial homeostasis.
Semin. Immunol.
PUBLISHED: 12-03-2011
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Only one single layer of epithelial cells separates the densely colonized and environmentally exposed intestinal lumen from the largely sterile subepithelial tissue. Together with the overlaying mucus and the subepithelial mucosal immune system the epithelium has evolved to maintain homeostasis in the presence of the enteric microbiota. It also contributes to rapid and efficient antimicrobial host defence in the event of infection with pathogenic microorganisms. Both, epithelial antimicrobial host defence and homeostasis rely on signalling pathways induced by innate immune receptors demonstrating the active role of epithelial cells in the host-microbial interplay. The interaction of epithelial cells with professional immune cells illustrates the integrated function within the mucosal tissue. In the present review we focus on structural and functional changes of the intestinal epithelium during the fetal-neonatal transition and infancy and try to delineate its role in the induction and maintenance of host-microbial homeostasis. We also address factors that impair epithelial functions and may lead to disruption of the mucosal barrier, tissue damage and the development of symptomatic disease.
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Establishment of intestinal homeostasis during the neonatal period.
Cell. Mol. Life Sci.
PUBLISHED: 09-06-2011
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The intestinal mucosa faces the challenge of regulating the balance between immune tolerance towards commensal bacteria, environmental stimuli and food antigens on the one hand, and induction of efficient immune responses against invading pathogens on the other hand. This regulatory task is of critical importance to prevent inappropriate immune activation that may otherwise lead to chronic inflammation, tissue disruption and organ dysfunction. The most striking example for the efficacy of the adaptive nature of the intestinal mucosa is birth. Whereas the body surfaces are protected from environmental and microbial exposure during fetal life, bacterial colonization and contact with potent immunostimulatory substances start immediately after birth. In the present review, we summarize the current knowledge on the mechanisms underlying the transition of the intestinal mucosa during the neonatal period leading to the establishment of a stable, life-long host-microbial homeostasis. The environmental exposure and microbial colonization during the neonatal period, and also the influence of maternal milk on the immune protection of the mucosa and the role of antimicrobial peptides, are described. We further highlight the molecular mechanisms of innate immune tolerance in neonatal intestinal epithelium. Finally, we link the described immunoregulatory mechanisms to the increased susceptibility to inflammatory and infectious diseases during the neonatal period.
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IFN-lambda determines the intestinal epithelial antiviral host defense.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 04-25-2011
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Type I and type III IFNs bind to different cell-surface receptors but induce identical signal transduction pathways, leading to the expression of antiviral host effector molecules. Despite the fact that type III IFN (IFN-?) has been shown to predominantly act on mucosal organs, in vivo infection studies have failed to attribute a specific, nonredundant function. Instead, a predominant role of type I IFN was observed, which was explained by the ubiquitous expression of the type I IFN receptor. Here we comparatively analyzed the role of functional IFN-? and type I IFN receptor signaling in the innate immune response to intestinal rotavirus infection in vivo, and determined viral replication and antiviral gene expression on the cellular level. We observed that both suckling and adult mice lacking functional receptors for IFN-? were impaired in the control of oral rotavirus infection, whereas animals lacking functional receptors for type I IFN were similar to wild-type mice. Using Mx1 protein accumulation as marker for IFN responsiveness of individual cells, we demonstrate that intestinal epithelial cells, which are the prime target cells of rotavirus, strongly responded to IFN-? but only marginally to type I IFN in vivo. Systemic treatment of suckling mice with IFN-? repressed rotavirus replication in the gut, whereas treatment with type I IFN was not effective. These results are unique in identifying a critical role of IFN-? in the epithelial antiviral host defense.
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Cesarean delivery is associated with celiac disease but not inflammatory bowel disease in children.
Gut Microbes
PUBLISHED: 03-01-2011
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The postnatal period represents a particularly dynamic phase in the establishment of the host-microbial homeostasis. The sterile protected intestinal mucosa of the fetus becomes exposed to and subsequently colonized by a complex and diverse bacterial community. Both, the exposure to microbial ligands and the bacterial colonization have been described to differ between neonates born vaginally or by cesarean delivery. These differences might influence the development of the mucosal immune system, the establishment of a stable intestinal host-microbial homeostasis, and ultimately contribute to the risk to acquire immune mediated diseases later in life. Indeed, an increased risk for atopic diseases such as allergic rhinitis and asthma was reported in children born by cesarean delivery. Our recent study described an association between cesarean delivery and celiac disease. Here we summarize the available information on postnatal microbial colonization and the influence of the mode of delivery on flora composition and host microbial homeostasis. We discuss possible consequences of the mode of delivery on epithelial barrier function and the establishment of the mucosal immune system and speculate on functional links between flora alterations and the development of inappropriate host immune responses that may contribute to enteric inflammatory diseases. 
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New antiseptic peptides to protect against endotoxin-mediated shock.
Antimicrob. Agents Chemother.
PUBLISHED: 07-06-2010
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Systemic bacterial infections are associated with high mortality. The access of bacteria or constituents thereof to systemic circulation induces the massive release of immunomodulatory mediators, ultimately causing tissue hypoperfusion and multiple-organ failure despite adequate antibiotic treatment. Lipid A, the "endotoxic principle" of bacterial lipopolysaccharide (LPS), is one of the major bacterial immunostimuli. Here we demonstrate the biological efficacy of rationally designed new synthetic antilipopolysaccharide peptides (SALPs) based on the Limulus anti-LPS factor for systemic application. We show efficient inhibition of LPS-induced cytokine release and protection from lethal septic shock in vivo, whereas cytotoxicity was not observed under physiologically relevant conditions and concentrations. The molecular mechanism of LPS neutralization was elucidated by biophysical techniques. The lipid A part of LPS is converted from its "endotoxic conformation," the cubic aggregate structure, into an inactive multilamellar structure, and the binding affinity of the peptide to LPS exceeds those of known LPS-binding proteins, such as LPS-binding protein (LBP). Our results thus delineate a novel therapeutic strategy for the clinical management of patients with septic shock.
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Cesarean delivery is associated with celiac disease but not inflammatory bowel disease in children.
Pediatrics
PUBLISHED: 05-17-2010
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The aim of this study was to analyze a possible association between cesarean delivery and enteric inflammatory diseases in children.
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miR-146a mediates protective innate immune tolerance in the neonate intestine.
Cell Host Microbe
PUBLISHED: 04-22-2010
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After birth, the intestinal mucosa undergoes a dramatic transition from a sterile protected site to an environmentally exposed and permanently colonized surface. The mechanisms that facilitate this transition are ill defined. Here, we demonstrate that microRNA-146a-mediated translational repression and proteolytic degradation of the essential Toll-like receptor (TLR) signaling molecule interleukin 1 receptor associated kinase 1 (IRAK1) is sufficient to induce intestinal epithelial innate immune tolerance and provide protection from bacteria-induced epithelial damage in neonates. Despite low IRAK1 protein levels, continuous TLR4- and IRAK1-dependent signal transduction induced by intraepithelial endotoxin persistence during the neonatal period maintains tolerance through sustained miR-146a expression. Strikingly, it additionally facilitates transcription of a distinct set of genes involved in cell survival, differentiation, and homeostasis. Thus, our results identify the underlying molecular mechanisms of intestinal epithelial innate immune tolerance during the neonatal period and characterize tolerance as an active condition involved in the establishment of intestinal mucosal homeostasis.
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Potentiation of epithelial innate host responses by intercellular communication.
PLoS Pathog.
PUBLISHED: 02-05-2010
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The epithelium efficiently attracts immune cells upon infection despite the low number of pathogenic microbes and moderate levels of secreted chemokines per cell. Here we examined whether horizontal intercellular communication between cells may contribute to a coordinated response of the epithelium. Listeria monocytogenes infection, transfection, and microinjection of individual cells within a polarized intestinal epithelial cell layer were performed and activation was determined at the single cell level by fluorescence microscopy and flow cytometry. Surprisingly, chemokine production after L. monocytogenes infection was primarily observed in non-infected epithelial cells despite invasion-dependent cell activation. Whereas horizontal communication was independent of gap junction formation, cytokine secretion, ion fluxes, or nitric oxide synthesis, NADPH oxidase (Nox) 4-dependent oxygen radical formation was required and sufficient to induce indirect epithelial cell activation. This is the first report to describe epithelial cell-cell communication in response to innate immune activation. Epithelial communication facilitates a coordinated infectious host defence at the very early stage of microbial infection.
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Internalization-dependent recognition of Mycobacterium avium ssp. paratuberculosis by intestinal epithelial cells.
Cell. Microbiol.
PUBLISHED: 08-13-2009
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Mycobacterium avium ssp. paratuberculosis (MAP) is the causative agent of Johnes disease, a highly prevalent chronic intestinal infection in domestic and wildlife ruminants. The microbial pathogenesis of MAP infection has attracted additional attention due to an association with the human enteric inflammatory Crohns disease. MAP is acquired by the faecal-oral route prompting us to study the interaction with differentiated intestinal epithelial cells. MAP was rapidly internalized and accumulated in a late endosomal compartment. In contrast to other opportunistic mycobacteria or M. bovis, MAP induced significant epithelial activation as indicated by a NF-kappaB-independent but Erk-dependent chemokine secretion. Surprisingly, MAP-induced chemokine production was completely internalization-dependent as inhibition of Rac-dependent bacterial uptake abolished epithelial activation. In accordance, innate immune recognition of MAP by differentiated intestinal epithelial cells occurred through the intracellularly localized pattern recognition receptors toll-like receptor 9 and NOD1 with signal transduction via the adaptor molecules MyD88 and RIP2. The internalization-dependent innate immune activation of intestinal epithelial cells is in contrast to the stimulation of professional phagocytes by extracellular bacterial constituents and might significantly contribute to the histopathological changes observed during enteric MAP infection.
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Intravenous tigecycline as adjunctive or alternative therapy for severe refractory Clostridium difficile infection.
Clin. Infect. Dis.
PUBLISHED: 05-14-2009
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Clostridium difficile infection (CDI) has become more refractory to standard therapy. We describe 4 patients with severe refractory CDI who were successfully treated with tigecycline. Symptoms improved within 1 week. No relapses were observed. This favorable outcome suggests that tigecycline might be a useful alternative for treating severe refractory CDI.
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O-antigen delays lipopolysaccharide recognition and impairs antibacterial host defense in murine intestinal epithelial cells.
PLoS Pathog.
PUBLISHED: 03-09-2009
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Although Toll-like receptor (TLR) 4 signals from the cell surface of myeloid cells, it is restricted to an intracellular compartment and requires ligand internalization in intestinal epithelial cells (IECs). Yet, the functional consequence of cell-type specific receptor localization and uptake-dependent lipopolysaccharide (LPS) recognition is unknown. Here, we demonstrate a strikingly delayed activation of IECs but not macrophages by wildtype Salmonella enterica subsp. enterica sv. (S.) Typhimurium as compared to isogenic O-antigen deficient mutants. Delayed epithelial activation is associated with impaired LPS internalization and retarded TLR4-mediated immune recognition. The O-antigen-mediated evasion from early epithelial innate immune activation significantly enhances intraepithelial bacterial survival in vitro and in vivo following oral challenge. These data identify O-antigen expression as an innate immune evasion mechanism during apical intestinal epithelial invasion and illustrate the importance of early innate immune recognition for efficient host defense against invading Salmonella.
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MicroRNA-146a-mediated downregulation of IRAK1 protects mouse and human small intestine against ischemia/reperfusion injury.
EMBO Mol Med
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Intestinal ischemia/reperfusion (I/R) injury causes inflammation and tissue damage and is associated with high morbidity and mortality. Uncontrolled activation of the innate immune system through toll-like receptors (Tlr) plays a key role in I/R-mediated tissue damage but the underlying mechanisms have not been fully resolved. Here, we identify post-transcriptional upregulation of the essential Tlr signalling molecule interleukin 1 receptor-associated kinase (Irak) 1 as the causative mechanism for post-ischemic immune hyper-responsiveness of intestinal epithelial cells. Increased Irak1 protein levels enhanced epithelial ligand responsiveness, chemokine secretion, apoptosis and mucosal barrier disruption in an experimental intestinal I/R model using wild-type, Irak1(-/-) and Tlr4(-/-) mice and ischemic human intestinal tissue. Irak1 accumulation under hypoxic conditions was associated with reduced K48 ubiquitination and enhanced Senp1-mediated deSUMOylation of Irak1. Importantly, administration of microRNA (miR)-146a or induction of miR-146a by the phytochemical diindolylmethane controlled Irak1 upregulation and prevented immune hyper-responsiveness in mouse and human tissue. These findings indicate that Irak1 accumulation triggers I/R-induced epithelial immune hyper-responsiveness and suggest that the induction of miR-146a offers a promising strategy to prevent I/R tissue injury.
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Innate immune signalling at the intestinal epithelium in homeostasis and disease.
EMBO Rep.
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The intestinal epithelium--which constitutes the interface between the enteric microbiota and host tissues--actively contributes to the maintenance of mucosal homeostasis and defends against pathogenic microbes. The recognition of conserved microbial products by cytosolic or transmembrane pattern recognition receptors in epithelial cells initiates signal transduction and influences effector cell function. However, the signalling pathways, effector molecules and regulatory mechanisms involved are not yet fully understood, and the functional outcome is poorly defined. This review analyses the complex and dynamic role of intestinal epithelial innate immune recognition and signalling, on the basis of results in intestinal epithelial cell-specific transgene or gene-deficient animals. This approach identifies specific epithelial cell functions within the diverse cellular composition of the mucosal tissue, in the presence of the complex and dynamic gut microbiota. These insights have thus provided a more comprehensive understanding of the role of the intestinal epithelium in innate immunity during homeostasis and disease.
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Bacterial cell wall compounds as promising targets of antimicrobial agents II. Immunological and clinical aspects.
Curr Drug Targets
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The bacterial cell wall represents the primary target for antimicrobial agents. Microbial destruction is accompanied by the release of potent immunostimulatory membrane constituents. Both Gram-positive and Gram-negative bacteria release a variety of lipoproteins and peptidoglycan fragments. Gram-positive bacteria additionally provide lipoteichoic acids, whereas Gram-negative bacteria also release lipopolysaccharide (LPS, endotoxin), essential component of the outer leaflet of the bacterial cell wall and one of the most potent immunostimulatory molecules known. Immune activation therefore can be considered as an adverse effect of antimicrobial destruction and killing during anti-infective treatment. In contrast to antibiotics, the use of cationic amphiphilic antimicrobial peptides allows both effective bacterial killing and inhibition of the immunostimulatory effect of the released bacterial membrane constituents. The administration of antimicrobial peptides alone or in combination with antibiotic agents thus represents a novel strategy in the antiinfective treatment with potentially important beneficial aspects. Here, data are presented which describe immunological and clinical aspects of the use of antimicrobial peptides (AMPs) as therapeutic agents to treat bacterial infection and neutralize the immunostimulatory activity of released cell wall constituents.
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Bacterial cell wall compounds as promising targets of antimicrobial agents I. Antimicrobial peptides and lipopolyamines.
Curr Drug Targets
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The first barrier that an antimicrobial agent must overcome when interacting with its target is the microbial cell wall. In the case of Gram-negative bacteria, additional to the cytoplasmic membrane and the peptidoglycan layer, an outer membrane (OM) is the outermost barrier. The OM has an asymmetric distribution of the lipids with phospholipids and lipopolysaccharide (LPS) located in the inner and outer leaflets, respectively. In contrast, Gram-positive bacteria lack OM and possess a much thicker peptidoglycan layer compared to their Gram-negative counterparts. An additional class of amphiphiles exists in Gram-positives, the lipoteichoic acids (LTA), which may represent important structural components. These long molecules cross-bridge the entire cell envelope with their lipid component inserting into the outer leaflet of the cytoplasmic membrane and the teichoic acid portion penetrating into the peptidoglycan layer. Furthermore, both classes of bacteria have other important amphiphiles, such as lipoproteins, whose importance has become evident only recently. It is not known yet whether any of these amphiphilic components are able to stimulate the immune system under physiological conditions as constituents of intact bacteria. However, all of them have a very high pro-inflammatory activity when released from the cell. Such a release may take place through the interaction with the immune system, or with antibiotics (particularly with those targeting cell wall components), or simply by the bacterial division. Therefore, a given antimicrobial agent must ideally have a double character, namely, it must overcome the bacterial cell wall barrier, without inducing the liberation of the pro-inflammatory amphiphiles. Here, new data are presented which describe the development and use of membrane-active antimicrobial agents, in particular antimicrobial peptides (AMPs) and lipopolyamines. In this way, essential progress was achieved, in particular with respect to the inhibition of deleterious consequences of bacterial infections such as severe sepsis and septic shock.
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Age-dependent TLR3 expression of the intestinal epithelium contributes to rotavirus susceptibility.
PLoS Pathog.
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Rotavirus is a major cause of diarrhea worldwide and exhibits a pronounced small intestinal epithelial cell (IEC) tropism. Both human infants and neonatal mice are highly susceptible, whereas adult individuals remain asymptomatic and shed only low numbers of viral particles. Here we investigated age-dependent mechanisms of the intestinal epithelial innate immune response to rotavirus infection in an oral mouse infection model. Expression of the innate immune receptor for viral dsRNA, Toll-like receptor (Tlr) 3 was low in the epithelium of suckling mice but strongly increased during the postnatal period inversely correlating with rotavirus susceptibility, viral shedding and histological damage. Adult mice deficient in Tlr3 (Tlr3(-/-)) or the adaptor molecule Trif (Trif(Lps2/Lps2)) exerted significantly higher viral shedding and decreased epithelial expression of proinflammatory and antiviral genes as compared to wild-type animals. In contrast, neonatal mice deficient in Tlr3 or Trif did not display impaired cell stimulation or enhanced rotavirus susceptibility. Using chimeric mice, a major contribution of the non-hematopoietic cell compartment in the Trif-mediated antiviral host response was detected in adult animals. Finally, a significant age-dependent increase of TLR3 expression was also detected in human small intestinal biopsies. Thus, upregulation of epithelial TLR3 expression during infancy might contribute to the age-dependent susceptibility to rotavirus infection.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.