Gap junction hemichannels contribute to the generation of diarrhoea during infectious enteric disease

ObjectiveThe attaching and effacing (A/E) pathogens enterohaemorrhagic Escherichia coli, enteropathogenic E coli and Citrobacter rodentium colonise intestinal tracts, attach to enterocytes, collapse infected cell microvilli and alter numerous host cell processes during infection. Enterocyte alterations result in numerous small molecules being released from host cells that likely contribute to diarrhoeal phenotypes observed during these infections. One possible route for small molecules to be released from intestinal cells may be through functional gap junction hemichannels. Here we examine the involvement of these hemichannels during the diarrhoeal disease caused by A/E pathogens in vivo.DesignMice were infected with the diarrhoea-causing murine A/E pathogen C rodentium for 7 days. Connexin43 (Cx43) protein levels and immunolocalisation in the colon were initially used to determine alterations during A/E bacterial infections in vivo. Connexin mimetic peptides and connexin permeable tracer molecules were used to gage the presence and function of unpaired connexin hemichannels. The role of Cx43 in diarrhoea generation was assessed by comparing infections of wild-type mice to Cx43 mutant mice and determining the water abundance in the colonic luminal material.ResultsWe demonstrate that Cx43 protein levels are increased in colonocytes during in vivo A/E bacterial infections, resulting in functionally open connexon hemichannels in apical membranes of infected cells. Moreover, infected Cx43 +/− mice do not suffer from diarrhoeal disease.ConclusionsThis study provides the first evidence that functional connexon hemichannels can occur in the intestine and are a novel molecular mechanism of water release during infectious diarrhoea.

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Mechanism of antrocytic connexin-43 autophagy degradation in oxygen-glucose deprivation and its effect on neuroinflammation

10.21203/rs.2.18795/v1 ◽

2019 ◽

Author(s):

Xinyu Wang ◽

Liangshu Feng ◽

Meiying Xin ◽

Yulei Hao ◽

Xu Wang ◽

...

Keyword(s):

Cerebral Ischemia ◽

Connexin 43 ◽

Cell Communication ◽

Glucose Deprivation ◽

In Vivo Model ◽

Oxygen Glucose Deprivation ◽

Cx43 Protein ◽

Protein Levels

Abstract Background : Connexin 43 (Cx43) are the most widely distributed gap junction proteins in the nervous system. Cx43 enables cell-to-cell communication and plays an important role in ion transport, substrate exchange and delivery of information , which have been implicated in cerebral ischemia injury. Our previous work revealed the relationships between Cx43 and glia-mediated neuroinflammation through the release of ATP in oxygen-glucose deprivation (OGD), which means degradation of Cx43 may improve neuroinflammatory damage during OGD injury . However, the roles of Cx43 degradation and neuroinflammation caused by OGD remain unclear. Methods: We used primary cultured astrocytes treated with OGD as an in vitro model of cerebral ischemia injury and we used middle cerebral artery occlusion (MCAO) model as an in vivo model of cerebral ischemia. HeLa cells were used in overexpression experiments. Cx43 protein levels were determined by western blotting. The interaction between Cx43 and related autophagy receptors was determined by co-immunoprecipitation and immunofluorescence. The gene knockdown (KD) of ATG5, OPTN, NDP52, PINK1 and Cx43 was applied by siRNA transfection. Related cytokines were detected by cytometric bead assay. Results: We found that Cx43 protein levels increased after ischemia in gene KD of ATG5, OPTN, NDP52 and PINK1 primary astrocytes. The interaction of Cx43 with OPTN, NDP52 and PINK1 was increased after cerebral ischemia injury in vitro and vivo. While the interaction was weakened after point mutation of Cx43 at Ser368, Tyr265 and Tyr247. Meanwhile, IL-10 upregulated during OGD after KD of ATG5, OPTN, NDP52 and PINK1 in astrocytes , while TNF downregulated during OGD after KD of ATG5, OPTN, NDP52 and PINK1 in astrocytes. Conclusions: Our results suggest that degradation of Cx43 is caused by selective autophagy during ischemia injury and the autophagy degradation of Cx43 plays important roles in neuroinflammation mediated by OGD injury. Treatment targeting Cx43 degradation pathway can improve neuroinflammation responses induced by OGD injury , which provide novel therapeutic strategies and crosstalk between autophagy and neuroinflammation.

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Pharmacological iron-chelation as an assisted nutritional immunity strategy against Piscirickettsia salmonis infection

Veterinary Research ◽

10.1186/s13567-020-00845-2 ◽

2020 ◽

Vol 51 (1) ◽

Author(s):

Mario Caruffo ◽

Dinka Mandakovic ◽

Madelaine Mejías ◽

Ignacio Chávez-Báez ◽

Pablo Salgado ◽

...

Keyword(s):

Bacterial Infections ◽

Iron Chelation ◽

Iron Chelator ◽

Host Cells ◽

Bacterial Disease ◽

Protective Capacity ◽

Intracellular Iron ◽

Piscirickettsia Salmonis

Abstract Salmonid Rickettsial Septicaemia (SRS), caused by Piscirickettsia salmonis, is a severe bacterial disease in the Chilean salmon farming industry. Vaccines and antibiotics are the current strategies to fight SRS; however, the high frequency of new epizootic events confirms the need to develop new strategies to combat this disease. An innovative opportunity is perturbing the host pathways used by the microorganisms to replicate inside host cells through host-directed antimicrobial drugs (HDAD). Iron is a critical nutrient for P. salmonis infection; hence, the use of iron-chelators becomes an excellent alternative to be used as HDAD. The aim of this work was to use the iron chelator Deferiprone (DFP) as HDAD to treat SRS. Here, we describe the protective effect of the iron chelator DFP over P. salmonis infections at non-antibiotic concentrations, in bacterial challenges both in vitro and in vivo. At the cellular level, our results indicate that DFP reduced the intracellular iron content by 33.1% and P. salmonis relative load during bacterial infections by 78%. These findings were recapitulated in fish, where DFP reduced the mortality of rainbow trout challenged with P. salmonis in 34.9% compared to the non-treated group. This is the first report of the protective capacity of an iron chelator against infection in fish, becoming a potential effective host-directed therapy for SRS and other animals against ferrophilic pathogens.

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Preconditioning in the absence or presence of sustained ischemia modulates myocardial Cx43 protein levels and gap junction distribution

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y01-004 ◽

2001 ◽

Vol 79 (5) ◽

pp. 371-378 ◽

Cited By ~ 27

Author(s):

Pascal Daleau ◽

Sophie Boudriau ◽

Monia Michaud ◽

Christine Jolicoeur ◽

John G Kingma Jr

Keyword(s):

Spatial Distribution ◽

Gap Junctions ◽

Gap Junction ◽

Ischemic Preconditioning ◽

Coronary Occlusion ◽

Progressive Increase ◽

Cx43 Protein ◽

Protein Levels ◽

Ischemic Region ◽

Initial Decrease

In the heart, brief repeated episodes of ischemia prior to a sustained occlusion (ischemic preconditioning; PC) significantly delay the onset of necrosis and arrhythmogenesis. Ischemia has been reported to influence gap junction organization and connexin43 (Cx43) content, but whether PC affects these structures is not known. We investigated the effect of PC (2 cycles of 5-min ischemia plus 10-min reperfusion) followed by prolonged reperfusion without concomitant regional coronary occlusion on the myocardial Cx43 content and its spatial distribution in rabbit hearts. We also compared the effect of sustained ischemia with or without PC on Cx43 spatial distribution. In experiments with PC only, there was an initial decrease in Cx43 levels within the ischemic zone followed by a progressive increase after 48 h reperfusion. Endtoend immunolabeling of Cx43 was augmented in the ischemic region between 24 and 48 h reperfusion; labeling was not uniquely confined to myocyte abutments, but was also dispersed along the sarcolemma. Cx43 immunolabelling was more intense and diffuse in hearts subjected to PC before sustained coronary occlusion (compared to non-PC). These data indicate that gap junctions are significantly altered during brief episodes of ischemia. Reorganization of the gap junction complex could contribute to PC-mediated reductions in cardiac arrhythmias.Key words: ischemic preconditioning, connexin43, gap junction, reperfusion, heart.

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Amiodarone and Bepridil Inhibit Anthrax Toxin Entry into Host Cells

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01184-06 ◽

2007 ◽

Vol 51 (7) ◽

pp. 2403-2411 ◽

Cited By ~ 21

Author(s):

Ana M. Sanchez ◽

Diane Thomas ◽

Eugene J. Gillespie ◽

Robert Damoiseaux ◽

Joseph Rogers ◽

...

Keyword(s):

Small Molecules ◽

Lethal Toxin ◽

Host Cells ◽

Anthrax Lethal Toxin ◽

Raw 264.7 Macrophages ◽

Endosomal Acidification ◽

A Cell

ABSTRACT Anthrax lethal toxin is one of the fundamental components believed to be responsible for the virulence of Bacillus anthracis. In order to find novel compounds with anti-lethal toxin properties, we used a cell-based assay to screen a collection of approximately 500 small molecules. Nineteen compounds that blocked lethal toxin-mediated killing of RAW 264.7 macrophages were identified, and we report here on the characterization of the two most potent antitoxic compounds, amiodarone and bepridil. These drugs are used to treat cardiac arrhythmia or angina in humans at doses similar to those that provide protection against lethal toxin in vitro. Our results support a model whereby the antitoxic properties of both drugs result from their ability to block endosomal acidification, thereby blocking toxin entry. Amiodarone was tested in vivo and found to significantly increase survival of lethal toxin-challenged Fischer rats.

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Development and Retranslational Validation of anIn VitroModel to Characterize Acute Infections in Large Human Joints

BioMed Research International ◽

10.1155/2014/848604 ◽

2014 ◽

Vol 2014 ◽

pp. 1-13 ◽

Cited By ~ 2

Author(s):

Ingo H. Pilz ◽

Alexander Mehlhorn ◽

David Dovi-Akue ◽

Elia Raoul Langenmair ◽

Norbert P. Südkamp ◽

...

Keyword(s):

Bacterial Infections ◽

Mononuclear Cells ◽

Joint Infection ◽

Joint Inflammation ◽

Synovial Fibroblasts ◽

Proteoglycan Synthesis ◽

Protein Levels ◽

Human Joints

Bacterial infections can destroy cartilage integrity, resulting in osteoarthritis. Goal was to develop anin vitromodel within vivovalidation of acute joint inflammation. Inflammation in cocultivated human synovial fibroblasts (SFB), chondrocytes (CHDR), and mononuclear cells (MNC) was successively relieved for 10 days. Articular effusions from patients with (n=7) and without (n=5) postoperative joint infection in healthy patients (ASA 1-2) were used as model validation. Inflammationin vitroresulted in an enormous increase in IL-1 and a successive reduction in SFB numbers. CHDR however, maintained metabolic activity and proteoglycan synthesis. While concentrations of bFGFin vivoandin vitrorose consistently, the mRNA increase was only moderate. Concurring with ourin vivodata, cartilage-specific IGF-1 steadily increased, while IGF-1 mRNA in the CHDR and SFB did not correlate with protein levels. Similarly, aggrecan (ACAN) protein concentrations increasedin vivoand failed to correlatein vitrowith gene expression in either the CHDR or the SFB, indicating extracellular matrix breakdown. Anabolic cartilage-specific BMP-7 with highly significant intra-articular levels was significantly elevatedin vitroon day 10 following maximum inflammation. Ourin vitromodel enables us to validate early inflammation ofin vivocell- and cytokine-specific regulatory patterns. This trial is registered with MISSinG, DRKS00003536.

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Interferons: Tug of War Between Bacteria and Their Host

Frontiers in Cellular and Infection Microbiology ◽

10.3389/fcimb.2021.624094 ◽

2021 ◽

Vol 11 ◽

Author(s):

Noémie Alphonse ◽

Ruth E. Dickenson ◽

Charlotte Odendall

Keyword(s):

Signaling Pathways ◽

Bacterial Infections ◽

Host Cells ◽

Host Responses ◽

Bacterial Invasion ◽

Type I ◽

Viral Immunity ◽

Diverse Range

Type I and III interferons (IFNs) are archetypally antiviral cytokines that are induced in response to recognition of foreign material by pattern recognition receptors (PRRs). Though their roles in anti-viral immunity are well established, recent evidence suggests that they are also crucial mediators of inflammatory processes during bacterial infections. Type I and III IFNs restrict bacterial infection in vitro and in some in vivo contexts. IFNs mainly function through the induction of hundreds of IFN-stimulated genes (ISGs). These include PRRs and regulators of antimicrobial signaling pathways. Other ISGs directly restrict bacterial invasion or multiplication within host cells. As they regulate a diverse range of anti-bacterial host responses, IFNs are an attractive virulence target for bacterial pathogens. This review will discuss the current understanding of the bacterial effectors that manipulate the different stages of the host IFN response: IFN induction, downstream signaling pathways, and target ISGs.

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Autophagy and Lc3-Associated Phagocytosis in Zebrafish Models of Bacterial Infections

Cells ◽

10.3390/cells9112372 ◽

2020 ◽

Vol 9 (11) ◽

pp. 2372

Author(s):

Salomé Muñoz-Sánchez ◽

Michiel van der Vaart ◽

Annemarie H. Meijer

Keyword(s):

Defense Mechanisms ◽

Bacterial Infections ◽

Shigella Flexneri ◽

Host Cells ◽

Bacterial Degradation ◽

Phagocytic Cells ◽

Infection Models ◽

Novel Therapeutic Strategies ◽

Ubiquitin Receptors

Modeling human infectious diseases using the early life stages of zebrafish provides unprecedented opportunities for visualizing and studying the interaction between pathogens and phagocytic cells of the innate immune system. Intracellular pathogens use phagocytes or other host cells, like gut epithelial cells, as a replication niche. The intracellular growth of these pathogens can be counteracted by host defense mechanisms that rely on the autophagy machinery. In recent years, zebrafish embryo infection models have provided in vivo evidence for the significance of the autophagic defenses and these models are now being used to explore autophagy as a therapeutic target. In line with studies in mammalian models, research in zebrafish has shown that selective autophagy mediated by ubiquitin receptors, such as p62, is important for host resistance against several bacterial pathogens, including Shigella flexneri, Mycobacterium marinum, and Staphylococcus aureus. Furthermore, an autophagy related process, Lc3-associated phagocytosis (LAP), proved host beneficial in the case of Salmonella Typhimurium infection but host detrimental in the case of S. aureus infection, where LAP delivers the pathogen to a replication niche. These studies provide valuable information for developing novel therapeutic strategies aimed at directing the autophagy machinery towards bacterial degradation.

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Polymicrobial Infection and Bacterium-Mediated Epigenetic Modification of DNA Tumor Viruses Contribute to Pathogenesis

mBio ◽

10.1128/mbio.01015-14 ◽

2014 ◽

Vol 5 (3) ◽

Cited By ~ 9

Author(s):

J. M. Doolittle ◽

J. Webster-Cyriaque

Keyword(s):

Bacterial Infections ◽

Kaposi's Sarcoma ◽

Epigenetic Modification ◽

Kaposi’S Sarcoma ◽

Host Cells ◽

Viral Reactivation ◽

Barr Virus ◽

Tumor Viruses ◽

Epstein Barr

ABSTRACTThe human body plays host to a wide variety of microbes, commensal and pathogenic. In addition to interacting with their host, different microbes, such as bacteria and viruses, interact with each other, sometimes in ways that exacerbate disease. In particular, gene expression of a number of viruses, including Kaposi’s sarcoma-associated herpesvirus (KSHV), Epstein-Barr virus (EBV), and human immunodeficiency virus (HIV), is known to be regulated by epigenetic modifications induced by bacteria. These viruses establish latent infection in their host cells and can be reactivated by bacterial products. Viral reactivation has been suggested to contribute to periodontal disease and AIDS. In addition, bacterium-virus interactions may play a role in cancers, such as Kaposi’s sarcoma, gastric cancer, and head and neck cancer. It is important to consider the effects of coexisting bacterial infections when studying viral diseasesin vivo.

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